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CONTE N TS.
PAGE
INTRODUCTION...                               I
BOOIK I.
FERMENTATION DUE TO CELLULAR ORGANISMS, OR DIRECT
FERMENTATION.
CHAPTER I.
HISTORICAL, L...   9
CHAPTER II.
ALCOHOLIC OR SPIRITUOUS FERMENTATION.  I8
CHAPTER III.
AILCOHOLIC FERMENTS....      a         34
CHAPTER IV.
ACTUAL COMPOSITION OF FERMENTS.  e... 63
CHAPTER V.
FUNCTIONS OF YEAST..,,         ~ 72
CHAPTER VI.
ACTION OF VARIOUS CHEMICAL AND PHYSICAL AGENTS
ON ALCOHOLIC FERMENTATION..         159
CHAPTER VII.
CAN NOTHING BUT ALCOHOLIC YEAST EXCITE ALCOHOLIC
FERMENTATION?..... 167




vi                 CONTENTS.
CHAPTER VIII.
PAG1E
VISCoUS OR MANNITIC FERMENTATION OF SUGAR.. 189
CHAPTER IX.
LACTIC FERMENTATION..                       193
CHAPTER X.
AMMONIACAL FERMENTATION., e    203
CHAPTER XI.
BUTYRIC FERMENTATION AND PUTREFACTION..   209
CHAPTER XII.
FERMENTATION BY OXIDATION...       228
CHAPTER XIII.
APPLICATION OF THE RESEARCHES AND IDEAS OF M.
PASTEUR.....              245
BOOK II.
ALBUMINOID SUBSTANCES-SOLUBLE OR INDIRECT FERIMENTSORIGIN OF FERMENTS.
CHAPTER I.
ALBUMINOID SUBSTANCES, OR PROTEIDS...        253
CHAPTER II.
SOLUBLE FERMENTS, AND INDIRECT FERMENTATION. 269
CHAPTER III.
ON TIlE ORIGIN OF FERMENTS.             ~ e    308




LIST OF ILLUSTRATIONS.
Fig. I. —Saccharomyces cerevisie - Sedimentary yeast,
X 400 diams.                                    45
Fig. 2.-Saccharomyces cerevisite-Sedimentary yeast, budding, X 400 diams.                              47
Fig. 3.-Saccharomyces cerevisiex-Surface yeast, budding,
X 40o diams.                                    47
Fig. 4.-Saccharomyces cerevisioe-Surface yeast, at rest,
X 400 diams.                                    49
Fig. 5.-Saccharomyces cerevisize-Sedimentary yeast, in a
growing state, X 40o diams..            49
Fig. 6.-Saccharomyces cerevisie — Formation of spores,
X 750 diams. a, b, c, d, e, successive phases of
the production of spores                        5I
Fig. 7.-Triads of spores, germinating, X 750 diams... I
Fig. 8.-Saccharomyces ellipsoideus, in process of budding,
X 6oo diams.                                    55
Fig. 9.-Saccharomyces ellipsoideus, development of Spores,
X 400 diams.                                    55
Fig. Io.-Saccharomyces ellipsoideus, group of spores in the
act of germinating, X 400 diams.....  55
Fig. II.-Saccharomyces Pastorianus, X 400 diams..       56
Fig. I2.- -Saccharomyces exiguus, X 350 diams...  56
Fig. I3.-Saccharomyces congloineratus, X 6oo diams...   6




viii              LIST  OF ILLUSTRATIONS.
PAGE
Fig. 14. —Saccharomyces  apiculatus  (Rees).   Carpozyma
apic. (Engel), apiculated ferment, X 600 diams.   57
Fig. I5.-Saccharomyces Reesii, ferment of red wine, x 350
diams....                     59
Fig. I6.-Saccharomyces mycoderma, X 350 diams.                    59
Fig. I7. —Saccharomyces mycoderma.,                        59
Fig. I. —Mucor racemosus, ferment in mass..         60
Fig. I9.-Apparatus for the measurement of oxygen dissolved in water.I.... 121
Fig. 20.-Viscous Ferments of Wine..          I90
Fig. 2 I.-Lactic Ferment..                                  I98
Fig. 22.-Mycoderma aceti..                                 239
Fig. 23.-Giret and Vinas' apparatus for warming wines            247
Figs. 24 and 25. —Organic corpuscules of dust, mixed with
amorphous particles...                    317
Fig. 26. -M. Pasteur's apparatus for the introduction of calcined air into flasks containing organic infusions  319
Fig. 27.-M. Pasteur's apparatus for studying the resistance
of germs and spores to temperatures more or less
elevated...                         323
Fig. 28.-M. Pasteur's flask to deprive the air of its germs. 323
NOTE BY TRANSLATOR.
The French words "invertir,"- inverti," "intervertir," "inversive," &e.,
p. 28, &c., may be rendered in English by the recognized terms, "inverted,"
"inversive," &c. Yet since these terms are not understood by all, and present
some difficulty, it has been thought better to adopt in the text the less technical
and more suggestive rendering " altered," "alterative," &c.; by altered sugar
being meant cane-sugar which has taken up a molecule of water and split up
into a mixture of glucose and levulose.




ON FERMENTATION.
INTRODUCTION.
FERMENTATION is only a particular instance, selected
from among the chemical phenomena of which living
organisms are the field; it, like all biological reactions,
comes before us as a manifestation of the special force
residing in these organisms, or rather in their cellular
elements.
If we leave the nature of the fermenting body, and
the products derived from it, in the background,
there is nothing to distinguish fermentation from the
otlier chemical transformations which take place in the
animal or vegetable economy.
The reason why the production of alcohol and carbon
dioxide at the expense of sugar, the conversion of
glucose into lactic and butyric acids, and other phenomena of the same order have been classed by themselves, is that the real cause of these curious transformations was long misunderstood.  It had not been
observed that they had as their origin the presence of
living organisms, or, at least, principles which are
directly derived therefi-om.




2               ON FERMENTATION.
There is, therefore, no longer any necessity, in the
present state of science, for grouping together under a
special name these various reactions; it is more convenient, on the contrary, to class them among the
general mass of chemical phenomena of the living
organism.
We must, consequently, do one of two things; either
cease to use the term fermentation, as a general expression applying to a certain order of phenomena,
or we must designate by it all those changes which,
by the special conditions under which they are produced, are evidently due to the intervention  of a
force differing from those which we handle in our laboratories.
It is true that the organisms which give rise to what
have been hitherto called fermentation are simple
elementary organisms reduced to a single cell; but a
plant or an animal of a high order is only the union,
under special laws, of different kinds of cells, each of
which acts in a certain determinable manner. When,
as M. Pasteur has remarked, we sow at the same time,
in the same saccharine medium, alcoholic, lactic, and
butyric ferment, we see three distinct reactions take
place, one of which splits up the sugar into alcohol and
carbon dioxide, the second converts it into lactic acid,
and the third into butyric acid.
The more simple an organism  is, the fewer special
kinds of cells it contains, the simpler are the chemical
reactions which take place in it, and the more easily are
they separated from each other, and isolated by experiment. On the contrary, in proportion as the histological
constitution is varied and heterogeneous, we see a




INTRODUCTION.                  3
greater number of distinct compounds, as the products
of the many chemical changes which take place in the
different tissues.
As a consequence of what we have just said, our plan
would be considerably enlarged, and the history of
fermentation would become that of the chemical phenomena of life.
We will not, however, give such a wide scope to this
work, but will confine ourselves to the examination of
the phenomena which have been hitherto designated by
the name of fermentation. Under these restrictions, the
history of fermentation may be considered as an introduction to biological chemistry.
In fact, it is easily seen, from  the preceding considerations, that the thorough study of ferments, properly
so called, or rather of elementary organisms, and of
their mode of existence, ought to precede that of the
more complete beings. We more easily understand the
properties of granite, and the influence exerted upon it
by water and atmospheric agents, when we have learned
that it is formed of crystals of quartz, felspar, and mica in
juxtaposition, and have studied the chemical characters
of each of these compounds. In the same manner, the
study of the chemical manifestations of the vital force
in cellular organisms is destined to throw a bright light
on the more complex functions of the higher plants and
animals. This has been recognized by M. Pasteur, and
by all those who have subsequently entered on the
physiological study of fermentation, and of the development of cellular organisms.
A living cell of beer-yeast possesses the property of
resolving into alcohol, glycerin, carbon. dioxide, and




4               ON FERMENTATION.
succinic acid the altered sugar which penetrates by
endosmose through its membranous envelope.
If we substitute for the cell of beer-yeast a cell of
lactic ferment, we still see the sugar disappear, but the
products into which the ponderable elements of the
glucose are resolved are different; instead of alcohol
and carbon- dioxide, we have lactic acid. The moduts
ficacienzdi of the vital force of this cell is evidently not
the same as that of the former one; but we cannot,
therefore, affirm that there are as many vital chemical
forces as there are reactions.
When a pencil of solar light passes through a prism,
the constituent parts of this pencil are isolated on
account of their unequal refrangibility.  The least
refrangible rays are revealed to us by the effects of
heat (the dilatation and change of condition of bodies);
next come the luminous rays, which excite on the retina
the impressions of colour forming the spectrum; and
then, beyond the violet, is a series of invisible rays,
which are revealed only by their decomposing action on
certain combinations (salts of silver, &c.).  But we
know now that all these calorific, luminous, and chemical rays, some of which give heat without light, and
others light without heat, or excite chemical reactions,
differ only in the rapidity of the vibratory movements of
the ether, and are essentially distinguishable from each
other only by their wave-length. It is possible. that an
analogous bond may unite the vital chemical forces of
the different elementary organisms.  Sand, sprinkled
uniformly on the surface of a vibrating plate, collects
in nodal lines of different forms, according to the sharpness of the note which we draw from this plate by




INTRODUCTION.                  5
means of a bow; in the same manner, chemical compounds may perhaps be resolved into more simple
combinations, varying in kind according to the vibratory
rhythm which starts them.
The transformation of sugar into alcohol and carbon
dioxide, and the conversion of the same body into
lactic acid are chemical phenomena which we cannot yet
reproduce by the intervention of heat alone, nor by the
additional agency of light or of electricity. The. force
capable of attacking, in a certain determinate direction,
the complex edifice which we call sugar, an edifice
composed of atoms of carbon, hydrogen, and oxygen,
grouped according to a determinate law-this force,
which is manifested only in the living cell of the ferment, is a force as material as all those which we are
accustomed to utilize.
Its principal peculiarity is, that it is only found in the
living organisms, to which it gives their peculiar character. We ought not to allow ourselves to be stopped by
this rampart, over which no one has hitherto been able
to pass; we ought not to say to the chemist, "You
shall go no farther, for beyond this is the domain of
life, where you have no control."
The history of science shows us the weakness of these
so-called impassable barriers.
Gerhardt, when he published his excellent treatise
on organic chemistry, thought himself justified in saying," It is vital force alone which acts synthetically
and reconstructs the edifice demolished by chemical
forces."
M. Berthelot, some years afterwards, in a brilliant
series of discoveries, made the first successful attempt to




6              ON FERMENTATION.
perform organic syntheses, and determined the principal
conditions under which they can be effected.
In a remarkable lecture on molecular dissymmetry
(Leqons de la  Societe Chimique de Paris, i86o),
M. Pasteur had established an important distinction
between artificial organic products and the compounds
formed under the influence of living organisms.
"The artificial products of the laboratory have coincident images (sont'a image superposable). On the contrary, most of the natural organic products-I might say
all, if I had only to allude to those which play an important part in the phenomena of vegetable and animal lifeall the products essential to life are unsymmetrical, and
unsymmetrical in such a way that their images cannot
be made to coincide with them."  And afterwards he
says, "We have not yet been able to realize the production of an unsymmetrical body, by the aid of compounds
which are not so themselves."
Nearly at the same time that these words were
uttered before the Chemical Society of Paris, two
English chemists, Perkin and Duppa, succeeded in
transforming succinic acid into tartaric  acid.  M.
Pasteur himself acknowledged that the artificial product of Perkin was a mixture of paratartaric acid and
of inactive tartaric acid. But paratartaric acid easily
splits up, as Pasteur's elegant experiments have shown,
into dextro-tartaric and  levo-tartaric  acid, and M.
Jungfleisch has shown that inactive tartaric acid, heated
with water at I75~, is partially converted into parae
tartaric acid.
The succinic acid employed by the English chemists
was formed by the oxidation of yellow amber. This




INTRODUCTION.                  /
was not a synthetical product; it might be thought
that, though it was inactive, it resulted, like racemic
acid, from the union of two active but opposed molecules. Jungfleisch has removed tlis last doubt. He
prepared  synthetic  succinic acid  by a well-known
method, by means of cyanide of ethylene and potassium.
This acid furnished paratartaric acid, like that produced
from amber.
Thus fell the barrier placed by M. Pasteur between,
natural and artificial products. This example shows us
how cautious we ought to be in making distinctions
which we seem justified in establishing between the
chemical reactions of the living organism and those of
the laboratory. Because a chemical phenomenon may
hitherto have been produced only under the influence of
life, it does not follow that it will never be effected
otherwise.
No one can any longer admit that vital force has
power over matter, to change, counterbalance, or annul
the natural play of chemical affinities. That which we
have agreed to call chemical affinity is not an absolute
force; this affinity is modified in numberless ways,
according as the circumstances by which bodies are
surrounded, vary.  Thus, the  apparent differences
between the reactions of the laboratory and those of
the organism ought to be sought for, more particularly
among the special conditions, which the latter alone has
been able hitherto to bring together.
In other words, there is really no chemical vital force.
If living cells produce reactions which seem peculiar
to themselves, it is because they realize conditions of
molecular mechanism which we have not hitherto suc



8              ON FERMENTATION.
ceeded in tracing, but which we shall, -withozut doubt, be
able to discover at some future time.  Science can
gain nothing by being limited in the possibility of the
aims which she proposes to herself, or the end which
she seeks.
If, in this work, we still employ the expression, "the
vital chemical force of an elementary organism," it will
be clearly understood that we intend these words to
signify the realization of the conditions of molecular
mechanism necessary in order to set up a certain reaction.
We will not delay any longer over these general
considerations, which are, after all, nothing but hypotheses, naturally suggesting -themselves to the mind of
him who seeks to explain the causes which produce
certain observed effects, but on which it is not necessary to dwell at the present stage of our inquiry; we
will therefore proceed at once to the examination of
facts.
The study of fermentation may be divided into two
parts, according to the nature of the ferment.  The
first will comprise the, fermentation due to the intervention of an organized ferment, having a determinate
form; the second will be reserved for fermentation
produced by soluble products, elaborated by living
organisms.




BOOK I.
FERMENTATION DUE TO CELLULAR ORGANISMS, OR
DIRECT FERMENTATION.
CHAPTER I.
HISTORICAL.
THE word fermentation is derived from fervere, to boil;
it evidently owes its origin to the reaction presented by
saccharine liquids, when they are left to themselves or
placed in contact with ferments. We observe, in fact,
in this case, a more or less abundant disengagement of
gas, which causes the liquid to effervesce or boil.  The
sugar disappears, and the product becomes spirituous.
The expression, fermentation, was subsequently applied
to other phenomena, in which an organic body, when
dissolved, is modified, changed, and transformed, under
the influence of a cause which remained for a long time
unknown and badly defined. Thus the acidification of
wine was called fermentation, although in this case
there was no effervescence. The analogy of the determining cause was considered, rather than the appearance of the phenomenon.
Alcoholic fermentation was the first known, and was




O             ON FERMEN TATION.
also more studied than the other reactions of this class.
Osiris among the Egyptians, Bacchus among the
Greeks, Noah, according to the Israelitish tradition,
taught men the art of cultivating the vine, and making
wine.  Moses, in his writings, draws a distinction between unleavened and leavened bread, and relates that
the Israelites were in such haste, during their flight from
Egypt, that they had no time to put leaven into their
dough. The ancients used as leaven for their bread
either dough that had been kept till it was sour, or beeryeast.
"Galliae et Hispanize frumento in potum resoluto,
spuma ita concreta pro fermento utuntur, qua de causa
levior illis, quam  ceteris, panis est," says Pliny, who
also adds, that in the fermentation of bread acidity is
the most active principle. From the earliest times certain fermented liquids were known, both in Egypt and
Germany, prepared from natural saccharine juices which
had been allowed to ferment; such as beer, hydromel,
palm-wine, and cider.
We find, in short, from all ancient documents, that
alcoholic fermentation was empirically known in its
principal effects, and utilized at a period far earlier than
that which has left written traces of its history.
Among the writings of the alchemists from the
thirteenth to the fifteenth century, we very frequently
find the expressions "fermentation  and  ferments"
(fermentatos et fermentum), without our being able to
ascertain clearly what precise ideas they attached to
them. They knew no distinction between mineral and
organic substances; the phenomena connected with
the changes in organic products were assimilated and




HISTORICAL.                  I I
confounded with the transformations of mineral compounds, and with the solution.of salts and metals. The
term "ferment" was often applied even to the philosopher's stone.
" Apud philosophos fermentum dupliciteo videtur dici;
uno modo ipse lapis philosophorum et suis elementis
compositus, et completusl in comparatione ad metalla;
alio modo illud, quod est perficiens lapidem et ipsum
complens.
"De. primo modo dicimus, quod sicut fermentum
pastae vincit pastam, et ad se convertit semper, sic et
lapis convertit ad se metalla reliqua. Et sicut una pars
fermenti pastae habet convertere partes pastae et non
converti, sic et hic lapis habet convertere plurimas
partes metallorum  ad se, et non converti."-PETRUS
BONUS of Ferrara,; 1330-1340.
We see that the writer is especially struck with this
fact, that a very small quantity- of leaven transforms
into fresh leaven an almost indefinite quantity of paste.
This property of transmitting a force to a large mass
without being itself weakened by the process, was precisely that which ought to characterize the philosopher's
stone which was so much sought after.
Basil Valentine, in his " Triumphal Car of Antimony,"
admits that yeast, employed in the preparation of beer,
communicates to the liquor an internal inflammation,
and determines thereby a purification, and a separation
of the clear parts from those which are troubled.
Alcohol, the presence of which in the fermented
liquid was known to him, was considered by him to
exist previously in the decoction of germinated barley;
but that it did not become active and susceptible of




12             ON FERMENTATION.
being separated by distillation until it had been cleared
from the impurities which accompany it, and mask its
special properties.
Libavius (Alchymia, I595) believed that, "Fermentatio est rei in substantia, per admistionem fermente
quod virtute per spiritum distribute totam penetrat
massam et in suam naturam' immutat, exaltatis." The'ferment must be of a similar nature to the matter which
enters into fermentation, and the latter must be either
liquid, or in a state of minute division; the principal
agent resides in the heat of the ferment.
Like the chemists of a later age, Libavius compares
fermentation to putrefaction, and considers them as different effects of the same cause.' He protests, on the
contrary, against the confused ideas which had been
entertained  concerning digestion  and  fermentation.
Digestion is, according to him, "motus ad mistionem,
non ad perfectionem," as fermentation is.
The iatro-chemical school attributed to fermentation
a preponderating power, and even confounded under this
term a great number of chemical reactions.
Thus Van Helmont expresses himself as follows in
his "Ortus Medicine " (648): " Docebo omnem transmutationem  formalem  praesupponere fermentum  corruptivum."
The formation of intestinal gasses, the production of
blood and animal fluids, spontaneous generation, the
effervescence of chalk under the influence of acids,
are phenomena with which fermentation has to do.
We may, however, say in passing that Van Helmont
had the merit of clearly distinguishing the production
of a special gas (gas vinorum) during alcoholic fermen



HISTORICAL.                   13
tation. He says that this gas vinorum is different from
spirit of wine, as he was able to prove by experiments.
It is impossible to discover from his writings whether
or no he recognized the identity of the gas vinorum and
the gas carbonum produced during the combustion of
charcoal.
In I664, Wren pointed out that the gas produced by
alcoholic fermentation can be absorbed by water, like
that which is disengaged by the action of an acid on
salt of tartar.
Silvius de la BoG (1659) no longer regarded the effervescence of the alkaline carbonates under the influence
of acids as a phenomenon of the same class as fermentation.
He supposed that in the former case there was combination, in the latter decomposition.
Lemery (Cours de Chimie, I675) is not so explicit
when he says: "The fermentation which occurs in paste,
wort, and all other similar things, is different from that
of which we have just spoken (effervescence), since it is
slower; it is excited by the natural acid salt of these
substances, which, becoming disengaged, and having its
energy increased by its motion, rarefies and raises the
gross and oily part which opposes its passage, and thus
we see the matter rise.
"The reason why the acid does not cause sulphurous
substances to ferment with as much noise and readiness
as alkalis, is that oils are composed of pliant parts which
yield to the points of the acid, as a piece of wool or cotton
would yield to needles pressed into it. Thus it seems
to me that we must admit of two sorts of fermentations;
one of acids with alkalis, which would be called effer



14               ON FERMENTATION.
vescence; and the other would be, when the acid rarefies
by degrees a solid matter like paste, or clear and
sulphurous like wort, cider, or other juices of plants;
we should call the latter sort fermentation."'
Lemery says again, when speaking of alcoholic fermentation; "In order to understand this effect, we must
know that wort contains much essential salt; this salt,
being volatile, makes an effort, during fermentation, to
detach itself from the oily particles by which it is, as it
were, bound; it penetrates them, divides and separates
them, until by its subtile and piercing points, it has rarefied them  into spirit; this effort causes the ebullition
which takes place in wine, and, at the same time, its
purification; for it separates and removes the grosser
parts in the form  of froth, a portion of which attaches
itself to the sides of the vessel and grows hard; the
other falls to the bottom, and is called tartar and lees.
The inflammable spirit of wine is therefore nothing but
an oil exalted, that is purified, by salt."
We find in the researches and writings of Becker
(1682), a very marked progress in the study of the products of fermentation. He was the first to bring forward the important fact that saccharine liquids alone
are capable of entering into spirituous fermentation.
He considers that alcohol does not pre-exist in the
wort, but is formed during the process of fermentation;
the intervention of air is necessary to set this action
going, which he considers analogous to combustion.
Becker brings together, under the name of fermentation,
the production of gas in the stomach of sick animals
(insumefactio), spirituous fermentation (proprie fermentatio), and acetification (acetificatio seu acescentia).




HISTORICAL.                  15
We owe: to Willis (I659), and to Stahl, the celebrated
originator of the theory of phlogiston (I697), the first
philosophical conception of the peculiar nature of fermentation, or rather of fer;enltations.  According- to
their views a ferment is a body endued with a motion
peculiar to itself, and it transmits this motion to the
fermentable matter.  Thus Willis says in his dissertation "De Fermentatione ": —
"Fermentatio est motus intestinus cujusvis corporis,
cum tendentia ad perfectionem ejusdem corporis vel
propter mutationem in aliud. Plures sunt modi quibus
fermentatio  promooctur.  Primus et principius erit
fermenti cujusdam corpori fermentando adjectio; cujus
particulae cum prius sint in vigore et motu positae, alias
in massa fermentanda otiosas et torpidas exsuscitant, et
in motum vindicant."
Stahl considered alcoholic fermentation as a phenomenon of the same class as putrefaction, and as only
a particular case of it. As there was at that time no
definite idea of the elementary composition of fermentable substances, and of the products of their fermentation, there evidently could not be established any correct
relation between these bodies, and any hypothesis could
be safely brought forward. Thus Stahl considers that
fermentable matter (sugar, flour, milk) is composed of
particles formed by the unstable union of salt, oil, and
earth; under the influence of the internal motion excited
by the ferment, the heterogeneous particles are separated from each other, and then recombined so as to
form more stable compounds including the same principles, but in other proportions.
From Stahl to Lavoisier we find no names of great




i6             ON FERMENTATION.
note, and no interesting discoveries with respect to
fermentation.
When chemistry underwent its great transformation
at the end of the last century, under the powerful influence of the genius of Lavoisier, fermentation necessarily attracted anew the attention of experimentalists.
Lavoisier himself studied it (Elemens de Chimie, vol. i.
p. I39, second edition), and as was the case with all
subjects which he handled, he threw a ray of light
upon the darkness. Proceeding in his usual manner,
balance in hand, by weight and measure, and applying
to the solution of the problem the new methods of
organic analysis which he had' invented, he endeavoured
to ascertain the bond or relation which exists between
the fermented matter, the sugar, and the products of
fermentation, alcohol and carbon dioxide.
From this moment we quit the domain of the history
of the science, and enter into that of the real and wellobserved facts which will be' treated of in the following
chapters.
We may say, in recapitulation, that, before the labours
of Lavoisier and his followers, the fermentable products
and the principal terms of their transformations (carbon
dioxide gas, alcohol, acetic acid, &c.) were known qzaclitatively. The distinction between the acid, or acetic
fermentation, and the alcoholic fermentation was known;
there was an idea of the analogy which exists between
putrefaction and alcoholic fermentation; and an explanation of the manner in which a ferment acts had
been sought.
The latter was known only as a kind of foam, deposit,
or paste, in which resided an occult and special force,




HISTORICAL.                 7
capable of determining chemical phenomena. We may
add that these phenomena were considered as distinct,
both in their action and exciting cause, from the
ordinary reactions of chemistry. This was, as one may
see, but a slight result of the many volumes that: had
been written on this subject.
Spirituous or alcoholic fermentation being, in every
respect, the part of this subject which has been the most
thoroughly studied, we will commence our monograph
by its examination.
2




I8             ON FERMENTATION.
CHAPTER II.
ALCOHOLIC OR SPIRITUOUS FERMENTATION.
PASTEUR, in his excellent memoir (Ann. de Chimie et
Physique, 3rd series, vol. lviii. p. 323), calls by the name
of alchoholic fermnetation that which sugar undergoes
under the influence of the ferment which bears the
name of yeast or barm.
We can only adopt this definition as applying, without any possibility of uncertainty, to a phenomenon
very limited in its cause and its effects; but we shall
have to inquire, in a later portion of this work, whether
alcohol cannot be produced at the expense of sugar
under other influences than those of the product known
as beer-yeast.
As we have before said, the splitting up of a molecule
of sugar into many more simple products, among which
we find alcohol and carbon dioxide, is the consequence
of a special mechanical action, exercised on the ultimate
particles of the compound matter. Whatever may be
the source, whether living organism or dead matter,
which realizes the conditions necessary for this rupture
of equilibrium, the phenomenon will be essentially the
same. In a general and philosophical point of view,
there is no more reason why we should separate alcoholic fermentation excited by yeast from that which
is due to any other agent, than  why we should




ALCOHOLIC OR SPIRITUOUS FERMENTATION.  19
distinguish cane sugar from that produced from beetroot.
While we restrict, with Pasteur, the expression " alcoholic fermentation," and do not include in it all the
phenomena of decomposition, in which alcohol is produced, we have to consider the body which ferments,
the sugar, or rather the sugars, the products of fermentation, among which alcohol takes its place in the first
rank, and then the determining cause of the fermentation of sugar, beer-yeast.
PRODUCTS OF THE REACTION.
Let us first consider alcoholic fermentation as an
ordinary chemical reaction; in other words, let us study
it by means of the body which is decomposed, and the
products which are derived from it; we will then
consider the cause of the decomposition, and the properties of this ferment, as well as those of analogous
products.
We said before that Becker was the first to recognize
the necessity of the presence of sugar in wines which
undergo spirituous fermentation, but that to Lavoisier
belongs the honour of having studied and demonstrated
the relations of their composition which connect sugar
with its derivatives.
Setting out with this principle, that nothing is created
either in the operations of art, or in those of nature;
that in every operation there is an equal quantity of
matter both before and after the operation; that the




20             ON FERMENTATION.
quality and quantity of the elements are the same, and
that there are only changes and modifications, this
illustrious chemist established by analysis the centesimal proportions of carbon, hydrogen;, and oxygen
contained in sugar, operating in the same manner on
the alcohol, the carbon dioxide, and acetic acid recognized by him as the products of the decomposition of
sugar; then, estimating by analysis the respective
quantities of these three bodies which are formed at
the expense of a known weight of sugar, he ascertained
the result of the reaction, and arrived at the. following
conclusions:
"The effects of vinous fermentation are reduced to
separating into two portions the sugar, which is an
oxide, and oxidizing one at the expense of the other,
so as to form  from  it carbon dioxide, in reducing
the other inri favour of the former, to form from it a
combustible substance, alcohol; so that if it were possible-to recombine these two substances, alcohol and
carbon dioxide, we should reform the sugar."  He had
really made a great advance on the conceptions of
Stahl, founded on a mixture of salt, oil, and earth.
The researches of Lavoisier may be summed up by
the following equation;95'9 parts of crystallized cane sugar contain 26'8 of
carbon, 7'7 of hydrogen, and 6I'4 of oxygen.
These are decomposed, and form 57'7 parts of
alcohol, containing I6'7 carbon, 9'6 hydrogen, and
3I'4 oxygen + 35 3 parts of carbon dioxide, containing 9'9 carbon, and 25'4 oxygen+2'5 parts of'acetic
acid, containing o'6 carbon, 0'2 hydrogen, and 1'7
oxygen.




ALCOHOLIC OR SPIRITUOUS FERMENTATION.  2I
Thus we find:Carbon of the sugar.          26-8
Hydrogen,,.  7 7
Oxygen,,,,    o,.   6I'4
Sum of carbon of the three products 27'2,,, hydrogen,,       9'    8,,,, oxygen    9     9,    58'5
Taking into consideration the imperfections of his
method of analysis, Lavoisier thought the agreement
between the two sides of this equation sufficient to
confirm the general principle announced above.
If we compare with these numbers those furnished
by the wonderfully accurate methods employed by
modern chemists, we shall see that, in reality, 95'9 parts
of cane sugar contain:44'4 carbon, 6'I hydrogen, and 49'4 oxygen;
and give,
5 I'6 of alcohol, containing26'9 carbon, 6'7 hydrogen and I8'o oxygen +
49'4 parts of carbon dioxide, containingI3'5 carbon, and 36'9 oxygen.
It was therefore only by compensation of considerable errors that Lavoisier was led to an approximate
solution.
Towards I815, the analyses so carefully made by
Gay-Lussac and Thenard, and by De Saussure, settled
in a determinate manner the composition of sugar and
of alcohol. These results, far from  invalidating the
conclusions of Lavoisier, gave them solid support.
Thus Gay-Lussac (Ann. de Chimie, vol. 95, p. 318)
wrote: "If it be supposed, now, that the products




22               ON FERMENTATION.
furnished by the ferment can be neglected, as far as
relates to the alcohol and carbonic acid which are the
only sensible results of fermentation, it will be found
that, given Ioo parts of sugar, 5I'34 of them will be
converted during fermentation into alcohol, and 48'66
into carbonic acid."
These  results, expressed in a chemical equation,*
give to cane sugar the formula C12 H24 012,-t and
we shall have C12 H24 012 =  4 C2H6 00   +  4 CO.++
The analyses of cane sugar made by Gay-Lussac and
Thenard themselves agree, as well as those since made
by a great number of chemists, with the formula
C12 H22 o.1~
In order to arrive at the error which we have just
pointed out, and which Messrs. Dumas and Boullay
showed in I828, Gay-Lussac supposed that his analyses
of cane sugar were imperfect, and he modified them
recklessly, in the proportion of 2 or 3 per cent., in order
to establish an agreement between the two sides of his
equation.
"The  theory of fermentation arrived at by GayLussac is still imperfect," said Messrs. Dumas and
Boullay, "but it is no longer so when we substitute
ether for alcohol in the theoretical composition of sugar.
The -most complete agreement is then established
between  theory  and  experiment."   The conclusion
which these two chemists deduced from this obser* These formulae are given by M. Schiitzenberger as original formulae. It may
be worth while, from an historical point of view, to preserve them as written by
their enunciators, in the " old notation," thus:t+ C12 H12 012.    + C12 H12 012 =    C4 H6 02 + 4 CO0,
~ C12 H11 O1.




ALCOHOLIC OR SPIRITUOUS FERMENTATION.  23
vation, was that cane sugar cannot ferment without
assimilating the elements of a molecule of water. In
other words, Gay-Lussac's equation, as a numerical
expression, is correct, but that it would be better to
write the first member of it under the formC12 H22 011 + H2 0 -4 C2 H0 0 + 4 CO2
cane sugar   water   alcohol  carbon dioxide.
A little later (I832), Dubrunfant observed that before
fermentation commenced, the cane sugar is transformed
into uncrystallizable sugar.
M. Berthelot proved afterwards that the taking up of
water by the cane sugar which precedes alcoholic fermentation is due to the presence of a soluble ferment
in the yeast; we shall return again to this important
point. Finally, in I833, Biot discovered the change of
sugar under the influence of acids.
Gay-Lussac's equation,, modified by Dumas and
Boullay, was generally admitted for more than twenty
years, as the mathematical expression of the decomposition of sugar by yeast.
Meanwhile, in I856, Dubrunfant, by making a quantitative analysis of the carbon dioxide disengaged by
fermentation, observed that it was not possible to make
experimentally the equation of fermentable sugars with
alcohol and carbon dioxide only. (Comp. Rend. de
l'Acad. des Sciences, vol. 42, p. 945.)
The latest important work on the qualitative and
quantitative analyses of the products of the alcoholic
fermentation of sugars is due to M. Pasteur. (Ann.
* C12 H, 01 + HO = 2 C4 H6 O02 + 4 CO2.
cant sugar  water   alcohol  carbon dioxide.




24              ON FERMENTATION.
Chimie et Physique, 3rd series, vol. 58, p. 330.)  By a
series of very interesting researches, and by irrefutable
experiments, this illustrious chemist proves: Ist. That
in every alcoholic fermentation, besides the principal
products, alcohol and carbon dioxide, glycerin and
succinic acid are formed; 2nd. That the glycerin and
succinic acid are produced at the expense of the elements of the sugar, and that the ferment takes no part
in it; 3rd. That, besides this, the sugar yields a certain
portion of its substance to the new ferment which is
developed; we shall return to the last point when we
more especially consider the ferment; 4th. That the
lactic acid, the production of which, in variable quantities, has been observed in alcoholic fermentation, is
the result of a special fermentation, differing from
alcoholic fermentation, and proceeding simultaneously
with it.
Let us say, in conclusion, in order to give to every
one his due, that the presence of succinic acid in fermented liquids had been observed, before M. Pasteur,
by Dr. Schmidt of Dorpat (Handwirterbuch der Chimie,
Von Liebig, Poggendorff, Ist edit., vol. 3, p. 224, I848),
and also by Schunck in the fermentation of sugar
by means of erythrozyme, the ferment derived from
madder.
These facts had:passed unperceived, and had been
forgotten, at the time when Pasteur returned to the
study of this subject.
Without entering into the details of the experiments
on which Pasteur's conclusions rest, and which will be
found in his memoir (loco citato), we will simply give
the results of his quantitative researches.




ALCOHOLIC OR SPIRITUOUS FERMENTATION.  25
Ioo parts of cane sugar C12 H22 0u, * corresponding
with Io5'26 grape sugar 2 C6 H112 0,'t give nearly,Alcohol   a. 5I' I
Carbon dioxide i 48'89 according to Gay-Lussac's equation
0'53 excess over,,,
Succinic acid. 0'67
Glycerin.. 3'I6
Matter  unitedl  I00
with ferments
I00'00
Thus, out of Ioo parts of cane sugar, about 95 parts
are decomposed, according to Gay-Lussac's equation;
4 parts disappear and form succinic acid, glycerin, and
carbon dioxide, and  I part is added  to the newlyformed ferment.
Pasteur endeavours to represent by an equation the
decomposition of the 4 parts of sugar, which yield
succinic acid and glycerin.  This expression is very
complex:49 (C,2 H2,2 01 + H2 O)
or 49 (C12 H24,,2) + 6o H2 0 - 24 (C4 H6 04) + I44 (C3 I118 03) +
succinic acid    glycerin
60 CO.2+
carbon dioxide
This equation can only be considered, as Pasteur
himself says, as a very approximate expression of the
numerical results of the analysis, and not as a mathematical expression of the reaction.
C12 H11 011.           t C12 H,2 012.,3 49 (C12 H,, 01, + HO), or 49 (C12 H,2 O12) + 60 HO -
12 (C8 H6 08) + 72 (C6 H8 06) + 6o CO2.




26              ON FERMENTATION.
M. Monoyer (These de la Faculte de Medicine de
Strasbourg) proposes a much more simple equation to
represent Pasteur's analysis:4 (C12 H22 011 + H2 0) *
or 4 (C12 H24 012) + 6 H2 0 = 2 (C4 H6 04) + 12 (C3 H8 03) +
4 C02 + 02.
He supposes, at the same time, that the excess of
oxygen serves for the respiration of the globules of the
ferment, a very plausible interpretation, as we shall
presently see.
In order to understand the chemical possibility of the
production of glycerin and succinic acid at the expense
of sugar, it is sufficient to remark that by adding together the formulae of glycerin and succinic acid, atom
by atom, we arrive at a sum in which hydrogen and
oxygen are in the proportions to form water:C4 H6 04 + C3 H8 03 = C7 H14 01.
On the other side we have,
C7 H14 07 + H2 0 -2 C3 H8 03 + C 02.
These two equations explain naturally enough the
formation of glycerin and succinic acid, at the expense
of glucose.
Even according to the researches of Pasteur, the proportions of glycerin and succinic acid, in relation to the
alcohol, furnished by the same weight of sugar, are not
absolutely constant. More glycerin and succinic acid,
and less alcohol are formed, according as the fermenta* 4 (C12 H  ~11 -- HO), or 4 (C12 H12 012) + 6 HO =
Cs H6 O8 + 6 C6 H8 O06 + 2 C2 04 + O2.




ALCOHOLIC OR SPIRITUOUS FERMENTATION.  27
tion is slower, or is made with more exhausted and less
pure yeast, supplied with but few alimentary principles,
and those unsuited for the multiplication of its globules.
Fermentation effected by sowing the ferment, in the
presence of more than a sufficient quantity of albuminoid and mineral matter suited to the nature of the
globules, furnishes less glycerin and succinic acid, and
more alcohol.
A feeble acidity of the liquor seems also to diminish
the proportion of the two secondary products; the contrary occurs if the medium be neutral. Yet Pasteur
himself says, that we usually find in wines a very large
proportion of glycerin and succinic acid, although the
fermentation of the must of the grape takes place in
an acid medium, in presence of a sufficient quantity of
albuminoid and mineral matters.
Whatever these variations may be, it is not less true
that glycerin and succinic acid were never deficient in
more than a hundred analyses of fermentation made
by Pasteur.
The following is the method followed by M. Pasteur,
to detect and measure quantitatively the glycerin and
succinic acid contained in a fermented liquid. The
liquid, when the fermentation is over, and all the sugar
has disappeared (which requires from fifteen to twenty
days under good conditions), is passed through a filter,
accurately weighed against another made of the same
paper. After having been dried at IooO C. (2I2~ F.), the
dried deposit of the ferment which is collected at the
bottom of the vessel is accurately weighed. The filtered
liquid is subjected to a very slow evaporation (at the
rate of from 12 to 20 hours for each half-litre).




28              ON FERMENTATION.
When it is reduced to Io or 20 cubic centimetres ('6I or
1'22 cub. in.), the evaporation is finished in a dry vacuum.
The sirupyresiduum in the capsule is treated several times
with a mixture of alcohol and ether, formed of one part
of alcohol at 300 or 32~, and I 2 parts of rectified ether.
After six or seven washings, there remains no more
succinic acid or glycerin. The etherized alcoholic liquid
is distilled in a retort, then evaporated in a water-bath
in a capsule, and afterwards in a-dry vacuum. Pure
lime water is added to the remainder, till it is neutralized; it is then evaporated afresh, and the dried mass
is again treated with the mixture of alcohol and ether,
which only dissolves out the glycerin, leaving the
calcium succinate in the form of a crystallized powder,
stained with a small quantity of extractive matter, and
with an uncrystallizable salt of lime. The calcium
succinate is easily purified by treating it with alcohol
at 8o per cent., which only dissolves out the foreignri
matters. The etherized alcoholic solution of glycerin
is evaporated, and weighed after dessication in a dry
vacuum.
Among the products met with normally and constantly in all alcoholic fermentations of sugar, we ought to
mention acetic acid. The formation of this body noticed
by Bechamp (Comp. Rend. de l'Acad., I863), was at first
attributed by Pasteur to a concomitant or subsequent
acetic fermentation, and to the presence of Mycodernza
aceti; but since the very precise and conclusive researches of Duclaux (Theses Presentees ta la Faculte
des Sciences, I865), it has been established: Ist. That
the acetic acid is never deficient, even in fermentations
conducted in the most careful manner, in order to pre



ALCOHOLIC OR SPIRITUOUS FERMENTATION.  29
serve them  from  contact with air; 2ndly. That the
proportion of this acid is remarkably constant, especially
if we are careful to stop the fermentation as soon as all
the sugar is transformed; besides this, it does not exceed 0o5 per cent. of the weight of the sugar. This
proportion of acetic acid is considerably augmented if
the fermentation be continued beyond the limits just indicatedo As we shall presently see that the ferment, when
left to itself, without sugar and without oxygen, can
form acetic acid, by acting on its own elements, we can
understand the observed augmentation in the weight of
this acid, and we shall be inclined to attribute its production in a general manner to the transformations
undergone by the ferment while it acts upon the sugar.
We may say the same thing of the leucine and tyrosine
found by B.hchamp in the extract of fermented glucose.
These are compounds in the production of which the
sugar and the fermentable matter take no part.
However, the later works of Bechamp on this question
are not favourable to this opinion concerning the acetic
acid (Comp. Rend., vol. 75, p. Io36). They tend to
establish: Ist. That the contact of air, far from augmenting the production of acetic acid, diminishes it.
A fermentation which in carbon dioxide produces from'25 to'40 of this body for Ioo of sugar, gives only'I in
contact with air; 2nd. That the acetic acid comes from
the sugar, and not from the ferment, for by arranging the
experiments suitably, we may obtain a weight of acid
greater than that of the ferment employed. In proportion as the ferment is well nourished, and the better
it multiplies, the less acetic acid it yields. That which
has only cane sugar for its nourishment is exhausted,




30              ON FERMENTATION.
and produces more acetic acid. The temperature and
the augmentation of pressure tend to increase this
phenomenon.
Most of the natural saccharine juices, such as those
of beet-root and grape-cake, give rise to the production
of small quantities of alcohols homologous with ordinary
alcohol. We find, in fact, when large masses of products are operated upon, in the arts, and crude alcohol
is distilled carefully, by means of suitable rectifying
apparatus, that there is a residuum less volatile than
ethyl alcohol, having a strong and disagreeable odour.
This oily residuum, known under the name of oil of
potatoes, has been made the object of many researches
by Chancel, Wurtz, Pelletan, Faget, and others; they
have generally found it composed in great part of
propyl alcohol, C3 HI  0, butyl alcohol C4 H10 O,
dominant amyl alcohol, C5 H12 0, caproic alcohol,
C6 H14 0, cenanthyl alcohol C7 H16 0, and caprylic
alcohol C8 H1s O0.
M. Jeanjean has, besides, ascertained, in the products
of the distillation of the water in which fermented
madder has been washed, the presence of camphyl
alcohol (camphor of Borneo, C10 H16 O).
We may ask whether these secondary products,
which  are  relatively not very abundant, owe their
origin to alcoholic fermentation properly so called, or
to distinct concomitant fermentations, having each a
special ferment; or whether, in fact, it is better to
attribute their appearance to special principles accompanying glucose in the natural saccharine juices.
* I have followed Schorlemmer in using the terms, ethyl alcohol, propyl
alcohol, instead of ethylic. The change is unimportant.




ALCOHOLIC OR SPIRITUOUS FERMENTATION.  31
The actual state of science does not allow us as yet
to answer these questions definitively.
M. Berthelot points out (Ch. Org. Fondee sur la Synthese, vol. 2, p. 63I) that the production of all these
homologues of ordinary alcohol at the. expense of sugar
may be formulated by the general equation:n1  C6 H12 06}- Cn H2l + 02
n       n1-2
+2 _C 02 +    H2 O.
OF THE FERMENTABLE BODY.
The progress of chemistry has taught us to distinguish many varieties of saccharine hydro-carbons,
differing either in their properties or in their composition; they do not all show the same characters, when
they are subjected to the influence of the special
alcoholic ferment, the yeast of beer.
Glucose (grape sugar or starch sugar) levulose or
the sugar of acid fruits, uncrystallizable sugar, maltose
or sugar of malt, formed by the action of diastase on
dextrin, lactose or sugar derived from sugar of milk
(lactive) by the action of acids, all have the same
formula, C6 H12 0.
An almost entire resemblance between their behaviour in presence of a ferment corresponds with this
analogy in their composition. These sugars are split up
progressively into alcohol and carbon dioxide without
undergoing any previous transformation-as Mitscherlich has observed (Ann. de Poggen., vol. 135, p. 95), the




32              ON FERMENTATION.
rotatory power of a solution of glucose diminishes in
proportion to the quantity of alcohol produced.
The equation of Dumas and Gay-Lussac, modified by
Pasteur, applies without any restriction to these various
saccharine matters. We will only add that, according
to the interesting observations of Dubrunfant, glucose
mixed with levulose, with the addition of yeast, ferments
sooner than the latter does when by itself. This is
what always happens when, having altered cane sugar
by an acid, we subject to fermentation the mixture of
equal weights of glucose and levulose which results
from this alteration; the glucose disappears before the
levulose, which, last of all, undergoes alcoholic decomposition. Dubrunfant has given this phenomenon the
name of elective fermentation.  The sugars whose
composition is represented by the formula C12 H22 Oil
can also be fermented, but only on the condition of
being previously hydrated, which converts them into
sugar with the formula C6 H12 0.
Saccharose or cane sugar is changed, when hydrated,
into two isomeric molecules, one of which crystallizes
and causes the plane of polarized light to deviate to
the right, and the other remains uncrystallizable, and
turns it to the left (levulose).  The two products of
this splitting up are fermentable, the hydratation, as
is well known is effected under the influence of acids,
of water only, of light, and of the lower cellular plants.
It may be understood, from this last observation, why
cane sugar can ferment: as soon as it is placed in
contact with yeast, it begins to alter, and afterwards
the glucoses produced form  alcohol.  The ferment
therefore plays a double part towards the saccharine




ALCOHOLIC OR SPIRITUOUS FERMENTATION.  33
matter, of which the first is much simpler than the
second. The alterative power is due to the presence in
the ferment of a soluble and inorganic nitrogenous
principle, formed at the expense of the proteids of this
organism. This active alterative substance accumulates
more especially in great proportion in ferment which
has been left to itself, and which has undergone the
phenomenon called softening.
The water in which such ferment has been washed
alters cane sugar with such rapidity that, when we'nix the two liquids (sweetened water and the water
in which the ferment was washed), the liquid rapidly
reduces Fehling's liquid poured into it some seconds
after.  We shall return to this order of phenomena
when we speak of indirect fermentation, called fermentation by a soluble ferment.
Meletizose, melitose, and lactine or sugar of milk,
are in the same category as cane sugar, and must be
previously hydrated. Berthelot observed this remarkable peculiarity in melitose; only half of this' sugar is
decomposed into alcohol and carbon dioxide, the other
is transformed into a compound, isomeric with glucose,
namely, eucalin, which' is not fermentable..
All bodies capable of producing glucose and its
congeners by hydratation, belong  to  the class of
indirectly fermentable substances, such as starch,
dextrine gum, glycogea, and the various glucosides
which are found in vegetable tissues.




34               ON FERMENTATION.
CHAPTER III.
ALCOHOLIC FERMENTS.*
WE have hitherto considered bodies susceptible of
alcoholic fermentation, and the details of the reaction
known by this name; there remains for us to speak of
the most interesting part of our subject, the exciting
cause of fermentation. It is more especially on this,
the most obscure and the most difficult part of the
question, that the most varied, and, we may say, the
most lively discussions have been raised. The other
parts of the problem required for their solution, in fact,
only good analysis and rigorous quantitative experiments.
Historical.-Leuwenhoeck was the first, in I68o, to
examine beer-yeast by the microscope, and to ascertain
that it is formed of very small spherical or ovoid
globules.  He could  not, however, determine their
nature.
In his memoir on fermentation, presented to the
Academy of Florence (I787), Fabroni compared ferments to  animal substances.  "The matter which
decomposes sugar is a vegeto-animal substance; it
resides in peculiar utricles, in grapes as well as in corn.
* Cfr.-Pasteur, "Annales de Chimie et de Physique," 3rdseries, vol. 58, p.
364; Monoyer, "These de MWdecine," Strasburg, i862; L. Engel, "These
pour le Doctorat Bs Sciences," Paris, I872.




ALCOHOLIC FERMENTS.              35
When the grapes are crushed, this glutinous matter
is mixed with the sugar; directly the two substances
come in contact, effervescence and fermentation commence."
The experiments and conclusions of Fabroni did not
seem sufficiently to elucidate the question, for in the
year VIII. the class of physical sciences of the institute proposed as the subject for the prize the following
question:" What are the characteristics which, in vegetable and
animal matters, distinguish those which serve as ferments from those which they cause to undergo fermentation? " Three years afterwards,* Thenard presented a
remarkable memoir on alcoholic fermentation and ferments. He arrived at the conclusion that all natural
juices, when spontaneous fermentation has set up, give
a deposit which has the appearance of beer-yeast, and
like it, is able to ferment pure sweetened water. This
ferment is of an animal nature; it is nitrogenous, and
gives over much ammonia when distilled. When
Thenard said that ferment is of an animal nature, he
considered only its chemical composition, and made no
allusion to the organization of the ferment. We shall
return to the labours of this investigator when we
come to study the transformations undergone by the
ferment during the act of fermentation.
Gay-Lussac proves, by well-known experiments, that
fermentation is only developed in the must of grapes
when it has been placed for a moment in contact with
air; he concludes, from his experiments, that oxygen
"Ann. de Chimie," vol. 26, p. 247




36'ON FERMENTATION.
is necessary to commence the fermentation; but that
it is not required to continue it.
In I828, Colin made many experiments which seem
to show that a great number of organic nitrogenous
substances differing from yeast, and in process, of
change, are able, when placed in sweetened water, to
set up alcoholic fermentation at the end of some hours;
at the same time the fcetid odour of putrefaction is
changed into the agreeable smell of the must of wine
(Colin, Ann. Chim. Phys., 2nd series, vol. 28, p. I28,
1828).
The question of fermentation had reached this point,
and yeast was regarded as an immediate principle
of plants, having the property of becoming precipitated in presence of fermentable sugars, when Cagniard de Latour took up the incomplete microscopical
observations of Leuwenhoeck, which had been so long
forgotten.
tI-e observed that yeast is a mass of organic globules,
susceptible of reproducing themselves by means of
buds, or seminules, which appeared to belong to the.
vegetable kingdom, and not to be simply organic or
chemical matter, as had been supposed.  He concluded that it is very probably by some effect of their
vegetation that the globules of yeast disengage carbon dioxide from the saccharine liquid, and convert
it into spirituous liquor. (Ann. Chim. Phys., 2nd
series, vol. 68.)
The discovery of Cagniard de Latour was again
made almost at the same time, but independently,
by Dr. Schwann at Jena, and by Kiitzing at Berlin
(Schwann, Poggen. Ann., 1837, vol. 41, p. I84; Kuit



AL.COHOLIC FERMENTS.              37
zing, Journ. fur -Prakt. Chem., p. 35); confirmed
by the observations of Quevenne (Journal de Pharm.
2, vol. 24), of Turpin (Comp. Rend. de l'Acad., 4, p.
369), of Mitserlich (Poggen. Ann., 55, p. 225), it led,
without any possible contradiction, to the following
conclusions respecting the nature of yeast. This body
was considered to be a mass of organized and living
cells, composed, like vegetable or animal cells, of an
envelope and granular contents.
From  the very commencement, it was not agreed
what place should be assigned to this new form of life.
Some- saw in it a fungus without a mycelium, others
looked upon it as one of the algae.
Thus Turpin (Comp. Rend., p. 379, I838) placed
the cells of yeast in the genus Torula of Persoon, in
which " sporm in floccos moniliformes concatenatre, dein
seudentes."  These cells were thus compared to spores,
without'considering their mode of production, which is
quite different, and without remarking that the Torula
has a mycelium, which is never found in ferments.
The discovery of true spores has since proved that
ferments cannot be placed in the family of the Torulaceae. Meyen (Pflanzen Physiologie, vol. 3, p. 455),also considered that yeast was a fungus, and created a
new genus for it, under the name of Saccharomyces.
This name was adopted by Rees, Engel, &c.; Kiltzing, on the contrary, with many other authors, maintained that ferments are alga, which he arranged in a
separate genus, cryfpto-cocczus.
The opinion of men of science who wvisli to assimilate yeast and ferments in general to algae, was founded
on the observation that these cells multiplied by budding




38              ON FERMENTATION.
only. But we shall soon see that by placing these ferments under certain conditions, we succeed in causing
them to fructify; besides this, algae almost always contain chlorophyll, which fungi and ferments do not.
It is now, therefore,. very generally admitted that
ferments are fungi.  Without in the least detracting
from the merits of Cagniard de Latour, we ought to
say that he had been anticipated in the field of microscopic observations, not only by Leuwenhoeck, but
also by Kieser (I814, Schweigger's Journal, I2, p. 229),
who describes it as formed of small transparent motionless spherical corpuscules, all of nearly the same size;
also by Desmazieres, who, in 1826 (Annales des Sciences
Naturelles, vol. Io, P. 4), examined the pellicle formed
on the surface of beer, and called by Persoon mzycoderma cerevisice.
Desmazieres was the first to give a representation of
the globules which he had observed, and having seen
in them a particular movement, which is nothing more
than the Brownian movement, which at that time was
unknown, he arranged these globules among the animazlcula monadinfa. Astier, as early as I8I3 (Ann. de
Chimie, vol. 87, p. 271), did not hesitate to affirm that
ferment, recognized as an animal substance by Fabroni,
was alive, and derived its nourishment from the sugar,
whence resulted the rupture of equilibrium between the
elements of this body. By this theory, it is easily explained, said he, that all the causes which kill animals,
or hinder their development, must be opposed to fermentation.
The observers who had demonstrated the organic
nature of ferments established at the same time that




ALCOHOLIC FERMENTS.             39
in a great number of liquids in alcoholic fermentation
(such as natural saccharine juices, solutions of sugar
with albumin, &c), globules of ferment are formed, as
Thenard had observed. Schmidt of Dorpat arrived at
the same conclusions by repeating Colin's experiments
on the fermentation excited by albuminoid matter in
the process of decomposition. Microscopic observations revealed to him the development of globules of
ferment whenever there was the production of alcohol.
From all these successive observations, which were
complementary to each other, arose the opinion generally
admitted, that yeast accompanies every well marked
alcoholic fermentation; it seems, therefore, that the
theory propounded by Astier and Cagniard de Latour
concerning fermentation, ought to have prevailed, and
to have been received by men of science. But this was
not the case. From the very commencement of the
discussion, the conclusions of Cagniard de Latour and
of Schwann found a powerful opponent. Liebig, whose
name was then an authority in chemistry, had a decided
theory concerning the phenomena of fermentation in
general, and he defended it with vigour, even after the
experiments of Pasteur, who admits that alcoholic fermentation is an act connected with life, and with the
organization of globules.
"My most decided opinion," says Pasteur, "on the
nature of alcoholic fermentation is the following: The
chemical act of fermentation is essentially a correlative
phenomenon of a vital act, beginning and ending with
it. I think that there is never any alcoholic fermentation
without there being, at the same time, organization,
development, multiplication of globules, or the con



4.0'ON FERMENTATION.
tinued consecutive life of globules already -formed."
As to the hypotheses which tend to go more deeply into
the physiological cause of decomposition, M. Pasteur
neither admits nor rejects them, at least in his first
memoir. Thus the ideas of Pasteur confirm and extend
those of Cagniard de Latour.
As to the theory of Liebig, it does not differ from
that of Willis and Stahl. According to the German
chemist, the cause of fermentation is the internal
molecular motion, which a body in. the course- of decomposition communicates to other matter in which
the elements are connected by a very feeble affinity.
"Yeast, and, in general, all animal and vegetable matters
in a state of putrefaction, will communicate to other
bodies the condition of decomposition in which they
are themselves placed; the motion which is given to
their own elements by the disturbance of equilibrium is
also communicated to the elements of the bodies which
come into contact with them. (Liebig, Ann. de Chimie
et de Phys., 2nd series, vol. 7I, p. I78.) This very
philosophical and seducing explanation of an obscure
phenomenon obtained greater credit among men of
science because it gave the key, not only to alcoholic fermentation, but also to other phenomena of the same
kind, such as the transformation of sugar into. lactic
and butyric acids, in which organic products had not'hitherto been observed, and which seemed only to be
the results of a conflict between a fermentable substance
and a nitrogenous substance in process of putrefaction.
Fremy and Boutron supposed that, in matters capable
of acting as ferments, the character of the fermentation
varies with the degree of decomposition of the sub



ALCOHOLIC FERMENTS.               41
stances. This would be successively alcoholic, lactic, or
butyric ferment according to the more or less advanced
state of its decomposition.
It is thus that the recognized invariable presence of
an organic body in every liquid in process of alcoholic
fermentation began, by degrees, to be considered as a
fact of slight importance with regard to the reaction;
the latter is excited, not by the direct action of the
globules of ferment, considered as a living organism,
but by the decomposition of the proteic nitrogenous
matter of this ferment, regarded only as nitrogenous
substance.   The  experiment of  Gay-Lussac was
naturally interpreted by this opinion; the momentary
presence of oxygen was indispensable to set up the
molecular disturbance of the albuminous matter of
the must of grapes.
Berzelius, for his part, treated the organic nature of
yeast as a poetico-scientific reverie, and, rejecting the
doctrine of Liebig, borrowed from Willis and Stahl,
would only see in fermentation an act of contact due to
catalytic force, and in yeast an amorphous principle.
Mitscherlich supported the ideas of Berzelius, while he
admitted the organic nature of the ferment.
However, the clear and well conducted researches of
Pasteur on fermentation, and especially on alcoholic
fermentation, had partly reconciled men of science to
the physiological theory; wherefore Liebig thought right
to recommence the contest in favour of his own ideas.
In I870, he published a long memoir on fermentation
and the source of muscular force (Ann. der Chemie
und Pharmacie, vol. 153, P. I), a memoir in which he
sought to show that the principal experiments of Pasteur
3




42              ON FERMENTATION.
are not conclusive. He first demonstrated that the
physiological theory of Pasteur, which explains the decomposition of sugar by the nutrition and development
of an organic substance, is not opposed to the mechanical doctrine of which he is the champion. This avowal
was already an enormous concession made by the German chemist, almost an avowal of defeat, for this language is very different from that which he had formerly
used.
However, the attack was sufficiently powerful to
induce Pasteur to reply to it (Ann. Chimie Phys., 4th
series, vol. 25, p. I45, I872), and Dumas to undertake
a series of experiments, in order to ascertain if it were
possible to verify the consequences of Liebig's theory.
Dumas, by means of ingenious experiments, conducted with his never-failing precision (Ann. de Chimie
et de Physique, 5th series, vol. 3, p. 69), succeeded in
proving irrefutably; Ist. That saccharine liquids are
not influenced by ferment, even through the shortest
columns of liquid, the thinnest membranes, or even without any separating medium; and that its immediate
and direct contact is necessary; 2nd. That sonorous
vibrations have no influence on the movements of
fermentation; 3rd. That no chemical action, among a
great number of those which have been tried, has been
able to effect the decomposition of sugar into alcohol
and carbon dioxide.
These negative results, without bringing about any
decisive solution of the question, are, however, contrary
to the opinion of a transmitted movement.
We may thus sum up these three great theories of
fermentation: Ist. The vitalist theory, formulated by




ALCOHOLIC FERMENTS.              43
these words of Turpin, "Fermentation as effect, and
vegetation as cause, are two things inseparable in the
act of decomposition of sugar," maintained by Astier,
Cagniard de Latour, Schwann, KiUtzing, Turpin, Bouchardot, Van de Brock, Shroeder, Pasteur, and Bichat;
2nd. The mechanical theory of Willis, Stahl, and
Liebig, admitted by Gerhardt; 3rd. The theory of
catalytic forces, and of acts of contact, maintained by
Berzelius and Mitscherlich.
Various mixed opinions range themselves by the side
of these three theories. Thus M. Berthelot considers
fermentation as produced by the action of a substance
elaborated by organic ferments, comparing, with this
idea, the alcoholic and lactic fermentations to the conversion of starch into dextrine and sugar under the
influence of diastase, a soluble inorganic ferment. The
learned chemist supports his opinion by experiments
which prove that, in certain cases, there may be the
production of alcohol without the formation of ferment
(Chimie Organique Fondee sur la Synthese); which does
not exclude the fact, now distinctly established, that
fermentation may be excited, and is indeed energetically
originated, by special organic substances.
As to a more precise relation between chemical
phenomena and the physiological functions of the
organic ferment, it is still to be discovered; and all
that has been said, written, and brought forward to
decide the question needs experimental proof, and can
only be considered by us in passing.
No one doubts that, in organic living cells, whether
they be isolated, like those of yeast, or form an integral
part of a more complicated organism, there resides a




44              ON FERMENTATION.
special force, capable of producing chemical reactions
under conditions quite different from those which we
employ in our laboratories, and to produce results of the
same class. This force, which we imagine to'be as
material as heat, reveals to us its activity by decompositions effected on complex molecules. Whether we
reduce the problem to the action of a soluble product
elaborated by the organic ferment, and to which it has
communicated its power, or suppose that the whole of
the ferment exercises an action of this kind, we ultimately
arrive at a motion communicated, more or less directly,
by vital force, and dependent upon it.
We must not confound this interpretation of the
phenomena with Liebig's theory; the German chemist
held that the albuminoid principles, in decomposing
spontaneously, produce the molecular motion which is
transmitted to the sugar to split it up; here, on the
contrary, it is the living organism which develops force,
by borrowing it from the great external reservoir and
transforming it. This force may act chemically, either
directly or indirectly by means of a soluble ferment.
Descriptioln of thze Frmeinzt.-We will give, with Rees,
the name Saccharomyces cerevisiw, to the alcoholic
ferment of beer; it is the ferment which has been
most thoroughly studied, and which is the most easily
procured.
There are three methods of causing the wort of beer
to ferment; the two first are generally employed; these
are surface and sedimentary fermentations.
In the third method, employed in Belgium only, the
wort is left to itself in a locality situated above the
level of the ground, and the spontaneous development




ALCOHOLIC FERMENTS.                  45
of fermentation is waited for; in the two former instances, the action is excited by mixing with the wort
a suitable proportion of yeast arising from an anterior
operation of the same class.
In beer brewed by surface fermentation, the starch of
the malt is changed into sugar by its being steeped
several times successively; fermentation takes place in
casks at a comparatively high temperature, from I5~ to
18~ C. (59~ to about 650 Fahr.)  The yeast, in this
case, as it is formed, rises through the bung-holes,
at the upper part of the cask. In England, this fermentation is carried on in large open vats; the yeast
then floats on the surface of the liquor, and can be
skimmed off.
In the manufacture of beer brewed by sedimentary
fermentation, the saccharification is effected by deFIG. x..-Saccharomyces cerevisize-Yeast of sedimentary beer, X 400o.
coction, and the transformation takes place in open
vats, at a temperature not exceeding from I2~ to I4~ C.
(from about 53~ to 580 Fahr.)  The yeast is deposited
at the bottom of the vats, and adheres in the form of a
pasty mass.  When  once the first and most active
fermentation is over (it usually lasts two or three days
for the surface fermentation, and eight or ten days for
the sedimentary), the clear liquid is drawn off, and kept




46              ON FERMENTATION,
in casks, glass, or stone bottles. In the meantime the
separation of the yeast has not been completed; it
continues to act on the still unmodified sugar; therefore
the amount of alcohol and carbon dioxide yielded
increases with the time of keeping, and at the same
time the liquid becomes turbid by the production of
fresh yeast.
The yeast greatly exceeds (seven or eight times), in
weight and in volume, that which had been previously
introduced into the wort. To this fact, which we now
merely notice in passing, we shall return presently; it
is explained by the multiplication by buds, which takes
place whenever the cells of yeast are placed in a saccharine medium suitable to their development; and
the wort of beer affords excellent conditions in this
respect. After the sedimentary fermentation, the yeast
found at the bottom of the vat is composed, almost
entirely, of cells of a single species of alcoholic ferment,
the Sacc/laromzyces cerevisive (Fig. I). The microscope will
show, but in a very small proportion, granules of lupulin, crystals of calcium oxalate, spores and mould.
This deposit is of the consistence of a paste of a yellowish white, or yellow-ochre colour.
The cells are round or oval, from  x~  to _-_0of a millimetre (from'ooo3I to'00035. in.) in their
greatest diameter.  They are formed of a thin and
elastic membrane of colourless cellulose, and of a protoplasm, also colourless, sometimes homogeneous, sometimes composed of small granulations. We find in the
protoplasm one or two vacuoles, of various sizes, containing cellular juice. The cells are either separate, or
united two by two.




AL,COHOLIC FERMENTS.                 47
When these cells are deposited in a fermentable
liquid, we soon see at one, and more rarely, at two points
of their surface, vesicular prominences arise, the interior
of which is filled at the expense of the protoplasm of
the mother-cell; these prominences enlarge, and at last,
having attained the size of the original cell, they lessen
in diameter at the base (Fig. 2). They usually originate
at the widest side, but more rarely at the extremities.
As soon as the formation of this kind of neck takes
place, the new cells separate with considerable rapidity
from the mother-cell, in which the protoplasm, given up
to the young cell, is replaced by one or two vacuoles.
If the conditions are favourable, the same cell is able to
produce several generations of cells; but, by degrees, it
loses all its protoplasm, which at last unites in granules
swimming in the midst of superabundant cellular juice.
The cell then ceases to reproduce, and. even to live; the
membrane is ruptured, and the granular contents are
diffused in the liquid.
FrI. 2. - Saccharomyces cerevisix-    FIG. 3. — Saccharomyces cerevisia —
Yeast of sedimentary beer, budding, X   Yeast of surface beer, budding, x 400.
400.
When the SaccSaromtyces cerevisic  is not in contact




48              ON FERMENTATION.
with a fermentable liquid, it may remain for some time
without becoming modified.
The isolated cells of the surface ferment (Fig. 3) do
not differ sensibly from those of the sedimentary ferment; and although it has been maintained that the
larger and oval forms are more prevalent in it, it is difficult to establish any distinction of this kind, for we find
in the two varieties all the intermediate forms between
the two extremes.
Besides, an elevation of temperature above 14~ C.
(57'2 Fahr.) during the fermentation is sufficient to
augment considerably the size of the sedimentary
cells, to cause them to attain a large diameter, from
&-c, to     4  of a millimatre (00ooo05I to'ooo55 of
an in.), giving them a long oval form; at the same
time, we see two circular vacuoles make their appearance; one, large, situated near the larger end;
the other, smaller, is found in the narrow part of the
cell.
The surface Saccaoaromyces buds much more quickly
than the other variety, when placed in a fermentable
liquid (Fig. 3). This budding is very rapid; the different
cells which issue from  each  other remain  attached
together, forming small ramified chains, composed of
from six to twelve, and even more, individual buds. It
may be easily understood that the bubbles of gas
adhering to these chaplets have greater hold upon them
than on an isolated cell; this causes the newly-formed
yeast to be raised towards the surface of the liquid, and
this is effected the more rapidly when the fermentation
is more active.  In these chaplets, the cells have an
elliptical form.  The only well-ascertained difference




ALCOHOLIC FERMENTS.                  49
between the two kinds of yeast is, therefore, the rapidity
with which buds are formed, and the greater activity of
FIG. 4. —Saccharomyces cerevisix-Sur-  FIG. 5.-Saccharomyces cerevisie —Sediface yeast, at rest, x 400.  mentary yeast, in a growing state, x 400.
the ferment in the one case; but this does not authorize
us to consider these two ferments as belonging to different
species.  We are able, indeed, though with great difficulty, by changing their conditions of existence, to
transform one into the other.
Multiplication by buds is not the only mode of reproduction of the Sacc/za-ominyces cerevisz.. It is true that
it alone appears as long as the yeast is in contact
with an appropriate fermentable liquid. We owe to
Rees (Botanische Zeitung, December, I869) the discovery of the fructification of yeast, and of ferments in
general; that is to say, their reproduction by means of
spores. As to the Sacc/aroonyces, the conditions which
appear to be peculiarly favourable to this special evolution of the fungus, are to deprive it suddenly of all
1nourishment, especially saccharine, and to expose it
to a damp atmosphere, or, still better, to place it on a
substance capable of affording it sufficient and constant
humidity.
We obtain, according to Rees, the richest production
of spores by leaving yeast previously washed several
times, for some days in contact with distilled water, and




50              ON FERMENTATION.
then, having decanted the greater part of the water,
later on removing day by day the small portions of
water which become separated from it. Under favourable conditions, we obtain, at the end of fifteen or sixteen days, a very rich formation of spores; but very
often this result is prevented by the putrefaction of the
yeast.
M. Engel, from whose essay we borrow the greater
part of these details, has also studied this question: he
proposes to cause the yeast to fructify, by the following
contrivance:We mix up some plaster, and allow it to run on some
polished, but not oily surface, such as window-gfass,
plate-glass, or marble. We make up the mass into
some form corresponding with the interior of the vessel
in which we intend to preserve it. The dimensions in
each direction ought to be about two centimetres
('78 in.) smaller than the internal dimensions of the
vessel, so as to allow a space between the sides and the
mass of plaster sufficient to pour in some distilled
water. We then take some very fresh yeast, decant as
far as possible all the supernatant fermentable liquid,
and dilute the yeast with distilled water, so as to bring
it to the consistence of very fluid broth; we pour some
drops of this liquid on the polished surface of the
plaster, inclining the mass in every direction so as to
spread the solution uniformly. This operation should
be performed quickly, for the plaster absorbing the
water very rapidly, the diluted yeast would become too
thick, would not spread with sufficient uniformity, and
the layer 6f ferment would be too thick in certain parts.
We then place the mass in the vessel, with the part




ALCOHOLIC FERMENTS.                  5I
covered with yeast upwards, and pour, by means of a
funnel, distilled water between the sides of the vessel
and the piece of plaster, until the liquid reaches above
a centimetre ('39 of an in.) beneath its upper surface.
The vessel is then covered with a plate of glass to
prevent, as far as possible, the contact of dust, and of
spores floating in the atmosphere.
Under these conditions, the vegetative life of the yeast
ceases suddenly, and in a few hours we see  great
changes take place in the protoplasm of the cells. The
oldest, and those which are least rich in protoplasm,
perish and break up. Others, on the contrary, grow
larger, their lacuna~ disappear, and the protoplasm is
diffused uniformly in the cellular juice. At the expiration of from six to ten hours, we notice the appearance,
in the midst of the protoplasm, of from two to four small
6islets," more brilliant and dense than the rest, around
which fine granulations collect. These dense spots do
not present any appearance of a nucleus, and they
become differentiated more and more till they are
FIr. 6.-Saccharomyces cerevisie —For-   FIG. 7. —Triads of spores, germinating,
mation of spores, x 750. a, b, c, d, e, suc-    X 750.
cessive phases of the production of spores.
exactly spherical (Figs. 6 and 7). Twelve or twentyfour hours later, each of these spherules is invested with
a membrane, very thin at first, but which thickens by




5'2             ON FERMENTATION.
degrees, and then shows a double outline, when magnified 6o00 diameters.
The spore is then ripe. The mother-cell thus contains from two to four spores. When there are but two,
they are situated in the greater diameter; when there
are three, they are usually arranged in a triangle; when
there are four, they are in the form of a cross, or three
of them form a triangle, on which the fourth is superposed in the manner of a tetrahedron.
During their evolution the spores touch each other;
a plane surface is therefore produced at the point of
contact; they remain attached to each other during
some time after maturity, and thus form combinations
of two, three, and four. The two spores connected together have only one plane surface, the triads have two,
inclined to each other at about I2o~, and in the tetrads
arranged in the form of a cross we observe, also, two
plane surfaces at right angles. When the spores ripen,
the thecme are moulded on them, and thus assume their
various forms. The theca of the diads is elliptical;
that of the triads is triangular, with rounded angles;
that of the tetrads, in the shape of a cross, is in the form
of a diamond with rounded angles; in the tetrads, piled
up on each other, the theca is tetrahedral; when in
complete maturity, the membrance of the spore case, or
mother-cell, which has become a fruit, is torn, and allows
the spores to escape. The theca: themselves vary from
I 3o to l5 - of a millimetre ('ooo39 to'ooo47 of an in.),
and the spores from -0v4 to -ijo of a millimetre (oooI 5
to ooo  9 in.)
The innumerable quantity of theca which we obtain
by the method of fructification on plaster, leaves no




ALCOHOLIC FERMENTS.                       53
doubt as to the origin of these organisms; besides, Rees
and Engel have often observed thecxe still attached to
vegetative cells, and those in process of budding, and
have thus recognized their relationship.
Hitherto, all that we have said applies to sedimentary
beer, properly so called, to the SacchaCromyces cerevisie;
but this special fungus is not the only one capable of
determining the alcoholic fermentation of glucose.
Microscopists who have studied this difficult question
distinguish, besides the special ferment of beer, other
varieties, for the most part belonging to the genus Sacchar-ormyces of Meyen.*
Under  the  name of Saccharolzyces  wnizor,  Engel
describes a kind of ferment obtained by him from the
leaven of flour, and to which it owes its activity.
The process of extraction is similar to that employed
by chemists to  separate starch from  gluten  in flour.
The liquid which passes through the sieve when bakers'
yeast is kneaded under a slight stream of water, contains starch and globules of yeast, which, on account of
their smaller density, are deposited last. We may thus,
by a series of washings, obtain a product very rich in
globules of yeast, and poor in grains of starch. Engel
proposes to employ sweetened instead of pure water in
these washings, in order not to diminish the physiological activity of the cells.
* Simple thecaphorous fungi, without a true mycelium. The vegetative
organs are cells, produced usually by buds from similar cells, and which, detaching themselves, sooner or later, from the mother-cell, multiply in the same
Inanner. A part of the cells thus formed are transformed, in another medium,
into naked sporiferous thece; unicellular spores, to the number of from one
to four in each theca. The germination of spores reproduces directly vegetative cells analogous to those which originate from buds.-E;NGEL, TIzesis of
the Faculty of Sciences of Paris, 1872.




5 4             ON FERMENTATION.
The ferment, examined by the microscope, is seen
under the form of isolated globules, double or sometimes united in threes. The largest of these globules
are 6 of a millimetre (about'ooo0003 I5 of an in.) in
diameter; their vacuoles are less apparent than those
of the yeast of beer.
This ferment, sown in the most favourable saccharine
medium, prepared according to the formula of M. Pasteur, has only produced a very slow fermentation.  By
renewing the experiment seven times, and only making
use each time of the ferment obtained in the preceding
experiment, we see no apparent modification in the form
and dimensions of the globules. The budding of this
species is effected in the same manner as in the yeast of
beer.
Placed under the conditions favourable to the formation of spores, of which we have spoken before, the
Sacc/zaromyces mninZor is transformed into sporiferous
thecae of spherical, and occasionally though rarely of
ovoid form, of  ofY  to  7_  of a millimetre in diameter.  The spores are only x-fo3   of a millimetre in
diameter, and are united in diads or triads. In fact,
except their form, which is always spherical, their smaller
dimensions and greater activity, the ferments of bread
resemble that of beer.
The Saccharomyces ellipsoi'deus of Rees is nothing but
Pasteur's ordinary alcoholic ferment of wine (Etudes
sur le Vin, Figs. 8, 9, and  I I);  it ought not to
be confounded with the Cryptococcius vini of Kiitzing.
The adult cells have an ellipsoidal form T-o6 of a millimetre in length by T-4, or i 5  in breadth (o00024 by
about'OOOI76 in.), with an oval vacuole. The sporula



ALCOHOLIC FERMENTS.                     5 5
tion and budding differ in no respect from the analogous
phenomena which are observed in yeast (Figs. 8, 9,
and Io)o
Rees gave the name of Saccharomyces Pastorialr'ns
(Fig. I ), to a variety of alcoholic ferment of wine
observed by Pasteur (Fig. 7, Etudes sur le Vin). The
cells are oval, pyriform, or elongated like a club.
The ovoid  cells are 1-0  of a millimetre in  length
('ooo236 in.), those that are club-shaped, which are seen
to proceed  like buds from  ovoid cells, reach,,  or
FIG. 8.-Saccharomyces ellipsoideus,   FIG. 9. —Saccharomyces ellipsoideus,
in process of budding X 6oo.   development of spores, x 400.
o~-o  of a milliimetre in length, by T        or   10  i
breadth at the larger end (o00064 in. to'ooo78 in. by'ooo314 in. to'0ooo0397 in.); they are united in flakes
containing from  three to seven articulations.
FIG. Io,
The Saccharomnyccs  exizuus (Rees), Fig.  I 2, is met




55                ON FERMENTATION,
with, like the preceding, in the juices of fermented fruits.
The cells are only x-   of a millimetre in length, by
o a. in width at the larger end ('oooI I8 x'oooo98 in.);
it multiplies by budding and sporulation, like all the
other varieties of this species.
The Saccharomyces conglorneralus of Rees (Fig. I3), is
rather rare; it is met with in the must of wine towards
the end of the fermentation. It has spheroidal cells
of x-6d, millimetre in diameter ('0oo236 in.).
FIG. 12.-Saccharomyces exiguus, X    FIG. 13.-Saccharomyces conglomera350.                          tus, X 600oo.
When the first cell has budded, this bud attains the
size of the mother-cell, without being detached; there
originate first in the inner angle formed by two cells,
and then on different parts of their surface, a considerable number of new cells, which, instead of forming a
chaplet or flakes, are entirely conglomerated.
The apiculated ferment (Carpozyma) does not belong
to the genus Saccharomyces, according to the observations of M. Engel.  This is the most abundant alcoholic




ALCOHOLIC FERMENTS.                  57
ferment. It is met with on all kinds of fruit, especially
on berries and stone-fruits, as well as in the greater
number of musts of wines in process of fermentation.
It has also been noticed in certain kinds of beer, as that
of Belgium, and of Obernai; those of Strasbourg do
FIG. I4.-Saccharomyces apiculatus (Rees). Carpozyma apic. (Engel), apiculated
ferment, X 600.
not contain it.  It is this which is usually the first to
appear and bud in these musts.
The adult and isolated cells (Fig. I4) have the form
of an ellipsoid, whose greater diameter is T-f-6  of a
millimetre ('000236 in.), and the transverse diameter
one-half of the larger one. At each extremity there is a
small projection or minute stalk, which give the cell the
appearance of a lemon. The interior encloses a spherical or ellipsoidal vacuole, around which there is a thin
layer of protoplasm, fringed towards the projecting parts.
The budding cells always present themselves at the
extremity of the projections.
When the development is normal, the new cells extend
in the direction of the principal axis of the mother-cell,
so that the three cells form a longitudinal row; but
when they have ceased to grow, they assume an elliptical form, and bend back at the point of their insertion,
so that their longer axis forms at last a right angle with




58              ON FERMENTATION.
the axis of the mother-cell; one of the cells turns to
the right, and the other to the left. They are then
detached; at this time they much resemble the cells
of Sacckacroryces ellizpsoideus; but we soon see the characteristic stalks appear.
When the apiculate ferment is deposited on damp
plaster, the transformation into thecae or sporanges
assumes phases very different from those which are
observed in the various species of Saccharomyces, and
resembles the evolutions of the Protomzyces mzacrosporus
studied by De Bary.
According to Engel, the apiculate ferment is a Protomyces without a mycelium; this botanist proposes to
give it the name of Carpozyma. Its thecm are spherical,
covered with a peritheca, and are hibernating. The
development of the spores is very slow, and the spores
numerous.
Rees met with a special form of ferment, which
accompanies the Saccharomyces ellipsoideus, in the fermented musts of red wine. It is composed of elongated
cylindrical cells. Although this ferment has not been
observed in the different evolutions of its vegetative
life, it has been thought necessary to establish for it
a special species, under the name of Saccharomyces
Reesii.
The Saccharomyces mycoderna (Figs. i6 and 17), called
flowers of wine, or flowers of beer, ought also, according
to M. Pasteur, to be placed among alcoholic ferments.
In fact, although it does not act in this manner, under
the ordinary circumstances of its development, when it
grows on the surface of fermented- liquors, perhaps
because the alcohol which it may then produce is




ALCOHOLIC FERMENTS.                  59
destroyed by a subsequent oxidation, M. Pasteur has
shown that the Mycoderrma vini, sown in sweetened
water, is able to set up in it alcoholic fermentation.
FIG. Is. —Saccharomyces Reesii, ferment   FIG. I6.-Saccharomyces mycoderma,
of red wine, X 350.          X 350.
It makes its appearance in all alcoholic liquids exposed to the air, when the fermentation is over or has
become languid. It grows with great rapidity; it is
sufficient to place a few of its cells on the surface of a
liquid that easily becomes alcoholic, and we shall find,
in less than forty-eight hours, the surface covered with
FIG. I7.-Saccharomyces mycoderma.
a thin pellicle, of a whitish or yellowish tint, at first
smooth, and then wrinkled.
M. Engel estimated, by calculation founded on his observations, that in forty-eight hours a cell of Mycoderma
viizi would produce about 35,378 cells. The cells of this
MAycoderma have various forms-ovoid, ellipsoidal, and
cylindrical, with rounded extremities.




60               ON FERMENTATION.
The ovoid cells have their greater diameter about f,-0
of a millimetre, and their smaller one 1-j-AT   (about
~000236 x oooI 57 in.). The cylinders have the longer
diameter about  b%-o  or 3f-l4  of a millimetre, and the
lesser one 0U-~- millimetre ('ooo47 x'oooI 8 in.).
The cells are generally poor in protoplasm; they
show in their interior from one to three brilliant points of
fatty matter. The budding is effected at the extremity,
by one or two buds originating at each end. Chaplets
and ramified and interlaced flakes are thus formed, giving
to the whole the appearance of a fine membrane.
When we dilute with a considerable proportion of
water the alcoholic liquid on which the mycoderma
FIG. I8.-Mucor racemosus, ferment in mass.
vegetates, the cells undergo great modifications. The
oldest are weakened and die, allowing their protoplasm
to escape. The others lengthen, acquiring a larger
diameter of from ~iL%-o to -o0j of a millimetre ('ooo629
to'ooo787 in.); their protoplasm  collects in different
points and forms spores. These are usually about three
or four in number, arranged in a longitudinal file in the
cell.  The spores are generally about X',O  millime tre
in diameter ('oooI  8 in.).
Many authors suppose the Sacczharomzyces itycodermaz
to be derived from the Sacc/h. cerevisiwe, one of the aerial
forms of which it represents. This question does not




ALCOHOLIC FERMENTS.               6I
seem yet definitely settled, although there is most
reason to suppose that it constitutes a separate and
independent species.
We shall have to return to the oxydizing properties
of the Mycodernza vizzi when we treat of acetic fermentation.
The Milzicor iziccdo and the Mzlcor raceimoszus (Fig. I8)
possess the property, when immersed in a solution of
sugar, and protected from  the access of oxygen, of
transforming or dividing their mycelium into joints
having the form of balls. These balls are multiplied
by budding, and excite alcoholic fermentation in sugar
as long as they are placed under these abnormal conditions.
This fact, which is indisputably proved, gives considerable support to the theories brought forward by
some men of science as to the transformation of ferments, from one to another, according to the conditions
under which they are placed. We see, in fact, the
Mitcor raccemnoszus completely change its mode of reproduction when it is placed, without access of oxygen, in
a saccharine medium. Analogous facts are known to
be produced in the case of other organisms.
These various kinds of ferments have been found, not
only in the must derived from fruits, but also on the
surface of their pericarps, to which they remain fixed in
a state of repose, until by the concurrence of suitable
circumstances, they are placed in contact with the
saccharine liquid contained in the cells. From this
moment they begin to develop by buds, and set up, at
the same time, alcoholic fermentation.
According to a recent work by Dr. de Vaureal, the




a62             ON FERMENTATION.
alcoholic ferment, with its envelope composed of noncontractile cellulose, and reproducing by gemmation,
which has been generally believed in, is inadmissible.
The supposed budding is only an optical delusion. The
utricle of yeast is allied to the spermogones of Tulasue;
the granulations or nucleolar elements are spermatia;
these elements when set free by the rupture of the
utricle, produce new ones.
This mode of multiplication explains the facility
with which the reproductive elements of yeast can be
carried by the air, when we cannot distinguish in airdust any characteristic globules of yeast.
In their mode of multiplication, ferments resemble
somewhat the zoospores of algxe; when they are not
too hybrid, like those of cider, which reproduce a
penicillium and an aspergillus, they arrange themselves
under the law of metagenesis, like the acalephae.
Especially in the genus Hydrodiction, we notice a great
similarity.
In fact, we see in this zoospores of two sorts; the
greater ones (macrogonidia) are true spores; they have
a rapid development and direct evolution; the smaller
ones (microgonidia) have a slow development; they do
not reproduce the plant, but produce in their interior
true zoospores. These are young spores, like ferments.




ACTUAL COMPOSITION OF FERMENTS.         63
CHAPTER  IV.
ACTUAL COMPOSITION OF FERMENTS.
BEFORE commencing this subject, in which we shall
have to consider, among other questions, the chemical
modifications which take place in ferments under the
different conditions in which they may be placed, we
ought to give a summary of the results obtained by
experimentalists who have devoted themselves to the
study of the chemical composition of these organisms.
Much has been done in this respect. Thus Schlossberger has published very careful researches on the
actual elementary composition of the two kinds of ferment of beer, freed as far as possible, by washing and
decanting, from the impurities which are found in the
crude yeast.
This observer found as the mean of two analyses:SURFACE        SEDIMENTARY
YEAST.            YEAST.
Carbon. 49'9.. 480
Resultsascertained, Hydrogen. 6'6.. 6-5
the ashes having  Nitrogen    21.'6.    9-8
been removed    Oxygen   3I-4            ~ 33'7
Ashes.'5                 3'5
Messrs. Mitscherlich, Mulder, and Wagner have published independently the following results:




64                ON FERMENTATIONo
SEDj.  SURF.  SURF.   SURF.
YEAST.  YEAST. YEAST.  YEAST.
(Wagner.) (Mitsch.) (Mulder.) (Wagner.)
( Carbon   44'4    47'0   50'8   49'8
Hydrogen  6'0    6'6      7'2    6'8
The  ashes  not Nitrogen   92    10       I 
separated.    Sulphur            of
Oxygen   35'8
M. Dumas (Traite de Chimic), finds:Carbon..o 5o'6
Hydrogen..3. 73
Nitrogen... I S-0
Oxygen     ]
Sulphur.27I
Phosphorus'
This result differs from  the others by a larger proportion of nitrogen; but we can understand variations
in the composition of such a product as yeast, which is
constantly undergoing a process of chemical evolution.
Thus, the smaller percentage of nitrogen and carbon
furnished by analyses of the sedimentary, as compared
with that of the surface yeast, is easily explained, if we
keep in mind the fact that the former remains much
longer in contact with the liquid after the fermentation.
Phenomena dependent on spontaneous change of condition may then take place, transforming into soluble
principles a portion of the nitrogenous albuminoid
products of the protoplasm, and allowing them to
escape into the surrounding liquid.
Schlossberger also endeavoured to isolate the various
ultimate principles contained in yeast. By treating it
with a very weak solution of potash, filtering and neu



ACTUAL COMPOSITION OF FERMENTS.            65
tralizing the liquid by an acid, he obtained a floccose
white precipitate, free from ashes, which, when analyzed
for its elements, gave the following numbers:MEAN OF TWO ANALYSES.
Carbon.       e ~.. 55'5
Hydrogen.. 75
Nitrogen.. -.     I3'9
Sulphur...     o'
The nitrogen is here present in too small a proportion for a normal albuminoid compound. The analysis
agrees tolerably well with that made by me of hemiprotein, one of the products of the decomposition of
albumin, by dilute sulphuric acid (see the chapter on
albuminoid substances). This similarity is the more
striking, since the hemi-protein is also soluble in dilute
alkalis, and precipitated by acids.  The precipitate,
when well washed, yields no ashes after combustion.
On the other hand, it is very probable that the yeast
acts on albuminoid substances by splitting them up
progressively, of which we shall find proofs farther on.
By saturating the yeast with acetic acid and precipitating the filtered liquor by ammonium  carbonate,
Mulder obtained a principle nearer in its composition
to albumin, and which gives,Carbon.. 53'3
Hydrogen... 7'0
Nitrogen   e.. I6'o
We may therefore admit the presence of one or more
albuminoid substances in the yeast-cell; in this respect
it does not differ from other vegetable cells.
The residuum insoluble in potass, in Schlossberger's
4




66               ON FERMENTATION.
experiment, was then saturated with acetic acid and
water. It then shows that it has a composition allied
to that of cellulose:
Carbon.         ~.         44'9
Hydrogen. 6'7
Nitrogen. 0'5
Remaining ashes.. I'This cellulose, boiled with sulphuric acid, is easily
converted into fermentable sugar. According to Liebig,
it is, not soluble in ammoniacal cupric oxide.  It
seems, therefore, to differ from  normal cellulose, soluble
in ammoniacal cupric oxide, and which dilute acids do
not transform into sugar.
Payen (Memoire des Savants Etrangers, vol. 9, p. 32)
gives the following direct analysis for yeast:Nitrogenous matter. 62'73
Cellulose (envelopes).. 2237
Fatty matter.,.. 2'Io
Mineral,,.        5'80
Other experimentalists, such as Pasteur and Liebig,
employing the methods of separation usual in the
analysis of the higher plants, have found only 18'5
per cent. of pure cellulose in fresh yeast. If we admit
that the nitrogen (II'8 to I2'5 per cent.) contained in
the yeast, forms an integral part of the albuminoid
matter, we can caculate, by simple rule of three, that
the yeast contains about 6o per cent. of proteids and
nearly 40 per cent. of hydrocarbons.
This method of! calculating the actual composition of
yeast is not quite legitimate.  We find, in fact, in the
washings of fresh yeast, performed in ice-cold water,
noticeable quantities of tyrosine, leucine, &c., which




ACTUAL COMPOSITION OF FERMENTS.            67
contain less nitrogen than albuminoid matters (io and
7'7 per cent., instead of from I5'5 to I6).
As direct analyses give only I8'5, or at most 30, per
cent. of cellulose, one is led to suppose that in the
yeast-cell other hydrocarbons are found, more easily
attacked by acids and alkalis than is true cellulose.
This opinion is corroborated by the  production of
alcohol during the digestion of yeast, without the
addition of sugar, although one cannot discover in it
the presence of glucose.  On the other hand, we find in
the extract of spontaneously decomposed yeast noticeable quantities of a special gummy substance (Bechamp,
Schutzenberger).  The origin of this gum, if it does
not pre-exist fully formed in the fresh cell, can only
be attributed to the splitting up of a compound of
the family'of glucosides, or to a molecular transformation of an insoluble hydrocarbon substance, different
from cellulose.  Pasteur (Compt. Rend., vol. 48, p. 64o)
obtained 20 per cent. of sugar by boiling yeast with
dilute sulphuric acid.
We owe to Mitscherlich (Ann. der Chemie und Phar
macie, vol. 56 ) some excellent analyses of the ashes of
yeast. The dried matter, placed in a silver crucible,
itself placed in one platinum, was burnt in a glass tube,
in a current of oxygen; the distillation of the organic
matter was commenced in an atmosphere of carbon
dioxide, and the combustion was finished in oxygen.
Quantity of ash of the yeast,:SURF. YEAST.   SED. YEAST.    SURF. YEAST.   SED. YEAST.
(Wagner and    (Schlossberger.)  (Bull.)     (Mitsch.)
Schlossberger.)
2'5 per cent.  3'5 to 4 per cent.  8'9 per cent.  7'7 per cent.




68                ON FERMENTATION.
SED. YEAST.          SED. YEAST.
(Mitsch.)           (Wagner.)
7'5 per cent.         5'3 per cent.
Taking only the results obtained by Mitscherlich,
which appear the most reliable, there would be no
difference with respect to the quantities of mineral
matters between the two kinds of yeast. The small
proportion of mineral matter found by Schlossberger
may have resulted from the fact that this experimentalist washed his yeast, an operation which, as Bechamp
has shown, gives rise to  a continuous elimination of
phosphates.
The ash of the ferment gives the following percentage:SURF. FERM.    SED. FERM.   SURF. FERM.
(Mitscherlich.)        OF PALE
ALE (Bull.)
Phosphoric acid.. 53'9.. 59'4   ~. 547
Potass... 398.. 28'3.  35'2
Soda... o05
Magnesia.. 6o            8-I.   4-I
Lime                     I     o  4'3   ~           4-5
Silica.... traces,.
Iron Oxide.., --.. -.. o-6
Sulphuric acid... -..
Hydrochloric acid., o'I
The predominant elements are, therefore, phosphoric
acid and potass, together with a little magnesia and
lime.
The analyses of Mitscherlich may be thus calculated:



ACTUAL COMPOSITION OF FERMENTS.            69
SURF. YEAST.  SED. YEAST.
Phosphoric acid.          4. I-8. 39'5
Potassa... 398.. 28-3
Soda..
Magnesium phosphate,       6'8         226
(Mg.3 2 P04).
Calcium phosphate
(Ca 2 P04) o23                        97
The estimates which we have just given of the direct
and elementary composition of the yeast are still, as
we see, incomplete; and this question is worth attentively studying again.
This opinion has already been brought forward by
Pasteur, who thus expresses himself (Ann. Chim. Phys.
(3) 58, p. 403):"Yeast contains several nitrogenous substances, and
also some not nitrogenous-substances distinct from each
other. It would be interesting to study this subject.
I have found that we should arrive at useful results by
examining separately the action of water, of dilute
sulphuric acid, and of potass. I think that an examination of yeast, made for this purpose, and of the different
materials which compose it, might reveal the secrets oi
certain changes which are observed in the nature of the
extract of the fermented liquid."
Notwithstanding the insufficiency of the data of this
question, we see that, considered qualitatively, the cells
of yeast resemble other cells which enter into the composition of larger plants, and of fungi in general; they
have envelopes formed of cellulose in different degrees
of evolution; and their contents are composed principally of an albuminous protoplasm, of hydrocarbons




70                 ON FERMENTATION.
analogous to gum, with fatty and even resinous substances.  Indeed, yeast contains a small quantity  of
bitter resin soluble in alkalis.
When examined quantitatively, it presents a special
character, and is very rich in nitrogen, far more so than
the vegetable tissues in general.  Thus in fungi, Messrs.
Schlossber-ger and Doepping found in Ioo grammes of
dry matter the following quantities of nitrogen:CANTHARELLUS         RUSSULA.      LACTARIUS
(CHANTARELLE).                     DELICIOSUS.
gram.              granm.       gram.
Nitrogen, per cent.. 3'22... 425.. 468
BOLETUS EDULIS.   AGARICUS CAMPESTRIS.
gram.                grain.
Nitrogen, per cent.. 4'7... 7'26
The composition of various fungi, according to Payen,
is the following:CULTIVATED  MORELLS.  WVIHITE     BLACK
MUSHROOMS.           TRUFFLES. TRUFFLES.
Water... 9IOI.  9g0       72'34. 72
Nitrogenous  compounds, with a
trace of sulphur. 4'68     4'4    ~ 996       o  8-76
Fatty matter.     o'40. o056. 0'44.  o56
Cellulose, dextrin,
sugars,  tertiary
matter..  3'45. 368. I516I               759
Salts, Alkalne, calcic,   magnesic,
silicic phosphates
and chlorides.. 046.'36. 2'Io.  207
Nitrogen per cent.
of the dry matter. 7'33     -. 1'532.'35




ACTUAL COMPOSITION OF FERMENTS.          71
Cultivated mushrooms are, therefore, almost as rich in
albuminoid matter as yeast, since IOO parts of dry
matter contain 52 parts of these substances. Yeast
contains 6o.




72              ON FERIMENTATION.
CHAPTER V.
FUNCTIONS OF YEAST.
YEAST is a living organism belonging to the family of
fungi, genus Saccuauron.yces, destitute of mycelium,
capable of reproduction, like all the elementary fungi,
by buds and spores; its composition, as we have just
seen, singularly resembles that of other vegetable tissues,
and especially of the plants of the same family. The
examination of its biological functions, studied more
particularly in their chemical aspect, shows us clearly
that this elementary form of life does not differ in
essentials from other elementary cells, unprovided with
chlorophyll, whether isolated or in groups, and belonging to the more complex organs. It breathes, transforms and modifies its proximate principles in a
continuous manner, and certainly in the same way as
other cells; like these, it can be multiplied by buds and
spores. The only important and decidedly distinctive
character which seems to render it a form of life
absolutely apart from  other forms in creation, was
removed from  it by M. Lechartier and M. Bellamy,
when these chemists succeeded in establishing that the
cells of fruits, seeds, and leaves, and even animal cells,
are capable of changing sugar into alcohol and carbon
dioxide.
From this time, therefore, the accurate study of the




FUNCTIONS OF' YEAST.              73
biological functions of yeast no longer appears to us
as an isolated chapter in the midst of those which compose general physiology, but, on the contrary, as a particular instance, from  which we may draw important
conclusions as to the chemical phenomena of living
organisms in general. Yeast offers this immense advantage to the observer, that it allows him  to make all
kinds of experiments on it with great facility. It is like
clay, which can be moulded at will, composed of one
and the same kind of elementary cells, which enables
us to avoid the complications due to the intervention of
complicated phenomena.
NAormal Conlditions of the L ife of Yeast.-The condition
which we shall call normal in the life-history of yeast
are those in which this form of life develops itself, and
increases with the greatest activity and energy. They
are of two orders, physical and chemical.
With respect to physical conditions, we have only to
notice the temperature. The temperature most favourable to the nutrition of yeast is also that which is found
advantageous to other cellular vegetable organs; between
250 C. and 35~ C. (77~ and 95~ F.). Above and below
these limits, the vital manifestations do not cease until
we descend below 9~ C. (49'6~ F.) or rise above 60~ C.
(i400 F.), the temperature at which albuminoid principles
begin to coagulate.
As to the chemical conditions, and the most favourable composition of the medium in which this organism
is to live and multiply, they are not so simple.
We owe to the learned and patient labours of Pasteur on
this question the best information whichwe possess on this
part of the subject. He has been followed in this interest



7-4             ON FERMENTATION.
ing investigation by some of his pupils (Duclaux, Raulin),
of whose researches we shall have to speak further on.
We can see, d priori, that the most favourable medium
is that which contains the most appropriate nutritive
elements. These elements ought to be those which we
find in the organism of the cell, or, at least, principles
susceptible of allowing the cell to form with them by
synthesis, its immediate component parts. Thus we
have seen that yeast contains water in greater or less
proportion, mineral salts, especially potassium, magnesium, and calcium phosphates. Water and the alkaline
and alkaline-earthy phosphates will therefore necessarily
form an integral part of the nutritive medium. We
find, besides, a great proportion of nitrogenous substances, either albuminous or otherwise. The food of
yeast ought, therefore, to include nitrogen. A question
then presents itself: Under what form ought nitrogen
to be supplied to the cell in order to be assimilated?
Experiment alone can reply.
We know, by the elegant researches of M. Boussingault, that the higher plants are not able to absorb the
free nitrogen of the atmosphere. This observer caused
a certain known weight of seeds, the average amount of
nitrogen in which he had previously ascertained, to
germinate in a calcined silicious soil, and watered them
with pure water. When the plant, in spite of these
unfavourable conditions, had attained a certain development, he ascertained the total quantity of nitrogen contained in the whole. The weight of this did not exceed
the original quantity contained in the seed. Yeast forms
no exception to this rule.
Assimilation of the Nitrogezn of Nitrates.-Agricultural




FUNCTIONS OF YEAST.                     75
experiments made on a large scale, and repeated for
successive years, as well as those made in chemical
laboratories, have established the efficacy of nitrates,
and salts of ammonium, among the nitrogenous aliments
utilized by plants in general.
The importance of saltpetre as a manure, has been
long known; however, with respect to biological phenomena, it is a question whether the nitrate introduced
into the soil does not, before it reaches the plant which
makes use of nitrogen, undergo changes which reduce
it to another form, that, for instance, of ammoniacal salts.
M. Boussingault has given a complete solution to
this question by experimenting on plants sown in a
medium or a soil absolutely deprived of organic matter,
and in which the reduction of nitrates is impossible.
The following table gives an idea of the influence of
nitrates on vegetation in a barren soil:Acquired by the plants
during 86 days
of vegetation.
Weight of the
dry produce, Vegetahle  Caibon dioxide  Carbon.  Nitrogen.
the seed   matter   decomposed in
being x.  elaborated.    24 hours.
grammes.  grammes.  gramrmes,
When the soil
had received nothing...    3'6. o'285 ~       245.    o'I44. 00023
When the soil
had received
phosphates,
ashes, and potassium nitrate. I98'3. 2I'IIl. I82oo. 8'446. o'I666
When the soil
had received
phosphates,
ashes, and potassiumbicarbonate.  46. o'39.  3'42     o' 56.  0'027




76              ON FERMENTATION.
These numbers show such important differences between the results obtained in the same time, with or
without the use of nitrates, all things besides being
equal, that an error of interpretation is impossible.
We may then say: In the larger plants the soluble
nitrate penetrates into the organism in a state of solution, and there it undergoes changes which finally
bring out its nitrogen, under the form of albuminoid
matter.
Is it the same with yeast? Can it elaborate its proteic matter by decomposing nitrates?
This question has been discussed.  On one side,
Dubrunfant (Comp. Rend. de l'Acad;, vol. 73, pp. 200,
263) says that he has observed a greater activity as a
ferment after the addition of potassium nitrate.
On the other side, Ad. Mayer (Lehrbuch  der
Gahrungs-Chemie, I874) affirms that he has obtained
only negative results in a whole course of researches,
directed to this end. Schaer arrived at the same results
as Mayer.
This decided difference between the phenomena of
the nutrition of yeast and of larger plants is still
more remarkable, since simple organisms, much allied
to the Sacc/zarozyces, act upon nitrates exactly in the
same way as terrestial plants of a higher order.
Thus, the mildews which vegetate on the surface of
liquids derive considerable nourishment from the dissolved nitrate.
Admitting the observations of Mayer to be correct,
we are compelled to interpret these negative results in
the following manner:In order to assimilate the nitrogen of a nitrate, the




FUNCTIONS OF YEAST.               77
plant must first reduce it. Therefore, either the cell of
the Sacch/aroznyces does not possess this reducing power
over the nitrates which we find in other isolated cells,
forming an integral part of more complex organisms,
or else the experiments have been made under conditions in which this reducing power has not been able
to manifest itself. However this may be, we have not
arrived at a final decision respecting the assimilation
of the nitrogen of nitrates; and before we determine
definitely in the negative it will be advisable to vary the
experiments.
Assimilation of the Nitrogen of Ammoniacal Salts.Agriculturists generally agree in recognizing the efficacy
of ammoniacal salts in vegetation. The experiments
of Sir H. Davy, Kuhlmann, J. Pierre, Lawes, and
Gilbert lead us to the same opinion. It appears probable, according to all the facts, that ammoniacal salts
may concur in the nutrition of plants, although it is
recognized that, with an equal weight of nitrogen, they
act in a less favourable manner than the nitrates. The
experiments of M. Bouchardat (Memoire sur lInfluence
des Composes Ammoniacaux sur la Vegetation), those of
M. Cloeiz (Leqons de la Soc. Chim., 186I, p. I67) tend,
on the contrary, to establish, Ist, that the solutions of
ammoniacal salts, usually employed, do not supply to
vegetables the nitrogen which they assimilate; 2ndly,
that if solutions at the rate of l   and even of l     of
these salts are absorbed by roots, they act as energetic
poisons, and kill the plant rapidly. M. Cloez cites, on
this subject, many instances, which leave no doubt as to
the reality of the fact.
If this be the case, we must, in order to make these




78              ON FERMENTATION.
apparently discordant results agree, either admit that
the ammonia, in order to be absorbed usefully, and without danger to the plant, ought to be presented to it
under a special form, perhaps very diluted, or in a
peculiar state of combination; or else it must previously undergo, in the soil itself, a transformation into a
nitrate.  This interpretation, due to M. Cloez, and
which is the reverse of the ideas of M. Kuhlmann (for
this observer maintains that, on the contrary, the
nitrates employed as manures are only active if they
are transformed, in the soil, into ammoniacal salts), is
by no means contradictory to what we know of the
phenomena of nitrification.
Let us return from the higher forms of plants to
ferments, as we have already done before. M. Pasteur
was the first to study the influence of ammoniacal salts
on the development and nutrition of the Sacckaronyces.e
After having ascertained, by experiments made on a
large scale in fermentation, carried on commercially, that
the ammonia of the ammoniacal salts contained in the
juices employed disappeared during the fermentation,
without disengaging any sensible quantity of nitrogen,
he adopted the following experiment, which may serve
as a type for all the rest.
In a solution of pure sugar-candy (we shall presently
see that sugar and its analogues are the food necessary for
yeast) we place, first, an ammoniacal salt-for instance,
some ammonium tartrate-and then the mineral matter
which enters into the composition of yeast, and add to
this a slight, we may say imponderable, quantity of
globules of fresh yeast. The globules sown under these




FUNCTIONS OF YEAST.              79
conditions are developed, multiply, and excite fermentation of the sugar, while the mineral matter dissolves by
slow degrees, and the ammonia disappears. In other
words, the ammonia is transformed into the complex
albuminoid matter which enters into the composition of
the yeast, whilst the phosphates supply to the new
globules their mineral principles: as for the carbon, it
is evidently furnished by the sugar.
This, for example, is the composition of one of the
liquids employed,IO grammes of pure sugar-candy,
Ashes of one gramme of yeast, obtained by means
of a cupel-furnace,
o'I gramme of ammonium dextro-tartrate,
Traces of washed beer-yeast, of about the size of a
pin's head, in a fresh condition, damp, losing 80o per cent.
of water at I00o C. (212~ F.)
In such a mixture, the vessel being filled up to the
neck, and well stopped, or furnished with a gas tube
dipping into pure water, the fermentation began. After
from twenty-four to thirty-six hours the liquor began to
give evident signs of fermentation, by a disengagement
of microscopic bubbles, which announced that the liquid
was already saturated with carbon dioxide. On the
following days the troubling of the liquor increased
progressively, as well as the disengagement of gas,
which was considerable enough for the froth to fill the
neck of the flask. A deposit began, by degrees, to
form at the bottom of the vessel. A drop of this
deposit, examined under the microscope, shows a beautiful example of yeast, very much ramified, extremely
young in appearance; that is to say, the globules are




So             ON FERMENTATION.
swollen, translucent, not granulated, and we distinguish
among them, with surprising facility, each globule of
the small quantity of yeast sown at the commencement
of the experiment. These latter globules have a thick
envelope, defined by a darker circle; their contents are
yellowish; they are granular; but the manner in which
they are sometimes surrounded by the newly formed
globules shows very clearly that they have given rise to
those among the latter which form the first links of the
chaplets. If the observations are made in the earlier
days of the formation, during the evening, and by gaslight, the old globules'are distinguishable among the
infinitely more numerous young ones, as you would distinguish a black ball in the midst of a great number of
white ones.
This fundamental experiment proves, then, that yeast
can bud, multiply, live and feed in a medium in which
nitrogen is only represented by ammoniacal salts.
Further, M. Pasteur draws this conclusion from it
that yeast causes with ammonia the synthesis of albuminoid substances.  In order to test this conclusion,
M. Pasteur weighed the ammonia which remained in the
liquid some weeks afterwards, and found it less —slightly
so, it is true ('0062 grammes). The weight of the newly
formed yeast rose to'o43 grammes.
In his last memoir on the origin of muscular force,
(Ann. Chim. Phys. [4], vol. 23, p. 5, I87I), Liebig energetically attacks the conclusions and results announced
by Pasteur.  He absolutely denies the formation of
yeast, and its increase in weight, under the conditions
of Pasteur's experiments, saying that he had never
succeeded in realizing it; while, by substituting for the




FUNCTIONS OF YEAST.              8I
ammoniacal salts water in which yeast had been washed,
the formation of fresh yeast was very evident. One of
the arguments of Liebig, on which he relies most, is the
absence of sulphur in the nutritive medium used by
Pasteur; the albuminous substances contain some, and,
therefore, yeast cannot elaborate proteinic matter under
these conditions.
Let us remark, on this head, that sulphur occupies
but little space in the complex molecule of albumin,
and that nothing proves that this body is indispensable
to the constitution of albumin.
In reply to this attack, M. Pasteur could only repeat,
with forcible conviction, his former affirmations, and
propose to his opponent to have the facts investigated
by scientific referees. We will dwell no longer on this
dispute, which was unfortunately terminated by the
death of one of the most distinguished chemists of our
age. We will only remark that fresh facts, studied with
great attention, especially by M. Raulin, have, by
analogy, given complete comfirmation to M. Pasteur's
opinions concerning the nutrition of simple organisms
in general, and yeast in particular. We shall allude to
these facts later on.
M. Pasteur's experiment, as described by him in his
memoir on alcoholic fermentation (Ann. de Chimie et
de Phys., vol. 58 [3], P. 390), may be liable to this
criticism-that, in proportion to' the small quantities of
yeast at first employed, the weight of ammonia that
had disappeared, and that of the newly formed yeast,
are very minute; and that they might be considered
as falling within the limits of experimental errors, if one
did not know the great ability of this eminent observer.




8 3              ON FERMENTATION.
These very small initial quantities of yeast were
employed to avoid the suspicion that the nourishment
f the new cells had been effected at the expense of the
soluble principles excreted from the old ones (the water
in which yeast has been washed is, in fact, very effectual
in giving activity to the multiplication of the cells of
Saccharomyces).  The experiments made by Duclaux
in a different manner prove that these objections have
no real foundation, and  that the ammonia of the
medium actually disappears.
This skilful chemist introduced into a certain volume
of a solution of sugar, 2'50I grammes of yeast, containing
0o215 grammes of nitrogen; the liquid also contained
I gramme of ammonium tartrate, corresponding to o' I52
grammes of ammonia:  Af'ter the fermentation, 2'326
grammes of yeast, containing 0'I48 grammes of nitrogen,
were obtained from  it. The liquid contained 0o055
grammes of ammonia, and 0'170 grammes of nitrogen
in the form of organic compounds; which gives the
following balance of nitrogen:Nit. before     Nit. after
fermentation.  fermentation.
In the yeast...  o215   a.    o'I48
In the composition of ammonia.. o'I52,. 0045
Under the form of nitrogenous
organic matter, dissolved in
the liquid....a  0o'20
Total  0..0'367              o'363
The two  amounts agree, within  an  error of 4
milligrammes (o06 grains Eng.). We see clearly that




FUNCTIONS OF YEAST.                     83
three-fourths of the  ammonia  has disappeared, and
we find it in the yeast and  the surrounding liquid,
under the form of nitrogenous organic combinations.
This experiment having since received the confirmation
of every well-observed fact, we may admit, with confidence, that yeast can effect the synthesis of its proteinic
materials, at the expense of sugar and ammonia.
M. Mayer has proved, as  a  complement of the
experiments of M. Pasteur and M. Duclaux, that the
ammonium tartrate may be replaced by other ammoniacal salts, as  the  nitrate  oxalate,  &c.,*  without
any disadvantage to the nutrition and to the disappearance of ammonia; and thus, far from being decomposed, in order to supply ammonia during its
development, and during the fermentation, as D6bereiner
had asserted, the yeast consumes the ammonia contained
in the liquids which are fermenting.
Although a salt of ammonia may serve for the
nutrition and development of ferment, it is proved,
however, by generally observed facts, that it is not the
especial nitrogenous aliment for this simple organism.
If we substitute for the ammoniacal salt in Pasteur's
experiment natural juices (as that of grapes, or of
beetroot, or the water in which yeast has been washed),
containing nitrogenous organic substances, the quantity
of yeast formed and decomposed in the same time
* In all the observations made by Pasteur and the other experimentalists,
an increase or decrease of the rate of development of the cells, their multiplication and their nutrition, was always accompanied by a variation in some
direction in the energy with which the sugar (one of the essential nutritive
elements) was resolved into alcohol and carbon dioxide. We will discuss
presently what explanation may be given of the correlation between the two
phenomena.




84             ON FERMENTATION.
is much greater, and the decomposition of sugar is
more active.
There exist, therefore, nitrogenous carburetted substances which are better suited to the nutrition of
yeast than ammonia. What are they?
The natural juices which we have just mentioned,
and more especially yeast-water which shows itself to
be particularly active, contain different kinds of nitrogenous matter, and particularly albuminoid substances.
Direct experiments alone can determine whether we are
to attribute the active part played by these juices to
proteinic principles, or to more simple compounds.
Pasteur found that the albumin of the white of egg
was entirely unfit for the nourishment of the globules
of yeast. Even at an earlier period M. Thenard and
M. Colin had observed that albumin does not begin
to excite alcoholic fermentation (or, which is the same
thing, to produce the nutrition and development of
the yeast globules) till the end of three weeks or a
month, when left at the temperature of 30o C. (86~ Fahr.),
when it undergoes, under the influence of infusoria and
mucidines, which are developed in it, a greater or
less degree of decomposition. The serum of blood
encourages the nutrition, of the globules, without requiring to be itself previously decomposed; but it is
not the serine which is active in this case; for, if
we eleminate it by coagulation, the boiled and filtered
liquid, when sugar and yeast are added, rapidly
produces an energetic fermentation (Pasteur).
M. Mayer has made many experiments, with the
view'of throwing light on the question of the nutritive
part played by albuminoid substances. He has found




FUNCTIONS OF YEAST.                85
that the inactivity of the greater part of them (albumin,
casein, &c.) arises especially from their not being
diffusible  through the organized membranes of the
cells. We know, in fact, that most of these bodies
belong to the class of colloid substances, not diffusible
through porous membranes, and we can easily understand that, not being able to penetrate into the interior
of the cell, but remaining imprisoned in the surrounding
liquid, they will not be in a condition to give any
powerful aid to the development, the nutrition, and
the  multiplication  of the  globules.  The  diffusible
products formed by stomachic, intestinal, or artificial
digestion, show themselves to be eminently suited to
nourish the cell of the Sacczlaromzyces.
It is the same with regard to diastase, and the
different kinds of pepsin; but, as the nutritive activity
of these ferments remain after they are cooked, it
must by no means be attributed to their specific
properties as soluble ferments (for these properties
are destroyed by heat), but rather to products analogous
to peptone, which  always accompany these soluble
ferments, and of which it is very difficult to get rid
-syntonin, and, in  a less degree, allantoin, urea
guanine, and uric acid, increase the fermenting
power of yeast; that is to say, give nourishment to
the organism.
Other nitrogenous substances, which we ought also
to  consider as compound  ammonias, have  shown
themselves to be but slightly, or not at all, active.
Such are creatin, creatinine, caffeine, asparagin, leucine,
hydroxylamine.
It is, then, very probable, according to these results,




86             ON FERMENTATION,
that natural juices, the wort of beer, and water in
which fresh }/east has been washed, owe their power
of nourishing the cells of yeast, not to albuminoid
principles, properly'so called, which are indiffusible,
but to allied nitrogenous compounds analogous to peptones, which have the'property of passing by osmose
through membranes.
Yeast thus affords us an evident and striking example
of vegetable cells, which assimilate their nitrogen under
the form of complex combinations, allied by their constitution to the higher forms of albuminoid substances.
There is no proof that similar phenomena are not produced in plants of higher organization.
Agricultural and physiological experiments do not
establish so clearly the assimilation of nitrogenous
organic combinations as that of nitrates and ammoniaca! salts; although the few observations made on
this subject are rather favourable than otherwise to
a positive solution of the question. But, if it were
otherwise, it would not be logical and prudent to seek
to draw a sharp distinction between the phenomena
of nutrition of the Saccharomyces, and those of larger
plants, and to say that the former may find nitrogenous
nourishment in the organic nitrogenous combinations
allied to albuminous substances, and that the latter do
not.
Plants of complex organization are, in fact, formed
by the union of cellular elements of various kinds,
fulfilling different functions, and whose conditions of
nutrition and development are not identical; among
which it is probable that some  are to be found
susceptible of assimilating complex nitrogenous or



FUNCTIONS OF YEAST.                 87
ganic materials, that have been elaborated elsewhere
at the expense of ammoniacal salts, or of nitrates.
Assimilation of Minizeral Princziles.- Vegetables, in
general, always leave, after combustion in air or in
oxygen, a fixed mineral residuum, whose weight varies,
within certain limits, from  one vegetable species to
another, and also, especially, from one organ to another,
as well as for the same organ, according to its age.
Thus, the leaves of the pear-tree yielded to M. Violette,
7' I 8 grammes of ash per cent. of dried matter:
The extremity of the twigs  bark.  o    3454
twood     o        0'304
(bark.. 3'682
The middle part.       bawood             3 6834
wood.           o'I34
The lower part        [bark.             2 903
iwood. 0'354
The trunk             Ibark. 2'657'wood. 0o296
The roots              bark.             I127''wood              0. 0'234
In proportion as the plant grows older, the weight of
the ash increases.
Do, then, mineral principles, which are found in all
vegetables, from the highest point of the scale to the
lowest-for we have already seen, by analysis, that yeast
forms no exception —do, then, mineral principles play
an important part in the biological phenomena of the
nutrition  and  development of the plant, or do they
only make their appearances as useless, but not
injurious elements, inevitably introduced by the fluids
from which the plant derives its nutritive principles?




88                 ON FERMENTATION.
The remarkable constancy of the chemical composition of various kinds of ash, especially with respect to
their constituent elements, as well as the most complete
agricultural experiments, have proved in the most positive manner that the greater part of saline compounds,
found by analysis, are  necessary to vegetation.   We
have also learned to classify them  in the order of their
nutritive importance, with reference to the entire plant
and its different constituent parts (leaves, stalks, seeds,
grain, &c.).
Thus the phosphates preponderate remarkably in
grain, and form by themselves the whole of the mineral
mass found after incineration, as shown in the following
results, published by Berthier:PHIOSPHATES IN  100 PAITS OF ASH.
Nature of Phosphate.   ct
Potassium phosphate   5'oo 48'50o 52'50  7'50 240 41'50 42o70 6670o 6170
Calcium,,      2200 29'20 15'00 i6'50 24'10 i8'5o  8'40 22'20 6'50
Magnesium,,     2800  -  2500 2000 240 3800 14'30 6 60 i9'60
Manganese,,           8'30  -   -   -
Total...    I oo'oo 96'00oo 92'50 44'00 72'3o0 65'50 98'09 95'50 87'80
We also give, for reference, from  the same author,
the analyses of the ashes of various parts of plants.




FUNCTIONS OF YEAST.                             89
STALKS, I00 PARTS OF AsEI CONTAINComposition of the Ashes.   " 0 
Potash,......... 340
Potassium  and sodium car-a    16'40             - - -
bonates...           I'40      -       -        444    1220
Potassium chloride.            220    0'78    2'90o    1'90       3'64,,    sulpate....  4440      3'40    0 30    a266    I*30,,    phosphate..             - 
silicate,.4'00
Lime......                    15'70
Calcium carbonate...       49'82    6'co     -       64'26   22'62
Magnesium  3,,                  385      -       -        607     6 39
Carbon dioxide     I,     *  I 00                         - 
Iron oxide......        -        -       2'60     -
Calcium phosphate...          570     660      ~oo      843    1131
Magnesium,,               I                             -'
Iron                          I83       -        -        -
Manganese,,
Phosphoric acid...          -        -       I-20     -
Silica......    580o   78'22    73'90       2'24   39'80
BULBS AND ROOTS GAVE, FOR I00 PARTS ASIIMadder   Jerusalem    Potatoes.   0ni0n1s
0 Madder  Jerusalem
Root.   Artichoke.  Potatoes..
Potassium  and sodium car-}
bonates......           31         35o        42.43       i6o
Potassium chloride...3...4                7'50       4'00       2'20
Sodium......
Potassium sulphate....    3 93       6 oo       2'80       4o00
phosphate...                30         34'70       -
Calcium carbonate               35-01        -         2'80       200oo
Magnesium                        4r3              _    -         I0.00
Calcium phosphate                 971      I6'50       6'87      38 -oo
Magnesium      -,,....  8'5         2'50
Iron,,....    5'o9            -          I'70
Silica.........    788                  250.. i~~




co                 ON FERMEN TATION.
In  the leaves, the calcium  carbonate and the silica
preponderate, and form in themselves from 6o to 90 per
cent. of the total weight of ash.
Pilne Leavnes. Vine Lieaves.  M|Leaves.
Calcium carbonate......      68374      51'00      53'00
Silica.6'43                              I0'20      27'70
Total,,.....         75I7      6i'20      80'70
Let us compare with this the composition of the ash
of beer-yeast (Mitscherlich) 
Surface Yeast.  Sedimentary Yeast.
Phosphoric acid.. 4I18         @    39'5
Potassa...      39'8   e 3        28'5
Soda..,.
Magnesium phosphate.. I63..    22'6
Calcium,, ~.  2-3..     9'7
If mineral salts really play an active part in vegetation, we may foresee, from these analyses, —
First. That their relative importance will vary with
the respective weights of these different bodies, found
in organized tissues; that, in consequence, the phosphates, potash, soda, magnesia, and the sulphates will
occupy the first place.
Secondly. That according as the soil or the medium
in which the plants grow is more especially manured by




FUNCTIONS OF YEAST.               91
any one of these salts in particular, the development
either of leaves, stalks, or seeds will be favoured. These
results have all been verified by direct experiment, but
as it does not enter into our plan to study agricultural
chemistry with reference to mineral manures, and as
our intention is merely to compare the nutrition of
yeast, and of analogous organisms, with that of other
plants of a higher order, we will return to the history of
the Sacc/aromzyces. We would remark that the composition of its ash, entirely composed of phosphates,
approaches more nearly to that of seeds, to which it
is also allied by the analogy of function and general
chemical composition.
We owe to M. Pasteur the proof of the absolute
necessity of mineral salts (phosphates of the alkalis,
and the alkaline earths) for the development and
nutrition of the yeast-cell.  If, in his experiment, in
which the ferment is sown, in an imponderable quantity,
in a medium entirely composed of pure sugar-candy,
ammonium tartrate, and ash of yeast, we omit the
latter element, the fermentation and development of
cells which ought to precede it (i.e., the fermentation),
do not take place. M. Pasteur went no farther in these
researches; absorbed by the pursuit of a different aim,
he did not endeavour to ascertain what were the most
favourable mineral substances; he only used in his researches ash of fresh yeast as an inorganic element,
rightly thinking that, at all events, he should find in it
that which agrees best with the mineral nutrition of the
fungus.
M. Mayer, following up this work, and seeking to
ascertain by direct experiment which among the salts




92             ON FERMENTATION.
generally contained in varying proportions in vegetable
ashes clearly favour the development of yeast, arrived
(loc. cit.) at the following conclusions:I. Preparations of iron, employed in very small quantities, seem to have no influence; in larger proportions
they are injurious.
2. Potassium phosphate shows a preponderating
favourable influence. It may be employed in a liquid
medium, in a high percentage, without its fertilizing
influence being destroyed; while, for plants of a higher
order, so great a concentration would become a serious
cause of pathological disturbance.  Potassium phosphate is not only favourable, but indispensable.
In fact, if from a medium formed of sugar-candy,
ammonium nitrate, traces of yeast, and a mixture of
acid potassium phosphate, magnesium sulphate, and
tricalcic phosphate, a medium which ferments with
considerable activity, we omit the acid potassium
phosphate, fermentation and the development of yeast
are not produced.
Potassium phosphate can by no means be replaced by
sodium phosphate, which is inactive.
The absence of calcium phosphate from the medium
causes much less injurious consequences than the
omission of the before-mentioned salt.
The result is, that potassium and phosphoric acid are
indispensable elements, whilst lime may be omitted
without any great inconvenience, as we might have foreseen from the results of the analysis.
Magnesium, on the contrary, appeared in Mayer's
experiments to be a very useful, if not an indispensable,
element. It is immaterial ivhether this metal is supplied




FUNCTIONS OF YEAST.               93
under the form of sulphate, or of ammoniaco-magnesian
phosphate.
The combinations of sodium present no material
effects, conformably with what has been already observed in plants of a higher order.
The sulphur, administered to yeast under the form of
sulphates, or soluble sulphites, appears not to be assimilated. At least, the presence or absence of these two
classes of salts seems to have no influence.  Yet yeast
contains sulphur in appreciable proportions, which we
even find combined intimately in the products of its
dis-assimilation (sulphuretted pseudo-leucine of Heintz).
We are unable to say what the origin of this normal
sulphur may be.
M. Raulin, in a remarkable investigation, has studied
with particular care, and by an excellent method, the
influence of the mineral components of the medium on
the development of a cellular plant, the Aspergillzs nizger.
As the results obtained may be interesting with respect
to the question, rather as a general one than specially
relating to yeast, which we are now considering, we will
enter into some details on this point, more especially
since the experimental method employed by M. Raulin
may perhaps serve as a model for other researches of
this kind.
First of all, an artificial medium is prepared, exclusively formed of definite chemical compounds suitable
for the vegetation of a particular plant. In order to study
the influence of various physical or chemical circumstances on the development of this plant, a vessel is
filled with the artificial mixture and placed under the
most favourable conditions for its vegetation. The seeds




94              ON FERMENTATION.
of the plant are sown in it, and they are allowed to grow
during the necessary time; this trial, which is reproduced exactly in the same manner in each series of
experiments, is the typical trial, with which all others
are compared.
Another experiment is arranged in every respect like
the first, with the exception of the single circumstance
which it is proposed to study. The two crops, obtained
at the same time, are dried and weighed separately, and
the numerical ratio of the weight of these two results
will be the measure of the influence of the condition to
be examined.
The degree of perfection of the method depends upon
three general conditions:
I. It is absolutely necessary to find, in the first place,
an artificial medium suited to the development of the
plant to be studied. M. Raulin found the ground quite
prepared in this respect, thanks to the labours of
M. Pasteur: the latter had observed that the mucidines
(Penicilfiumzn) can be developed in a medium exclusively
formed of definite artificial substances.
Water, sugar, ammoniacal salt (bitartrate), and ash
of yeast. No portion whatever of the constituent parts
of this medium can be omitted, without giving a complete check to the development.
2. The weight of the crop which the medium intended
for the typical experiments can yield in a given time,
with a constant weight of nutritive substances, ought, all
other things being equal, to be as great as possible.
3. The typical experiments placed under the same
conditions ought to yield crops whose numerical ratios
differ but little from that assumed as unity: the ratio




FUNCTIONS OF YEAST.                   95
which differs the most fixes the relative maximum error
of the process.
At the beginning of his researches, M. Raulin, relying on the data of M. Pasteur, made use of a typical
medium composed ofWater   e.        *   a. 2,000
Sugar    e.             70
Ammonium  nitrate,               3
Tartaric acid...    2
Ammonium phosphate..
Potassium carbonate.'      small
Calcium,,.           quantities.
Magnesium,,              
Aspergillus sowed at 20~ C. (68~ Fahr.).
With such a medium, the variation in weight of the
crop, between one typical experiment and another, was
so considerable, that it was not possible to ascertain the
influence exercised by the omission of certain elements,
which entered into the mixture only in small proportions.: Thus, after forty-eight hours' vegetation, the
weight of two typical crops were found equal toGra m es,                Grammes.
No...   319.         No. 2.      I'77.
By the omission of all the mineral elements he arrived
at the resultGrammes.                Grammes.
No. I..    o'Io.     No. 2.    0'-87.
The omission of potassium carbonate alone gaveGramlnes.               Grammes.
No. I...    2'29.       No. 2            III.




96               ON FERMENTATION.
The action of the whole of the mineral salts comes
out strongly; that of the potassium  carbonate is not
perceptible, for the number 2-29 lies between 3'I9 and
I'I77, the numbers found for the typical experiments.
Besides, in these first experiments, M. Raulin ascertained that the development of the mucidines was
fairly rapid in the first few days, and then grew indefinitely slower. While seeking to find the causes of this
disturbance by tentatively modifying the conditions of
the medium, especially by adding to it sulphur, zinc,
iron, and silicon, in the form of salts; by modifying the
proportions of the essential elements, raising the temperature to 35~ C. (950 F.); and, finally, by employing
vessels of considerable area and small depth, he succeeded in finding a typical medium, giving for the same
length of time a result fifty times greater than that of
the first experiments. Under these conditions, the ratio
of the typical experiments, instead of varying from I'o
to i'8, acquired a remarkable constancy, and did not
vary now more than 2-I4 of its value.  It is evident that
the favourable influence of any particular substance will
then show itself in a much more defined manner.
The experiments, as far as Aspergillus niger is concerned, must now be conducted in the following
manner:In the vessel intended for the typical experiment, the
following chemical substances are brought together:W\Tater. 1,500
Sugar-candy..               70
Tartaric acid..                4
Ammonium nitrate'.           4
9)   phosphate..   o60o




FUNCTIONS OF YEAST.                97
Potassiuml carbonate,.mu  o'6o
Magnesium,,...    o40
Ammonium sulphate...   025
Zinc       0,..      07
Iron,,               0-07
Potassium silicate.0  o'07
This mixture is left to itself for several hours, and
then stirred with a porcelain spatula.
In order to sow the fungus, it is sufficient to pass over
the whole surface the end of a camels'-hair brush with
which spores have been collected from a very pure, and
not too dry, vegetation of Aspergfillus.
When we are not yet in possession of any Aspergfillzs,
it is sufficient, in order to procure this plant in a pure
state, to leave exposed to the air certain natural substances, such as the water of acidulated yeast, damp
bread, or slices of lemon.  The spores of Aspergillus
which exist among the germs in the atmosphere may
fall on these matters, and develop themselves there,
mixed with other organisms.
When we see Aspergillus make its appearance, which
is immediately distinguishable by its black fructification,
it is again sown on an artificial liquid, and we at last
obtain it free from mixture. The typical experiment
being thus prepared, it is placed in a stove at 35~ C.
(95~ F.), and constantly supplied with damp air. The
spores develop, and at the end of twenty-four hours the
filaments of the mycelium  form  a continuous whitish
membrane on the surface of the liquid  At the end of
forty-eight hours this membrane has become very thick,
and turns to a deep brown colour; after three days it has
become quite black on the upper surface, which colour is




98               ON FERMENTATION.
due to the appearance of spores. The thick membrane
is then removed by the fingers, squeezed, and then spread
upon a plate to dry. New spores are sown upon the
liquid, and after three days we obtain a second crop,
weaker than the first.
The typical mixture, and that which is to serve for
the experiment, and which differ from each other only
in the single element, the influence of which we wish to
ascertain, are placed together in the stove. This influence is measured by the ratio of the weights of the
two first crops, or, still better, by the amount of the
first and second crops obtained in six days.
E.-aiple. — The following trial mixtures were placed
in the stove:No. I. Typical medium.
No. 2.,,,,       less the potass.
No. i.   No..    No. 2
Grammes. Grammes.
First crop (after 3 days).    14'4    oSo
Second crop (after 3 other days)   Io'o    O'I2
Total 24'4   0o92
Ratio of the two first crops l-= —i-; that of the
weights of the whole crops 942= 2, numbers which prove
plainly, in the most complete manner, the utility of the
potass.
The results obtained by this remarkable method are
the following:
I. All the elements of the typical artificial medium
concur simultaneously in the development of the plant,
for if we omit each of them in turn, the weight of the
crop undergoes a diminution, which is usually somewhat




FUNCTIONS OF YEAST.                  99
considerable, and which cannot be attributed to experimental errors.
2. The mineral oxides of the artificial medium cannot
be substituted for each other.
3. Nitric acid may be used instead of the ammoniacal
salts as the nitrogenous aliment.
Finally, the following are the ratios -found between
the typical and the experimental trials:Omission of the oxygen..    very great,,,,,, water.         infinite,,,,,, sugar                       65
I,,, tartaric acid.         infinite,, ammoniacal salt, or nitrate. I53,,, phosphoric acid. I82,,,, magnesia..      9,, potash.....  25, sulphuric acid.  24
zinc oxide                      10
9,,,iron oxide..            2-7
99silica                             I.. I'4
The nutritive elements of the artificial medium  are
some indispensable, which are those found in large proportions in it, and others are useful, but apparently not
indispensable: these only enter into the composition of
the typical medium in very small proportions.
It is probable that some of these, such as sulphur,
exist accidentally in very small quantities in the artificial
media, to which they have not been added, and may
thus set up a sluggish development.   Thus, Mayer
asserts that he has been.unable by repeated crystallizations, and even by precipitating the liquor by barium
chloride, to obtain sugar free from  sulphur; it is to the




100            ON FERMENTATION.
presence of this sulphur that he attributes the introduction of this element into the newly formed yeast.
It is evident that the results obtained by M. Raulin,
and especially his method, may be applied to the search
after better conditions for the maximum development
of other kinds of vegetation, or simple organisms. M.
Pasteur, who in his laboratory at the "Ecole normal"
discovered new methods for the production of pure beeryeast on a large scale, had to make preparatory trials
analogous to those of M. Raulin. His researches also
establish an important fact, that an artificial medium,
suitably prepared, may be as favourable to the development of vegetation, and even more favourable, than
the most fertile natural media. We may thence draw
conclusions of great importance as to the cultivation of
larger plants, and may suppose that chemical manures,
suitably chosen, may be substituted for natural ones in
agriculture, with great advantage.
This is what several men of science, who make a
study of agriculture, have already attempted; the great
point is to determine carefully the useful composition of
these manures; unfortunately, we must admit, experiments on larger plants are not so simple and so easily
managed as those on mucidines.
Sufgar.- Pasteur and Raulin have demonstrated. the
preponderating influence of, and'the necessary part
played by, sugar or analogous bodies in the vegetation
of AspcrgiliD s and of mucidines.  This influence is as
powerful in the development of the yeast of beer.
Without sugar, without hydrocarbonate substances,
yeast can neither reproduce nor be nourished. An
important difference is thus, at first sight, established




FUNCTIONS OF YEAST,             IOI
between simple organisms, such as ferments, mildews,
&c., and the larger plants, which derive the organic
elements of their constitution from the simplest compounds of carbon, such as carbon dioxide.
This distinction, however, loses its force after a more
complete examination.
If the larger plants derive nourishment at the expense
of carbon dioxide, it is because, in their leaves and other
green parts, there are found organs suited to the utilization of the active force of the luminous rays sent by
the sun or other source of light. The carbon is set free
directly, and the oxygen is disengaged. It appears very
probable, that at the moment when the carbon is separated from the oxygen, under a special condition as
yet unknown, and very different from that of black
amorphous carbon, or of the diamond or graphite (forms
under which we know this element), that it unites with
the elements of water to form a hydrate of carbon
(starch, sugar?), or at least a body which can be converted
into these principles by ulterior transformations.
If we were able to effect the decomposition of carbon
dioxide under the influence of light outside the animal
economy, I have no doubt but that (if the experiment
were made in the presence of water) there would be
found a hydrocarbon compound.  I have even been
able to give a slight experimental confirmation of this
theoretical opinion.
If we treat, in the cold, coarsely powdered white cast
iron (which is known to contain an iron carburet), with
a solution of cupric sulphate, the iron of the white cast
iron is entirely dissolved, without disengagement of any
carbon or other gas; after having washed it, we may




102             ON FERMENTATION.
eliminate the deposited copper by placing it in contact
with a solution of iron perchloride.  The copper is
rapidly dissolved; there remains a pulverulent black
mass, which, after dessication at So0 C. (I76~ Fahr.), in a
vacuum, resembles carbon.  But this carbon contains
water in combination, which is suddenly disengaged when
it is heated to about 2500 C. (480~ Fahr.); it is easily dissolved in nitric acid, becoming oxidated, yielding, yellow
or orange-yellow substances containing nitrogen. This
residuum, when analyzed, gives a quantity of water,
which is in a tolerably constant proportion to that of
carbon.
It therefore represents a true and definite carbon
hydrate. It is evident that the condition of the carbon
in the cast iron must be very different from that of the
carbon of carbon dioxide, and that the hydrates which
arise from the separation of these forms of carbon may
differ greatly. Nevertheless, the experiment which I
have just described gives material support to the idea
which physiologists entertain of the successive chemical
metamorphoses of the carbon compounds in plants.
When the carbon hydrate is once formed in the leaf,
it is carried into the other parts of the plant, to serve
there as nutrition, for the development of cells containing no chlorophyll, and whose biological functions
closely resemble those of cellular organisms.  That
which takes place during the germination of seeds, up
to the moment when the new plant becomes provided
with aerial leaves which have become green under the
influence of light and ail; and begins to utilize carbon
dioxide, leaves no doubt as to the scientific value of this
interpretation.




FUNCTIONS OF YEAST.               103
We see here the newly formed cells successively
developed, and superposed so as to form radicles, stalk,
cotyledons, and leaves, and the germ procures the necessary materials for its development from the organic
principles which the seed has accumulated, and among
which hydrocarbons are always predominant.
We must, therefore, admit that the phenomena of
nutrition of the larger plants do not seem to differ
much, when examined in detail, from those of the more
simple ones.
The former are provided with special organs which
enable them to elaborate for themselves the hydrocarbon substances which they require for the development of the rest of their organism. The  inferior
cellular plants, and in fact generally all those which are
unprovided with cells containing chlorophyll, are necessarily parasites, which must borrow their hydrocarbonate nourishment, directly or indirectly, from plants
furnished with these cells.
Besides these general considerations, founded on the
phenomena of nutrition observed in plants, the experiments of M. Pasteur establish with certainty, that in all
alcoholic fermentation a part of the sugar is fixed in
the yeast, in the state of cellulose or some analogous
body. In fact, since infinitely small quantities of yeast,
sown in a medium entirely formed of pure sugar-candy,
of ammonium tartrate or nitrate (Mayer), and ash of
yeast, develop and give rise to very ponderable proportions of yeast, considerably greater than the original
quantities, it cannot be doubted that the hydrocarbon
principles of this new vegetation  (cellulose, &c.), are
furnished by the elements of the sugar.




104            ON FERMENTATION.
The following experiments lead to the same result:M. Pasteur submitted to fermentation Ioo grammes
of sugar, about 750 cubic centimetres of water, 2'626 of
yeast (weight of the dried matter).
After the fermentation, which lasted twenty days, he
collected 2'965 grammes of yeast (dried matter).
He likewise boiled for six or eight hours a determined
weight of fermented yeast, and also of the same yeast
before fermentation, with sulphuric acid, diluted with
twenty times its weight of water (fermented yeast 1707
grammes, and unfermented yeast I'73 grammes, dried at
a temperature of Io000 C. (2I20 F.).
The insoluble residues were weighed on filters,
the weight of which had been estimated; they were
then washed, dried at Ioo~ C. (212~ F.), and weighed.
The filtered liquids were neutralized with barium
carbonate; the quantity of sugar formed by the action
of the sulphuric'acid on the cellulose was ascertained,
either by means of Fehling's liquid, or by fermentation.
He thus found, by calculating the results obtained for
the two weights, 2'626 grammes, and 2'965 grammes of
yeast employed, and yeast obtainedI. That the 2'626 grammes of crude yeast employed
gave an insoluble nitrogenous residuum equal to 0o'39I
(14-8 per cent.), and 0'532 of fermentable sugar.
2. That the 2'965 grammes of yeast found after fermentation leave a nitrogenized  residuum  of o'634
grammes (about 2I14 per cent.), and o09IS grammes of
fermentable sugar.
There was, therefore, fixed in the fermentation of
Ioo grammes of sugar with 2'626 grammes of yeast,
04 grammes of hydrocarbon matter, transformable, by




FUNCTIONS OF YEAST.               105
dilute sulphuric acid, into fermentable sugar; there
was also a sensible augmentation of nitrogenous matter,
insoluble in dilute sulphuric acid.
On the other hand, in order to verify, by a second
experiment, the value of these conclusions, M. Pasteur
made use of the process of separating cellulose from the
albuminoid substances indicated by Payen and Schlossberger.  This process consists, as is well known, in
treating yeast with dilute solutions of potass.
In three careful experiments, M. Pasteur found a
residuum, insoluble in potass, formed of cellulose, transformable into sugar by being boiled with dilute sulphuric acid of I7'77, 19'29, and I9'21 per cent. of the
dry yeast experimented on.
But the 0'532 grammes of sugar, produced without
the intervention of potass, by 2'626 grammes of the
same yeast, correspond to 20 per cent. of yeast. It is,
therefore, proved that boiling in sulphuric acid had
removed all the cellulose.
Let us also notice that the 2'965 grammes of yeast
found after fermentation, giving og918  grammes of
sugar, ought to contain 3I'9 per cent. of cellulose, a
quantity i I per cent. greater than there was before
fermentation. This considerable augmentation of the
weight of the cellulose in the yeast, while it exercised
its action on the sugar, is a point worthy of remark,
since it proves that, in accomplishing one of its principal
functions, yeast undergoes very marked evolutions in its
composition.
The following experiment of M. Pasteur's proves,
besides, that during fermentation the yeast itself forms
its fatty matter by the help of the elements of sugar.




IoG             ON FERMENTATION.
Let us first call to mind that Payen's analyses show
2 per cent. of fatty matter in the ye'ast, and that the lees
of wine also contain fatty matter. It had been thought
that this fatty matter was furnished by the fermentable
medium.  Pasteur mixed sweetened water (prepared
with pure sugar-candy) with the watery extract of
limpid yeast, heated;. several times with alcohol and
ether. He sowed in it an imponderable quantity of
fresh globules. These multiplied, and caused the sugar
to ferment.  He succeeded thus in preparing some
grammes of yeast from substances completely without
fatty matter. But this newly formed yeast does not
contain less than from I to 2 per cent. of fatty saponifiable matter, yielding crystallized fatty acids.  The
same fact is observed with yeast which has been grown
in a mediumcomposed of sugar, water, ammonia, and
phosphate. It is, therefore, from the elements of the
sugar that the fatty matter is obtained.
These facts confirm  the views of M. Dumas as
to the possible formation of fatty matter from
sugar.
Water.-Water is, we need hardly say, quite as indispensable for yeast, and the elementary organisms, as for
higher forms of life.
According to Wiesner, the cell of yeast manifests its
activity, develops, and is nourished within the limits of
hydratation comprised between 40 and 8o per cent. of
water.  Yeast dried with precaution may regain its
power when moistened afresh. It may be understood
from this why a solution of sugar, the concentration of
which exceeds 35 per cent., is not changed by ferment:
such a solution takes from the cells, by osmose, a suffi



FUNCTIONS OF YEAST.              107
cient quantity of water to lower their hydratation below
40 per cent.
Wiesner's researches have also shown that there are
two states of concentration in which the phenomena of
the fermentation and nutrition of yeast attain their
maximum value. One of these maxima corresponds to
a solution of from 2 to 4 per cent. of sugar; the other
to a solution of from 20 to 25 per cent. These facts
require confirmation; at all events, there is at present
no conclusion to be drawn from them.
Orygenz.-The cells of the Saccharovmyces cerevisic,
introduced into a liquid medium  containing oxygen in
solution (pure water, a saccharine solution, with or
without nutritive mineral and nitrogenous elements),
absorb oxygen with great rapidity, and develop a
corresponding quantity of carbon dioxide. This fact,
which constitutes true respiration, comparable to that of
animals, has-been brought to light by M. Pasteur. An
excellent method of obtaining water completely deoxygenized, much more efficaciously than by boiling,
consists in diffusing through the water one or two
grammes (from I5'4 to 30'8 grains) per litre (I'76 pints
English) of fresh yeast in the form of paste, and
leaving the liquid undisturbed for from one to two hours
at a temperature of 25~ to 300 C. (770 to 86~0 F.). Powdered zinc shaken up with water, containing air in
solution, gives the same results.
I determined, by the help of M. Quinquand, the
weight of oxygen absorbed by the unit of weight of
yeast, in the unit of time, when this organism is placed
in water containing air in solution, without any mixture
of nutritive materials. These measurements were taken




io8             ON FERMENTATION.
by an oxymetrical process which I have invented, with
the assistance of M. C. Risler, one of my pupils. As
this method seems to me likely to be serviceable in
researches of this kind, and in the study of biological
phenomena, I think I ought to give the description of it
here, even at the risk of introducing a foreign element
into the examination of the facts which now occupy our
attention.
Process for the Volumetric Estimation of Dissolved
Oxiygen.-The process of measurement of the quantity
of oxygen dissolved in water, by means of a standard
liquid, which was proposed by M. Gerardin and myself,
(Comp. Rend., vol. 75, p. 879), and which I have
since improved, with the assistance of M. C. Risler,
depends essentially on the energetic reducing properties
of sodium hyposulphite. This salt* is obtained with the
greatest facility by the action of a solution of sodium
bisulphite on zinc, either in plates, shavings, or powder.
Its formation is much more rapid when the zinc employed is finely divided, and the points of contact
between the metal and the solution of bisulphite are
more numerous. Thus, with powdered zinc employed
in sufficient quantity, and a very concentrated solution
of bisulphite (marking 35~ Beaune, and requiring from
5 to 7 per cent. of its weight of powdered zinc), an
agitation of from three to five minutes is sufficient to
complete the reaction. This takes place, with elevation
of temperature, according to the following equation:3 (SO. NaO. HO) + ZN2 = S. NaO. HO + SO (NaO)2 + SO (Zn 0)2 + H2 0.
Sodium bisulphite.  Sodium hyposulphite.  Sodium sulphite.   Zinc sulphite.  Water.
* P. Schutzenberger on a new acid of sulphur, Ann. de Chim. et de Phys.,
vol. 20, p. 251 (4).




FUNCTIONS OF YEAST.               og9
If the bisulphite used in the experiment is concentrated, there are deposited, a short time after the cooling
of the liquid, crystals of the double zinc and sodium
sulphite, while the hyposulphite formed remains in solution, still mixed with the sulphites. The impure solution of hyposulphite (a mixture of hyposulphite and of
sulphite of soda and zinc) may be employed as it is for
the estimation; but it will only keep for any length of
time when protected from the air, and in a very dilute
state.
By adding to this liquid a suitable quantity of milk
of lime, we precipitate the zinc oxide, and by filtration
we obtain a solution very slightly alkaline, endowed,
like the former, with very decided reducing power;
possessing the property of keeping for a longer time,
when not exposed to the air, especially in a state of
great dilution, the form under which it is always used in
the quantitative estimation of oxygen. Without going
at farther length into the properties of sodium hyposulphite (see the memoir in the Ann. de Chim. et de Phys.,
before cited), I ought to particularize those which are
especially utilized in this process..- The sodium  hyposulphite, not saturated by lime,
absorbs oxygen with great rapidity, whether in the
form of gas, or in solution; its action is, in this respect,
similar to that of sodium pyrogallate.
By fixing the oxygen the hyposulphite becomes acid,
and is converted unto sodium bisulphite.
S. (Na. O) (H O) +- O = S O. (Na. O) (H O).
Hyposulphite.      Bisulphite.
When saturated by lime, it still acts in the same
manner on the gaseous oxygen, but more slowly; while




I IO            ON FERMENTATION.
it absorbs dissolved oxygen, instantaneously, and removes it from the oxygenated liquid with which it is
mixed.
2. The hyposulphite, poured into a solution of
ammonio-cupric sulphate, reduces the cupric oxide to
the state of cuprous oxide, destroying the colour of
the liquid, and then it reduces the cuprous oxide in its
turn, precipitating metallic copper; the reduction is
made at two intervals of time, and we are able, by employing more or less hyposulphite, to stop the process
at the first stage, shown by the decolouration of the
liquid.
3. The hyposulphite, whether acid or neutralized,
instantly destroys the colour, by reduction, of the solution of Coupier's blue (aniline blue), and of sodium
sulphindigotate (indigo carmine). These bleached solutions resume their blue tint when exposed to the
air.
4. If to water containing oxygen in solution, and
coloured blue by aniline blue'or indigo carmine, we
add, little by little, a dilute solution of hyposulphite,
either saturated or not with lime, the reducing agent
acts at first upon the dissolved oxygen, and does not
destroy the colour until it has absorbed it.
Thus, by taking two equal volumes of water tinted
blue by either of these colouring matters, saturating one
with oxygen by agitation with air, and depriving the
other of its oxygen by sufficiently prolonged boiling,
we shall find that the latter loses it colour after the
addition of a few drops of the hyposulphite, while
the second requires, in order to effect this result,
a much greater quantity of the reducing solution,




FUNCTIONS OF YXEAST.              II
and one in proportion to the quantity of dissolved
oxygen.
Here, however, a very remarkable peculiarity presents
itself, one which we ought specially to point out, because our ignorance of it would entail grave errors in the
analysis.  If we have some aerated water, and a suitably
dilute solution of the hyposulphite, either saturated or
not with milk of lime, we may previously determine the
oxymetric value of the hyposulphite, that is to say, the
volume of oxygen which is required to saturate the
unit of volume of the solution; it is only necessary for
this purpose to prepare a solution of ammonio-cuprous
sulphate, containing 4'46 grammes (68'826 grains) of
pure crystallized cupric sulphate to the litre (1'76
Eng. pints).  Such a solution having been brought
exactly to the bleached state, without precipitation of
metallic copper, that is to say, being brought back to the
state of solution of ammonio-cuprous-oxide, will have
yielded to the reducing liquid half of the oxygen corresponding with the cupric oxide which it contains,
about I cub. centimetre of oxygen ('o6  cub. in.) for
each Io cub. cent. ('6I cub. in.) of the solution.
It is sufficient, therefore, to determine with precision
the volume of the hyposulphite necessary to decolourize
completely Io cubic centimetres of the cupric liquor
without precipitation of metallic copper; this volume
will correspond to one cubic centimetre of oxygen.
This being determined, let us colour with a little
indigo carmine or aniline blue (just sufficiently to render
the tint perceptible) a certain quantity (for instance,
one or one half litre) of our aerated water, and let us
pour in the hyposulphite with a burette; the moment




I 12           ON FERMENTATION.
will come when the last drop will effect the decolouration of the liquid, which will rapidly pass from blue to
yellow. In this state, the clear yellow solution is a very
delicate test of free oxygen; it requires only the slightest
bubble of air, of the size of a pin's head to produce
very evident blue streaks. We are therefore induced
to admit that the dissolved oxygen has been completely utilized by the reducing liquid. This is not,
however, correct.  If we calculate, according to the
quantity of hyposulphite fixed by the cupric solution,
and the volume of the reducing liquid employed to
change the colour of the blue liquid, the quantity of
oxygen contained in a litre of water, we find, as nearly
as possible, half the oxygen really contained in this
water, and the mercurial pump or boiling could disengage from it. This remarkable result has been determined by a great number of experiments. What, then,
has become of the other half?
M. Risler and I thought, at first, that the products
of oxidation of the -sodium hyposulphite were not the
same when the oxidation took place under the influence
of free oxygen, as under that of the ammonio-cupric
oxide; however, after having ascertained that in both
cases sulphite was formed, and nothing but sulphite, we
were obliged to abandon this interpretation; we could
only believe that the diluted hyposulphite, acting, in the
cold, on the dissolved oxygen, divides it into two equal
parts, one of which is fixed in the reducing liquid, and
the other unites with the water, forming oxygenated
water or some analogous compound. This second half
of the oxygen, which has been, as it were, rendered
latent, acts no longer either on the hyposulphite or on




FUNCTIONS OF YEAST.'I I3
the indigo (discoloured carmine). When I say that it
no longer acts, I mean under the conditions of the experiment, which may be considered to be instantaneous,
and at a low temperature.
In fact, if we keep the bleached liquid-provided that
we have not used too little indigo (carmine)-for some
time from access of air, especially if we raise its temperature to 50~ or 60o C. (122~ to I40o F.), we see it become
blue again instantaneously, and throughout the whole
mass. The experiment may be made in a vessel filled
with an atmosphere of pure hydrogen to which is
fixed the extremity of a Mohr's burette, containing the
hyposulphite. If we now add a fresh quantity of the
reducing fluid until the second decolouration, the same
effect will be produced again, and until we have introduced a volume of hyposulphite nearly equal to that
employed in order to attain the first term of decolouration.
These experiments are delicate. To make them
succeed they must be completely protected from the
access of atmospheric oxygen; hyposulphite which has
been neutralized by lime, should be employed, and there
should be a sufficient quantity of indigo. (For further
details on this subject, see the Bulletin de la Sociedt
Chim. de Paris, vol. 20, p. I45, I873.)  They prove that
the first action of the acid hydrosulphite (which may be
considered instantaneous) on the aerated water coloured
by indigo, only removes half the oxygen. The other
half acts much more slowly on the reduced indigo, and,
by its inztervention, on the hyposulphite in excess. This
action does not manifest itself at all if the solution is
even slightly acid. In this case, the latent oxygen
6




I 14           ON FERMENTATION
may remain almost indefinitely in the presence of a
great excess of hyposulphite, or of the reduced solution
of indigo carmine, without being fixed by it.
By employing water coloured blue, to which has been
added a little oxygenated water (H2 O2), we produce
with the hyposulphite alternate decolourations, followed
by spontaneous recolourations, of the whole mass, which
resemble, so as to be indistinguishable from them, those
which take place in the experiments already described.
This similarity, added to the want of any other plausible
explanation, makes me think that the latent oxygen is
really found in the liquid under the form of oxygenated
water.
If we operate on a liquor rather acid than neutral, or
on a neutral liquor employing only hyposulphite not
saturated with lime, which becomes acid as it oxidizes;
finally, by taking note of the previous observation in
our calculation, that is, multiplying the quantity of
oxygen found by 2, we arrive at very close and satisfactory results.
I will first describe a rough method, susceptible of
being used anywhere, on the banks of a river, or in the
country, but which can furnish only approximate indications, by giving the amount of oxygen within a quarter
of a cubic centimetre per litre ('oI5 cub. in. per I'76
pint).
Some acid hyposulphite may be prepared, instantaneously, by agitating with zinc powder a diluted
solution of sodium bisulphite, prepared with supersaturating sodium carbonate by a current pf sulphurous
acid (the bisulphite at 350 Beaurne is a commercial product, and may be used). This bisulphite at 35~ Beaume




FUNCTIONS OF YEAST.              I I5
is previously diluted with four times its weight of water
and for Ioo grammes of the diluted solution we employ 2 grammes of zinc grey (powdered zinc). The
mixture and the agitation are to be made in a vessel
nearly filled with the liquid. After five minutes, the
solution must be filtered, and suitably diluted with water,
so that in a preliminary trial, one litre of water agitated
with air (saturated with oxygen under the pressure of
5 of an atmosphere at the ordinary temperature) and
tinted blue by some drops of a solution of aniline blue, or
indigo carmine, may be decoloured by about 25 or 35
cubic centimetres (I'525 to 2'I35 cubic in.) of the solution of hyposulphite.
The analysis requires nothing but a vessel with a
large mouth, (a wide-mouthed bottle), holding about
IS  litre, a stirrer which will allow us to mix together
the different layers of liquid without disturbing the
surface too much, one of Mohr's burettes, furnished
with a narrow tube at one end, fixed to the indiarubber tube of the. pinch-cock, and arranged so as to
be held mid-way in the water; also a glass bottle or
jar holding a little more than two litres, graduated so
as to indicate I litre.  A  litre  (about I    pints)
of the water to be tested, is introduced into the widemouthed bottle, tinted with aniline blue or indigo
carmine; then, the burette being filled with hyposulphite, and its lower end previously filled with the
liquid plunging midway into the water in the widemouthed bottle, we allow the reducing liquid to flow
in slowly, agitating the contents with the stirrer up
and down, so as not to disturb the surface too much;
the experiment is stopped at the moment that the




I6            ON FERMENTATION.
decolouration takes place, and the volume employed is
read off.
Immediately after, we proceed to the estimation of
the hyposulphite exactly in the same manner, by
employing I litre of the same kind of water which
served for the first experiment, but after having previously agitated it for some minutes with air, in the
large bottle, and taken its temperature. Under these
conditions, whether the original water be above or
below the limit of saturation for oxygen; we always
succeed quickly in having water saturated with oxygen
at the pressure of I of an atmosphere (the pressure of
oxygen in the air), and at the temperature which has
been read off. Tables of solubility, notably those of
Bunsen (Me'thodes Gazometriques,Traduction Frangaise
de T. Schneider) give the amount of oxygen.
Thus, in two experiments made under conditions
identically similar, we have the volume of the reducing
fluid required by the water whose oxygen is unknown,
and that required by the water whose oxygen is known.
A simple proportion will give the value of x in the problem. This process of estimating the hyposulphite, on
account of its simplicity and certainty, is preferable to
the employment of an ammoniacal solution of copper,
which M. Gerardin and I had proposed; it was suggested by M. Raulin, assistant director of the labora-:tory of M. Pasteur. As the operation is performed with
contact of air, it is necessary to make the measurements as quickly as possible, and to operate on a large
quantity of water (a litre), in order to counteract as far
as possible the influence of the oxygen of the air.
Besides, the method of analysis described above has




FUNCTIONS OF YEAST.              117
the effect of neutralizing almost entirely this cause of
error; the two operations being made under the same
conditions, the error can only proceed from a slight
difference between the conditions of the two experiments, such as their. duration, or the greater or less
agitation of the water.
I have succeeded, by the assistance of M. Risler, in
applying a similar.method of quantitative analysis.to
much smaller quantities of water, or oxygenated liquid;
and by modifying the process of the operation, I have
been able to ascertain by the reducing liquid, not the
hagf, but the whole of the dissolved oxygen; this
method is much preferable and more certain. In order.to attain this double result it is only requisite; Ist. To
make the analysis in a liquid completely protected from
access of the oxygen.in the air, by an atmosphere of
pure hydrogen: 2nd. To introduce the aerated water that
is to be tested, (a known volume, from 40 to Ioo  cubic
centimietres (2'44 to 6'I cub. in.) into a tepid, 400 or 500
C. (Io4~ to I220 F.) neutral, or very slightly alkaline,
but never acid, medium, formed by a solution of indigo carmine, just decoloured by hyposulphite which'has been previously neutralized or rendered slightly
alkaline by milk of lime. This yellow medium turns
blue under the influence of dissolved  oxygen; a
quantity of blue indigo is re-formed, proportional to
the amount of oxygen dissolved. If the preceding
conditions of temperature and neutralization have been
carefully observed, all the dissolved oxygen is utilized
in oxygenating the reduced indigo, and there only
remains to be estimated, by means of the hyposulphite,
the volume of this reducing agent necessary to de



I I8           ON FERMENTATION.
colour the blue liquid. The same experiment is repeated immediately after, with the same volume of
water, agitated at a known temperature, and calculation
will give, as before, the volume of oxygen sought. If
the liquid is acid, or becomes so in the process of
analysis, the conditions leading to the formation of
oxygenated water are at once present, and the results
are deceptive, and always too low, approaching more
or less nearly to the half of the whole amount of
oxygen.
If, then, we operate on acid liquids, we ought to
add previously to the yellow indigotic liquid a sufficient
quantity of a diluted solution of ammonia to correct
this disturbing condition.  The principles of the experiment being known, I will enter into some details
concerning the apparatus, and the preparation of the
reagents.
I. Sodizui- Hyposzdlphiie.-We prepare the acid hyposulphite in the manner described above.   This is
neutralized by milk of lime. For Ioo grammes of
concentrated bisulphite at 350 Beaume6, employed for
the preparation of the acid hyposulphite, we shall
make use, for this purpose, of 35 grammes of milk of
lime, prepared from 200 grammes of lime previously
slaked, to I litre of water. The saturation is effected
with the acid hyposulphite, diluted, as I have before
said, with four -times its weight of water, but the
quantity of milk of lime is calculated according to
the weight of concentrated bisulphite that is employed.
It is shaken up, left to settle, decanted, and then
filtered; the liquid is kept in full and well-stoppered
bottles.




FUNCTIONS OF YEAST.              I9When used, it is necessary to dilute this liquid with
distilled water, until 50 cubic centimetres (3'05 cub. in.),
of water saturated with oxygen require from 4 to 5
cubic centimetres ('244 to'305 in.), of the reducing
agent.
The solution thus prepared may be kept almost indefinitely at the required strength, if we take the following precautions.  It is to be placed in a bottle containing about I litre (I 4 pints) filled to the neck, closed
by a good india-rubber cork, perforated with two holes;
in one of them is fixed a tube bent at right angles,
whose extremity dips to the bottom of the bottle, and
whose other end has an india-rubber tube, furnished
with a Mohr's pinch cock; in the free end of the indiarubber tube, is fixed a piece of glass tube; in the
second hole is fixed another tube bent at right angles,
but which only goes about I or 2 centimetres ('39 or
~78 in.) past the cork. This tube is kept in permanent
communication by means of a sufficiently long indiarubber tube, with a gas-jet kept turned on. It is proper
to interpose between the gas-burner and the bottle a
glass vessel like those employed by chemists for drying
gases; this vessel is filled with pumice stone, saturated
with a concentrated solution of sodium pyrogallate.
By this means, we get rid of the oxygen which common
gas always contains (from I to 2 per cent.) The
burettes are filled with hyposulphite by aspiration, from
below upwards.  For this purpose, the india-rubber
pinch-cock tube, which, in order to serve another purpose, ought to be sufficiently long, is placed in communication with the tube which goes to the bottom of the
hyposulphite, and we draw in the liquid by the mouth,




120             ON FERMENTATION.
by means of an india-rubber tube, fixed by a cork and
a bent glass tube to the upper part of the Mohr's
burette. By this means, we avoid the agitation of the
liquid in contact with the air. With these precautions,
the strength of the test does not alter; it is, however,
prudent not to trust to this, but to ascertain it at every
fresh experiment, which is very easily done.
Indrigo.-We prepare beforehand ten litres of the
solution of indigo carmine, by dissolving in this quantity nearly 200 grammes (6-42 oz. troy), of indigo
carmine in the form of paste (sodium sulphindigotate).
The liquid should be kept in blue or black glass bottles,
sheltered from the light.
The estimation is made in a three-necked bottle, of
I or I   litres in capacity (Fig. I9). One of the three
lateral necks receives, through an india-rubber cork, the
tube, Hy., bent at right angles for the admission of
hydrogen. It is well that this tube should slide in its
cork without too much friction, in order that it may
be raised or depressed at will, without allowing air to
enter.
The other lateral neck has an india-rubber cork,
pierced with two holes., In one of these is fixed the
end of a small funnel with a glass stop-cock, E, a
bromide funnel; this end should be sufficiently long to
reach the bottom of the bottle.
In the other hole is fixed the tube which carries off
the hydrogen; this tube, being twice bent, has its free
extremity plunged into the mouth of a test-tube, T,
which is fixed there by means of a cork, also pierced
with two holes; the tube contains water, and  the
hydrogen in excess, after having passed through this




FUNCTIONS OF YEAST.                 121
water, escapes by a tube bent at right angles, fixed in
the second hole of the cork.
The middle neck of the bottle holds as a fixture a
stopper of cork or india-rubber, hermetically cemented
in, and pierced with two orifices, in which are also permanently fixed the two pointed extremities of two
Mohr burettes, H I, held above the bottle by a special
2g )             l-[y  -~  -— ~,, I —
FIG. I9.- Apparatus for the measurement of oxygen dissolved in water.
support, one by the side of the other. The india~rubber pinch-cock tubes at the lower end of these
two burettes ought to be long enough to allow the
bottle to be moved without the burettes being shaken;




122            ON FERMENTATION.
they are fixed to the burettes, and ought, on the contrary, to be able to be detached, at will, from the thin
glass ends which pass through the middle cork of the
bottle. It is well to have at hand a third burette, either
independent, or fixed to the same support by the side
of the others. It is intended as a reserve for the hyposulphite of the principal burette, H. One of the first
two burettes, I, contains indigo carmine, the two others
are filled with sodium hyposulphite.
This having been arranged, when we wish to measure
the oxygen dissolved in any particular specimen of
water, we introduce into the large bottle(I) About 50 cubic centimetres (3'05 cub. in.) of
solution of indigo.
(2) About 250 cubic centimetres (I5'25 cub. in.) of ordinary warm water, at 400 to 500~ C. (Io4~ to 122~ F.). We
adapt to the fixed sockets the burette, I, with the indigo,
and the inzdependelZt burette of hyposulphite; we charge
the two sockets with the contents of the burettes, and
allow the hydrogen to pass rather quickly. (The apparatus producing the hydrogen may be of any kind,
provided that we are able to regulate the disengagement
of gas by means of a single stop-cock; the hydrogen
is purified by water, and passes through a column of
caustic potash in plates.) When we suppose that almost
the whole of the air has been swept away by the current
of hydrogen, we allow the hyposulphite to flow in
slowly, holding the bottle in the hand, and giving the
liquid a gyratory motion sufficient to mix it, until it has
only a slight greenish tint, or reddish if it is alkaline;
then, without interrupting the current of hydrogen gas,
we detach the independent burette, and fix on the hypo



FUNCTIONS OF YEAST.             123
sulphite burette,.H, intended for the analysis. In doing
this, the test liquid contained in the socket escapes, and
often completes the decolouration of the water by turning it yellow. We charge the socket again by slightly
opening the pinch-cock, then by adding first a little
carmine, then a little hyposulphite, we bring the liquor
to a bright yellow colour, which verges towards green
or red by the addition of a single drop of carmine.
We notice that the yellow liquid does not turn blue at
the surface, which proves that the atmosphere of the
bottle is thoroughly deprived of oxygen. Then all is
ready for the analysis if we have taken care, at first,
to fill the lower end of the funnel with the same water
as that whose contained oxygen we wish to measure.
This estimation is made by proceeding in the following
manner, and in the order indicated:(I) Slacken the current of hydrogen, without stopping
it; (2) Raise the supply tube, so that the gas should
not bubble up; (3) Read off the graduation of the
hyposulphite burette, at which the surface of the liquid
stands; (4) Introduce into the funnel 50 or Ioo cubic
centimetres of the water to be tested, and allow this
water to run at once into the bottle, keeping the lower end
full from the stop-cock; shake the bottle; the standard
yellow liquid grows blue if the water is aerated, and
does not change its colour if it contains no oxygen.
We now need only allow the hyposulphite to run in,
drop by drop, shaking it, and noticing the exact limit of
decolouration which takes place, true to a single drop
(-0-' of a cubic centimetre) if the liquid is without colour;
Immediately after having noted the point at which
this takes place, without changing or disturbing any



124            ON FERMENTATION.
thing, fill the body of the funnel with water saturated
with oxygen, at a pressure of ~ of an atmosphere, and
at a known temperature; correct with some drops of
hyposulphite the slightly blue tint developed by this
operation, and proceed to estimate the value with 50
or Ioo cubic centimetres of saturated water.  A' rule
of three sum gives the quantity of oxygen in the first
portion of water.
Instead of testing the hyposulphite with saturated
water at a known temperature, we may estimate it by
allowing a known volume (25 cubic centimetres) of solution of indigo carmine to flow into the bottle after-the
first trial, and ascertaining the volume of the reducing
agent necessary to bring back the decolouratioh.
On the other hand, we have determined, once for all,
the volumetric ratio between the indigo employed, and
a solution of ammonio-cupric sulphate, containing 4'46
grammes of pure crystallized cupric sulphate to the litre;
this amounts to Io cubic centimetres = i cubic centim'tre of oxygen at the moment of complete decolouration. It is sufficient for this purpose to ascertain the
volumes of the same solution of hyposulphite necessary
to decolour equal volumes of indigo carmine and -of the
cupric liquid, which allows us to calculate the ratios of
volumetric equivalence of these two liquids, and consequently the volume of oxygen corresponding to I
cubic centimetre of the indigo solution.
This experiment ought to be made with great care,
for on its accuracy will depend that of the absolute
values of all the subsequent analyses.
We determine the ratio between the hyposulphite
and the indigo under the. same conditions as those of




FUNCTIONS OF YEAST.              I25
the preceding estimation, since these are the conditions
to which we shall always come back in future experiments.
The ratio between the same hyposulphite and the
cupric solution is determined by operating in an atmosphere of pure hydrogen. The cupric solution.(I5 to 20
cubic centimetres) is placed in a small bottle with three
necks, one for the admission the other for the exit of
the gas, this latter being furnished with a small stirring
apparatus, similar to that: on the. large bottle; on the
middle neck of the bottle is fixed, by an india-rubber
tube, the socket of the hyposulphite burette, lengthened
and drawn out to a fine point, and charged beforehand.
The reducing agent is allowed to run in as soon as the
air is expelled, and this is continued till complete decolouration takes place; this point is somewhat difficult
to ascertain.
Exammple.-By operating as above, we have found
that(I) 4'6 cubic centimetres of hyposulphite were equiva-'lent to 50 cubic centim'etres of indigo.
(2) I 5'  cubic centimetres of hyposulphite were
equivalent to 25 cubic centimetres of ammoniacal solution of copper; Io cubic centimetres of decoloured
cupric solution = I cubic centimetre of oxygen (at o~
C. (320 F.), and 760 millimetres of pressure). We easily
find that i cubic centimetre of indigo, corresponds to
0'OI52 cubic centimetres of oxygen.
On the other hand;
For Ioo cubic centimetres of water to be tested, we
have employed 5 7 cubic centimetres of some hyposulphite.




126             ON FERMENTATION.
20 cubic centimetres of indigo require 2,9 cubic centimetres of this hyposulphite.
We make the proportion:2-9: 20:: 5 7: =x=_ 20X9 57'  39'3 cubic centimetres.
The oxygen of Ioo cubic centimetres of water corresponds to 39'3 cubic centimetres of indigo. Nothing
remains but to multiply 39'3 by O'OI52- 0'59736, to
find that one litre of water contained 5'97 cubic centimetres of dissolved oxygen, measured at o~ C. (320 F.)
and at 760 millimetres pressure.
N.B.-It is important, when we make a series of
analyses, never to use the last cubic centimetres of the
burette of hyposulphite, which, having remained in contact with the air, have lost their reducing power. If we
leave more than half an hour's interval between the trials,
it would be better to renew entirely the contents of the
burette.
A complete experiment, including the preparation of
the reduced medium, does not require more than ten
minutes, and each of the succeeding measurements is
made in one or two minutes. Two trials of the same
kind, repeated, never differ more than -ix- of a cubic
centimetre.
This process of analysis allowing us to estimate the
oxygen dissolved in 50 cubic centimetres of water (3'05
cub. in.) with an approximation of o0005 cubic centimetre, and consequently of o'I cubic centimetre per
litre, we have been able to utilize it for the purpose of
studying the respiratory phenomena of yeast, and of
measuring their intensity under different conditions of
temperature. The rapidity of the estimates, which only




FUNCTIONS OF YEAST.              127
require three or four minutes, gave us the opportunity
of multiplying the experiments, and establishing the
results which are given below by a series of analyses,
the number of which was not obliged to be restricted.
The experiments to which I shall here allude apply only
to the case of fresh yeast in the form of paste, containing
from 29 to 30 per cent. of solid matter, diffused in pure
aerated water, without the addition of any nutritive
element.  We shall presently see what takes place
when the yeast is placed under conditions more favourable to its developement.
The method consists in leaving a known weight of
yeast for a known length of time in contact with a
known weight of water, under the conditions of temperature which we may wish to study. The oxymetrical
degrees of the water are measured at the commencement and the end of the experiment. Their difference
gives the oxygen absorbed.
The yeast of beer only shows the phenomenon of absorption of oxygen, with production of carbon dioxide.
All things else being equal, the respiratory intensity
is the same in the dark, in diffused, and in full light;
it is proportionate to the weight of yeast employed.
The original amount of dissolved oxygen does not
sensibly influence the results, except when it is below
i cubic centimetre per litre. We find, in this case, a
feeble diminution in the rapidity of the absorption,
which continues till complete deoxygenation of the
water.
Below Io~ C. (50~ F.), the absorbing power of the
yeast for oxygen is almost nil; it increases slowly to
18~ C. (about 65~ F.); from this point the increase is




l128            ON FERMENTATION.
rapid till about 35~ C. (95~ F.), a temperature at which
the respiratory intensity attains its maximum, which is
sustained sensibly till 500 C. (1220 F.). At 600 C. (I400
F.), the absorbing power is annulled and destroyed.
A specimen of yeast, sensibly fresh, containing 26
per cent. of solid matter, absorbed, per gramme and per
hour, at 9~ C. (about 490 F.), o'I4 cubic centimetre;
at II~ (520 F.), 042 cubic centimetre; at 220 C. (about
720 F.), I'2 cubic centimetre; at 330 C. (about 93~ F.),
2'I cubic centimetre; at 400 C. (Io40 F.), 2'06 cubic
centimetres; at 500 C. (I220 F.), 2'4 cubic centimetres; at
600 C. (I400 F.), none.
Another specimen of yeast, very fresh, and of very
good appearance, containing 30 per cent. of solid matter,
absorbed, per gramme and per hour, at 240 C. (about
760 F.), 2'2 cubic centimetres of oxygen; at 360 C.
(about 970 F.) Io'7 cubic centimetres.
The increase of absorbing power between 240 C. and
360 C. was therefore more considerable than with the
first yeast; this power is doubled in the one case, and
quintupled in the other.
The values of the quantities of the oxygen absorbed,
as well as the magnitude of the variations with relation
to the temperature, are by no means absolute; they
depend on a particular factor, inherent in the yeast,
which we may call the factor of vitality; but what-.ever this factor may be, the direction of the variations
is always the same, and susceptible of being represented by a curve starting from the line of abscissae, or
line of temperature, at about 9~ or Io~ C. (about 400 or
50~ F.), rising slowly up to I8~ C. (about 65o F.), thence
rapidly reaching, at about 350 C. (950 F.), a maximum




FUNCTIONS OF YEAST.              129
lieight, which it retains till nearly 600o C. (I400 F.), when
it returns suddenly towards the line of abscissae
Yeast can not only utilize and cause to disappear the
oxygen physically dissolved in water, but also oxygen
combined with hwemaglobin, which, as we know, can be
eliminated by a diminution of pressure.
Thus, when we diffuse fresh yeast, whether washed or
not, in arterial red blood, or in a solution of hoemaglobin
saturated with oxygen, we see the tint change rapidly
from  red to dark blue or black.  A simple agitation
of the blood with air is' sufficient to restore its ruddy
colour; then the phenomena of deoxygenation recommence; the same experiment maythus be repeated a great
number of times, especially with fresh and washed yeast.
Although, in this case, the yeast is in contact with a
medium infinitely more rich in oxygen than is aSrated
water (containing from 200 to 230 cubic centimetres of
oxygen to the litre, instead of from 6 to 7 cubic centimetres), the rapidity of the absorption is not increased,
if the conditions of temperature are the same.
One gramme of yeast absorbs as much oxygen in an
hour,. at the same temperature, whether it be mixed with
water containing 5 -or 7 cubic centimetres of oxygen per
litre, or in arterial blood containing 200 cubic centimetres
of oxygen.
In the experiment with blood, we might fear a direct
influence of the yeast, or of its soluble materials, on the
colouring matter of blood; this influence is, in fact,
produced, especially with solutions of hemaglobin; it
shows itself by the transformation of this primordial
colouring matter into haematin; but it only makes its
appearance after some hours.




130            ON FERMENTATION.
The behaviour of the yeast with reference to blood
may be explained in the following manner: The cells
of Sacc/zaromzyces diffused in the liquid breathe at the
expense of the oxygen physically dissolved in the plasm
or serum in the midst of which swim the red globules
of blood. In proportion as the plasmic liquid grows
less rich in oxygen, a portion of this body, feebly combined with haemaglobin, is separated, and enters into
physical dissolution, by a dissociation comparable to
that presented by potassium bicarbonate in a vacuum;
the process continues till there is a complete disappearance of the oxygen dissolved in the serum, and of that
which is fixed in the haemaglobin. If this explanation
is correct, the experiment ought to succeed, even when
the blood is separated from the yeast diffused in water
or serum by means of a membrane permeable to
gas and to liquids, but capable of preventing all direct
contact between the yeast-cells and the red globules.
This is, in fact, what takes place. I have thus been
able, by arranging a suitable apparatus, to imitate artificially that which takes place in the organs and tissues
of animals, when the red and oxygenated arterial blood
traverses the network of capillary vessels, and passes
out into the veins under the form of black and partially
deoxygenated blood.
For this purpose, it is only necessary to cause red
blood to circulate slowly through a sufficiently long
system of hollow tubes, the walls of which are formed
of thin gold-beater's skin, which is immersed in a mixture of yeast diffused in fresh serum, without globules,
kept at 350 C. (950 F.).
We see the red blood pass out black and venous at




FUNCTIONS OF YEAST.              131
the other extremity. A confirmatory experiment, made
at the same time, with a system of tubes precisely
similar, but immersed in serum without yeast, proves
that yeast is indispensable for thus rapidly effecting the
deoxidation of the blood. This experiment is the
exact representation of what takes place in the animal
organism, with the exception of the perfect method
employed by nature to multiply contacts and surfaces.
In the latter case, the cellular and histological elements of the tissues play the part of the yeast; they
absorb the oxygen dissolved in the plasmic liquids which
bathe them, and constantly tend to bring down to zero
their oxymetric condition. The oxgen, but feebly fixed
in the hmemaglobin, re-establishes the equilibrium by a
series of gaseous diffusions from the red globules to the
plasm of the blood, and from the plasm of the blood to
that of the organs. These continual diffusions are the
inevitable consequence of the disturbance of equilibrium
produced by the aeration of the organic cells, or of
the cells of yeast in the experiment just described.
All these facts, then, prove distinctly that yeast
breathes when placed in contact with dissolved oxygen
The measure of the respiratory power, under the most
favourable conditions, shows us this respiration to be
as active, and even more so, than that of fishes. This
cannot be considered as only a curious accessory fact,
of which we must take but slight notice, in the study
of the biological phenomena of yeast. For, a priori, it
is improbable that a function so distinct, so sharply
defined, is of no serious importance. On the other
hand, if we consider what is passing in other living
beings, from the highest to the lowest ranks in the scale




32            ON FERMENTATION.
of animal and vegetable- life, we see respiration-that is
to say, combustion- at the expense of oxygen, playing
a preponderating part.
Without dwelling on the animal kingdom, we may
remember that it has been long known that plants, in
darkness, absorb oxygen, and disengage carbon dioxide;
it was,even suspected,. with good reason, that this respiratory function, the reverse of that shown by parts:
exposed to the sun, was independent of the diurnal
respiration, and that it belonged to another class of cells
containing no chlorophyll. By operating on immersed.
aquatic plants, we can demonstrate this fact in the
clearest and most elegant manner. Let the fresh stalks
of Elodea be immersed in aerated water, of which the:
original oxymetrical degree is known. The flask, complctely filled, is placed in the dark.'At the end of two
or three hours, we test the quantity of oxygen, and
we'find a diminution, which, as was the case with the
yeast, at equal temperatures, is proportional to the
quantity of the plant, and the duration of the experiment, and. whose absolute amount varies with the temperature.
If,,now, we warm for a moment the water and. the
plant/ to about 500 C..(I220 F.), we shall destroy for ever
the activity of these cells containing chlorophyll, that is
to say, the powers.which the. plant possesses of decom,
posing carbon dioxide by its green parts; but without
lessening its activity in respiration or combustion. We
have seen, in fact, in yeast, that.this function is only
finally modified at about 60o  C. (I400 F).  The green
parts of the plant are dead, but it.is still capable of fulfilling certain biological functions. The. flask of aerated




FUNCTIONS OF YEAST.             I33
water may be exposed to the sun's rays; when, far
from observing an increase in the quantity of dissolved
oxygen, the opposite is seen.
It is especially in the parts of plants which have to
undergo a rapid evolution, and a marked cellular development, that the absorption of oxygen and internal
combustion show themselves to be very active.
Every one knows that in the germination of seed or
grain, that of barley in the manufacture of beer, for
instance, the internal combustion develops a considerable
quantity of heat.
The blossoming of flowers is also accompanied by a
very marked oxidizing respiration.
Returning to yeast, M. Pasteur has shown (Bullet,
Soc. Chimique, p. 80o, I86I) that beer-yeast, sown in an
albuminous liquid, such as the water of yeast, multiplies,
even when there is no trace of sugar in the liquor, provided always that the oxygen of the air be present in
large quantity; deprived of air, and under these conditions, the yeast throws out no buds.  The same
experiments may be repeated with an albuminous liquid
mixed with a solution of unfermentable sugar such as
crystallized sugar of milk; the results are of the same
kind.
The yeast formed thus, in the absence of sugar, has
not changed its nature, it causes sugar to ferment if it
has been made to react on that substance without the
access of air. We must, however, remark, that the
development of yeast is very difficult when it has nofermentable matter to nourish it.
On the other hand, this same observer has remarked
that, when in contact with air, and when it is extended




134            ON FERMENTATION.
over a large surface, alcoholic fermentation is more
rapid than when deprived of oxygen, and that the
budding is more active, since, notwithstanding the great
rapidity of the fermentation, the relation between the
newly-formed yeast and the decomposed sugar passes
from -8' to "- or -IM. Mayer (Landw. Versuchs., vol. I6, p. 290) performed experiments, from which it would appear that
oxygen has no influence, either on the rapidity of the
fermentation or on the quantity of newly-formed yeast.
However, his process of aeration of the liquids in
fermentation, which consists in allowing calcined air to
pass into the flask three times a day, appears to me to
be insufficient, the slowness with which the water absorbs
oxygen, and the rapidity with which yeast absorbs it,
being well known; we cannot, then, draw  from  the
experiments of this author the conclusions unfavourable to Pasteur's theory which he drew from them.
The results of all these facts are,-That yeast, like
ordinary plants, buds and multiplies even in the absence
of fermentable sugar, when it is furnished with free
oxygen; that this multiplication, however, is favoured
by the presence of sugar, which is a more appropriate
element than non-fermentable hydrocarbon compounds;
and also, that yeast is able to bud and multiply in the
absence of free oxygen, but that in this case a fermentable substance is indispensable.
We are, therefore, compelled to arrive at the conclusion which M. Pasteur has drawn from all these facts;
that saccharine matter can supply free oxygen in proportion to the yeast, and can excite it to bud. M.
Pasteur has gone farther; he thinks that the ferme nt



FUNCTIONS OF YEAST.                135
ing character of a cell is due to the power which it
possesses of breathing at the expense of sugar, without
the contact of air, and that the decomposition into
alcohol and carbon dioxide is the consequence of a
disturbance of equilibrium, due to this partial abstraction of oxygen. We may also interpret the facts in the
following manner:(I) The supply of oxygen, and the combustions to
which it gives rise are necessary for the development
and the reproduction of cell life. This fact is abundantly established for all the forms of life and organs
of the vegetable kingdom.
(2) Yeast possesses the power of resolving the sugar
which penetrates by endosmose into the interior of the
cell into alcohol, carbon dioxide, glycerin, succinic acid,
and oxygen. In fact, we have before seen (p. 23 of the
French) that M. Monoyer had proposed a very simple
equation to represent M. Pasteur's formula relative to
the formation of succinic acid and glycerin.
In this equation which we give below,4 (C12 Hll 1, +H O), or*
4 (C2 H12 012) + 6H O = C8 H   Os +6 C6 H8 O - 2 C2 04 + 02
we see an excess of oxygen make its appearance in
the second member of the equation, and M. Monoyer
adds, "we may suppose that this oxygen in excess
serves for the respiration of the yeast. After this, the
idea is not without foundation that fermentation is a
primary phenomenon, due to a special action of the yeast
and of other cells (Sechartier and Bellamy), and that,
* This is the old notation form of =" given by M. Schiitzenberger; the
new notation form will be found on the page referred to.




136            ON FERMENTATION.
as a consequence of this fermentation, there is some
oxygen to spare, which may serve the purposes of
respiration, and consequently may promote the budding
of the yeast.
In this manner of explaining the facts, the yeast
would not become a ferment because it breathes a part
of the oxygen of the sugar; but it may breathe a part
of the oxygen of the sugar, and consequently reproduce,
precisely because it sets free oxygen by decomposing
the sugar.
Looking at the question in a more general point of
view, we may also say that respiratory combustion is
to the living organism a source of energy necessary for
its development. In the decomposition of sugar there
would be, according to the calculations of M. Berthelot,
a disengagement of heat; the quantity of heat set free
in this phenomenon would be about -l. of the heat
disengaged by the complete combustion of the sugar
decomposed.: (Comp. Rend., vol. 59, p. 90I.) In this
estimate no account has been taken of the heat of
solution of the sugar which disappears, nor of that of
the solution of the alcohol formed, positive quantities
which would tend to raise the heat of fermentation. It
is not even necessary, therefore, to have recourse to
the hypothesis of a combustion at the expense of the
oxygen of the sugar, to explain how the phenomenon of
fermentation can feed the combustion, and become a
source of the energy indispensable to the development
of the plant.
However this may be, there is an evident correlation,
as M. Pasteur observes, beween fermentation and the
development, nutrition, and budding of yeast. In fer



FUNCTIONS OF YEAST.              I37
mentation without oxygen, the relation between the
new yeast and the decomposed sugar will be necessarily greater than in fermentation with oxygen, since
in the former case the budding takes place only under
the influence of the oxygen furnished by the sugar, and
in the second, of that furnished both by the medium
and the sugar.
We ought to examine what takes place when yeast is
left to itself, without nourishment and in a damp state,
without the intervention either of saccharine matter or
oxygen, before we proceed to the study of the chemical
modifications produced in yeast, while it is placed under
conditions -favourable to its nutrition and development.
These conditions, as we have seen, return always to
those of alcoholic fermentation, sugar being one of the
indispensable sources of nourishment of the Sacczarovnyces, and this cannot be in contact with it without
fermenting, provided that the other conditions of nutrition are fulfilled.  Does it preserve, in this case, its
original integrity, without any change in the chemical
composition of its immediate principles?  In other
terms, does its vitality remain in a latent state, to manifest itself afresh, as soon as sugar or oxygen is supplied?
A priori, this would appear improbable, if we regard
what takes place in the tissues of plants.  In fact,
experiment has proved that, under these conditions, the
yeast undergoes important modifications, in respect of
the composition of its organic principles.
This question has been studied by M. Pasteur first,
and then successively by M. Bechamp and by the
author of this book.
M. Pasteur having set 5 grammes of sugar to fer7




138               ON FERMENTATION.
ment with Io grammes of yeast, in the form  of paste,
(2'I55 grammes of dry matter), a much larger quantity
than is necessary for the complete decomposition of the
sugar, was surprised to see that this fermentation did
not entirely end, that it had a tendency to continue with
a weak disengagement of gas, and when Fehling's liquor
no longer revealed in it the slightest trace of sugar.
Having arranged in receivers reversed over mercury the
following fermentationsIst. Sugar-candy......'313
Yeast from wine (deposit in empty casks)  6'95o
Pure water.                 9'336
2nd. Sugar-candy...                  I'4425
Yeast from beer (2'I 5 grammes when dry) Io'o
Pure water.o   9'210
He obtained, in two days, while the gaseous disengagement was still sensible, from —
No. I. 360 cubic centimetres (2I'96 cub. in.) at zero (320 F.), and
at 760 millinmetres' pressure;
No. 2. 387'5 cubic centimetres (23'6 cub. in.) at the same temperature and pressure.
Of carbon dioxide, entirely absorbable by potass. The
theoretical quantities, even when a deduction is made of
the succinic acid and glycerini-that is to say, those
which correspond to Gay-Lussac's former equationthose which give the highest number for carbon dioxide
would beNo. I., 34I 8.
No. 2...      375'5.
By increasing still more the  amount of yeast, we
arrive at more conclusive results.




FUNCTIONS OF YEAST.             139
Thus,'424 gramme of pure sugar-candy, made to
ferment with a weight of damp yeast corresponding
with Io grammes of dry matter, furnished, at the end
of two days, 300' cubic centimetres of carbon dioxide;
the sugar alone could only furnish IIO cubic centimetres.
The liquid, carefully distilled, gave a little more than
o'6 gramme of absolute alcohol. The weight of alcohol
obtained was greater than the whole weight of the
sugar employed, and in proportion to the volume of
carbon dioxide formed.
This experiment proves that when yeast is mixed
with a comparatively very small weight of sugar, after
the latter has been decomposed, the activity of the
yeast continues, reacting on its own tissues and its
hydrocarbons with extraordinary energy and rapidity,
proceeding more and more slowly as the process
goes on.
If we put an end to the fermentation at the moment
when a volume of carbon dioxide is formed, equal or
very little greater than that which corresponds with the
weight of the sugar employed, wefied no more sugear iJn
th/e liqor. This observation is very important, because
it tends to prove that the action which the yeast exerts
on its own'elements does not commence till it is deprived of sugar.
It is not necessary to fulfil the conditions of the
preceding experiments (fermentation with excess of
yeast), in order to observe fermentation at the expense
of the elements of the yeast itself; it is sufficient, as
Pasteur has already shown, to mix fresh beer-yeast with
water at 250 C. (77 ~ F.); we soon see numerous bubbles




140            ON FERMENTATION.
of pure carbon dioxide gas rise, and it is easy, by distillation, to ascertain the production of alcohol. Hydrogen gas and signs of putrefaction do not appear till
long afterwards, when the microscope reveals the
presence of lactic ferment and infusoria.
M. Bechamp, who took up this question after M.
Pasteur, took care to avoid completely the formation
of infusoria, by employing creosote water; he also
ascertained that alcohol and carbon dioxide were
produced as a consequence of the vital activity of the
yeast when without nourishment (deprived of oxygen
and sugar). But this is not all, for this curious phenomenon is allied to other chemical reactions not less
interesting.
It is well known that, by preserving yeast, in the form
of damp paste, in a hot place (250 to 300 C., 77~ to
86" F.), it undergoes considerable softening, and entirely
changes its appearance.
This modification is not due to incipient putrefaction.
There is no foundation for this conclusion; nothing is
formed as a volatile product, except carbon dioxide and
alcohol. The microscope reveals no organism except
the Saccharornyces, especially if, as M. Bechamp suggests,
we make use of creosote water. If, now, we treat this
softened yeast with luke-warm water, we'can extract
from it a much greater quantity of soluble and diffusible principles than by employing fresh yeast.
Thus, Ioo grammes of fresh yeast, leaving after dessication at Ioo~ C. (2120 F.), a fixed residue of 30
grammes, give after washing, before the softening takes
place, only 23 grammes of dry residue. The loss by
washing is 8 per cent.




FUNCTIONS OF YEAST.             141
The same yeast, softened spontaneously for two days,
and then washed, leaves a residue weighing, when dry,
I4 grammes; the loss by washing is thus I6 grammes.
Yeast has therefore, by softening, transformed into
soluble principles 8 grammes of previously insoluble
principles, per cent. of damp yeast, or 26'66 grammes
per cent. of dry yeast.  During this internal reaction
of yeast, when kept damp, and without nourishment,
M. Bechamp observed the production of pure nitrogen.
The water by which it has been washed contains
acetic acid, a perceptible portion of soluble alterative ferment (zymase, of which we shall presently speak; see the
chapter on soluble ferments), an albuminoid principle,
soluble, coagulable by heat, and nearly allied to albumin,
from which it differs by rotating power; a gummy matter,
which nitric acid transforms into mucic acid, very like
arabin, from which, however, it differs by rotating power;
we find, also, tyrosine and leucine, and an uncrystallizable
sirupy matter, alkaline and alkaline-earthy phosphates
in observable proportions.
Resuming the question, the author confirmed the
results obtained by M. Bechamp, and added to them
new facts. The extract of softened yeast contains,
besides the principles mentioned above, nitrogenous
compounds of the sarcine group, which had not hitherto
been noticed in the vegetable economy.
The following is the analytical process which I have
followed. The yeast, digested without the addition of
any nutritive principles, is boiled with a considerable
quantity of water to coagulate the albumin; it is then
filtered. The liquid, which has a slightly acid reaction,
is concentrated in a water bath to a sirupy consistence;




142             ON FERMENTATION.
it assumes, as it grows cold, a gelatinous crystallized
form  composed of small crystals forming a kind of
paste in a brownish syrup. The whole, placed' in a
flask, is boiled for some time with a great excess of
strong alcohol (92 per cent.); a deep coloured pitchy
mass is separated, which sticks to the sides of the
flask. The alcoholic solution, when properly concentrated, furnishes, as it grows cool, an abundant crystalline deposit, which, after filtration, washing with cold
alcohol, and pressure, is almost white. This crystallized
mass, formed of very thin flakes or hyaline globules, as
well as that which is obtained by concentrating the
alcoholic mother-liquor, is almost exclusively formed of
sulphuretted pseudo-leuzcine, with a little tyrosine. The
mother-liquor separated at the second crystallization
is distilled in a water-bath to drive off the alcohol.
The remainder is diluted with water, and solution of
barytes is added to precipitate the phosphates.  The
excess of barytes is removed from the filtered liquid by
a current of carbon dioxide. The filtered liquid is then
boiled with an excess of cupric acetate. A brownish
flaky precipitate is formed, which contains carnine,
sarcine, xanthine, and guanine, combined with copper
oxide. The filtered liquor above this precipitate, which
is comparatively not very abundant, yields, when the
copper has been removed by sulphuretted hydrogen,
and it has been concentrated, a crystalline mass, from
which cold alcohol extracts a sirupy nitrogenous substance, of a sweet taste, leaving a crystallized mixture
of leucine and butalinine.
The cupric precipitate, produced by boiling in cupric
acetate, is thoroughly washed with hot water, then




FUNCTIONS OF YEAST.              143
treated, by the warm process, with dilute hydrochloric
acid. Nearly all of it is dissolved, with the exception
of black flakes of copper sulphuret.  The solution,
filtered while warm, deposits, when cold, a large part
of the copper combination, which had been dissolved.
This deposit, washed and decomposed by sulphuretted
hydrogen, furnishes carnine, which is purified by being
crystallized in water, being deprived of colour, if necessary, by a little animal charcoal. The hydrochloric
mother-liquor which deposited the copper combination
of carnine, having been deprived of its copper by
sulphuretted hydrogen, and then concentrated, gives
first crystals of xanthine hydrochlorate; then, by concentrating the decanted liquor in the cold, crystals of
guanine hydrochlorate, from which the guanine is extracted by precipitating it with an excess of ammonia,
which dissolves the xanthine, and leaves the guanine.
The sarcine is obtained by precipitating by ammonia
and silver nitrate the nitric solution of the first copper
precipitate; by washing with water of ammonia the
dirty-white gelatinous precipitate which is formed, and
then crystallizing it in boiling nitric acid at I20 Baume;
we thus obtain immediately the nitro-argentine combination of sarcine. This is decomposed by sulphuretted
hydrogen; the liquid, when filtered, and concentrated,
has ammonia added to it, which, by concentration,
leaves the sarcine in fineneedle-like crystals.
The pitchy precipitate formed at first, by the addition
of alcohol to the concentrated extract of digested yeast,
is in great part formed of earthy phosphates, tyrosine,
and gummy matter.
The leucine extracted from the yeast after its diges



144            ON FERMENTATION.
tion shows a peculiarity noticed by Hesse. It contains
a quantity of sulphur, which varies within certain limits,
and may attain to 4 per cent. My analyses, made from
very pure products, crystallized several times in alcohol,
and having the appearance of beautiful white nacreous
flakes, gave from 2'93 to 2'I4 per cent. of sulphur. This
cannot be eliminated by long boiling at I 00 C. (2120 F.)
with a mixture of potass and lead hydrate.  The
hydrogen of these leucines have always been found
to be somewhat deficient (9'34 or 9'6 per cent. instead
of 9'9).
It appears that the sulphur forms an integral part of
the molecule, and is not found in the state of a sulphuretted body mixed with leucine. At any rate, repeated
and very careful purifications do not succeed in splitting
up this mixture.
Let us now see what may be the signification of these
results.
All the nitrogenous compounds noticed as being the
products of spontaneous changes in the yeast have
been obtained directly by the splitting up of albumin
and albuminoid substances. (See the chapter on these
bodies.) Their origin is, therefore, not doubtful; they
are formed by the decomposition of certain insoluble
proteids in the yeast, and by a chemical process similar
to that which takes place in the animal tissues; for it is
impossible to mistake the great analogy of composition
which exists between the extract of digested yeast and
extracts obtained from animal tissues. This chemical
phenomenon is also comparable to those which are
observed by causing dilute warm sulphuric acid, or the
alkalis, such as potash and barytes, to react, under cer



FUNCTIONS OF YEAST.              145
tain conditions, on proteids; we thus obtain, in fact,
similar products.
Albumin, obtained in such large proportions in the
extract of digested yeast, must be one of the results of
the splitting up; it is probable that yeast has no action
on the one substance among proteids which resists so
strongly chemical agents.
This opinion is corroborated by the interesting observation of M. Gautier, who proved that fibrin splits up,
under the influence of salt water, into albumin and
another albuminoid principle. Zymase, or alterative
ferment, which exists in large proportions in the extract
of softened yeast, also represents one of the substances
derived from the decomposition of insoluble proteids.
I have before said that we succeed, by direct analysis,
in separating an uncrystallizable nitrogenous principle
of a sweet taste. This substance resembles in its properties the hemiproteidin on hemialbumin formed by
the action of boiling dilute sulphuric acid on albumin.
As to the tyrosine, leucine, butalanine, the sarcinic
bases, is evident that they are the directly der:ved products of albuminoid substances.
-With respect to the proceeds of the sugar which
furnish the alcohol and carbon dioxide of the spontaneous
fermentation of yeast, and of the gummy matter, their
source has not been ascertained with as great certainty.
It is generally admitted, in accordance with the views
of Payen and Schlossberger, that washed' yeast is composed of cellulose and insoluble proteids.
The question whether the sugar and gum which show
themselves in notable proportions in the extract of yeast,
when digested and washed, ought to be considered as
the products of the physiological decomposition of the




[46            ON FERMENTATION.
albuminoid substances, or the results of a transformation of cellulose, can be decided only by a series of quantitative experiments. The following results give only
an approximate solution, but they prove, at least, that
the greater part, if not the whole, of the solid principles
contained in the extract of yeast when washed in cold
water and digested, must be derived from proteids.
Ioo grammes of fresh yeast contain 30 grammes of
solid matter. This includes 9'28 per cent. of nitrogen;
whence it results, that Ioo grammes of fresh yeast
contain 2'78 grammes of nitrogen.
ioo grammes of fresh yeast, washed with boiling
water until it runs off clear, leave 20 grammes of solid
matter.  This contains IO'I7 per cent. of nitrogen.
The result is that Ioo grammes of yeast, washed with
boiling water, contain 2o03 grammes of nitrogen.
ioo grammes of yeast, digested for fifteen hours at
350 C. (950 F.), and then washed with boiling water,
contain I2-5 grammes of solid matter, yielding 7'55 per
cent of nitrogen.
0oo grammes of yeast, digested and then washed
with boiling water, contain, then, 70'94 grammes of
nitrogen. The loss of nitrogen arising from the digestion and washing is I182 - 0'75 - I'o7.
The loss of solid products due to digestion and subsequent washing amounts to I7'5 - Io  7'5.
But proteids contain, on an average, I5'5 per cent. of
nitrogen; I'o07 gramme of nitrogen correspond to 69
grammes of proteinic matter. Consequently, out of 7'5
grammes of the principles which have become soluble by
the process of digestion, 6'9 grammes or -4 are derived
from albuminoid substances.
On the other hand, a direct quantitative estimate of




FUNCTIONS OF YEAST.             I47
the nitrogen in the dry extracts gave I2'5 per cent. of
nitrogen. It is, then, evident that the yeast must have
given up part of its non-nitrogenous elements; this last
calculation would lead us to the proportion of x ploteids,
and 5 hydrocarbon matter, if we take no account of
the elements of water which are necessarily united with
the albuminoid matter at the time of its splitting up
and transformation into bodies such as leucine.  By
making an approximate estimate of this water of hydratation, we should find {o of albuminoid and -rl of
hydrocarbon matter. It is not very probable that the
gum has an albuminoid origin; it could at most be
derived only from the decomposition of a substance
analogous to tunicin or chitin.
The principles yielded to water by fresh yeast in a
much smaller proportion are of the same nature as
those which we have found before; it cannot, indeed, be
otherwise. The conditions under which M. Bechalmp
and I carried on our researches necessarily exaggerated
the effects of a continuous cause. As soon as the yeast
finds no more sugar, it reacts on its own elements, and it
is difficult to find yeast which has not been placed in this
situation for a longer or shorter time. It is probable
that all kinds of yeast give reactions of this nature in
the fermenting vats, when the sugar begins to fail. M.
Bechamp says that he has found tyrosine and leucine in
the aqueous extract of all fermentations which he has
examined ad hoc. Before, therefore, we draw too positive
conclusions, we must know whether the fermentations
have been studied immediately after the total decomposition of the sugar.
M. Pasteur has shown us that spontaneous fermentation does not appear till this moment; it would be




148            ON FERMENTATION.
interesting to ascertain whether the formation of nitrogenous excremental substances, such as leucine or tyrosine, takes place during fermentation or no; we should
thus see whether this phenomenon is allied to spontaneous fermentation, or is independent of it.
M. Bechamp, considering the spontaneous fermentation of yeast and its concomitant phenomena (the
production of acetic acid, tyrosine, &c.) has given a
physiological theory of fermentation which may be
shortly stated thus:Yeast, like every living organism, shows phenomena
of two kinds; those of nutrition and assimilation, which
are subordinate, to the presence of its nutritious principles (sugar, nitrogenous compounds, mineral salts).
These various principles, penetrating by endosmose
into the cell, undergo there suitable transformations,
and are converted into tissues of recent formation in
the new cells which are formed by budding. Together
with these phenomena of nutrition, and side by side
with them, other inverse reactions, those of disassimilation, take place, by which the tissues are changed into
excrementitious products, unsuited to the life of the cell,
and these are eliminated. The production of carbon
dioxide and of alcohol are the consequences of this
process, and belong to disassimilating reactions. In this
theory, M. Bechamp develops the ideas of M. Dumas
as to the part played by yeast and other ferments. It
is certain that the production of carbon dioxide and
alcohol without sugar, at the expense of the constituent
elements of the yeast itself, gives some support to the
opinion of Bechamp, whether or no we admit that this
production is the result of the formation of buds, in which
the new cells are nourished at the expense of the old ones.




FUNCTIONS OF YEAST.              149
On the whole, this theory throws but little light on
the question relating to the very essence of the phenomenon. Whether the sugar is decomposed with or without previous assimilation is of little importance; we are
no nearer knowing why it is decomposed, and scarcely
any one now doubts that the decomposition of sugar
is a biological phenomenon. We must now examine
some special points relating to alcoholic fermentation.
It has been long thought that alcoholic fermentation
could take place under two distinct circumstances, according as yeast is added to a solution of pure sugar in
water, or to sugared water containing the nitrogenous
and saline principles necessary for its nourishment. In
the first case, it was thought that the ferment acts without reproducing itself; while in the second case, which is
that occurring in the brewing of beer, it acts and reproduces itself. More than this, Thenard had observed
that, in the fermentation of 20 parts of yeast and Ioo
parts of sugar, there only remain, after all the disengagement of carbon dioxide has ceased, I3'7 parts of
an insoluble residue, which may be reduced even to Io
by fresh contact with sugar. Thus yeast, while exciting
the fermentation of pure sugar, partially destroys itself.
M. Pasteur, on the contrary, admitting it to be
proved by his experiments that the budding and multiplication of yeast are phenomena which, in a constant
manner, accompany all alcoholic fermentation; explains
the formation of new globules in a solution of pure
sugar in water by nutrition at the expense of the nitrogenous soluble substances of the original yeast.  In
this case, whatever soluble nitrogenous aliment there is
in the ferment employed becomes fixed in an insoluble
state in the newly-formed globules. It is certain that




150                ON  FERMENTATION.
yeast formed in a suitable medium, such as the wort of
beer, is gorged with soluble nitrogenous principles, which
it can yield to water, and which are eminently suited to
the nutrition of new yeast.
In order to remove all doubt as to the reality of this
explanation, it was necessary for Pasteur thoroughly to
explain away those of Thenard's results, quoted above,
which seem  to contradict his opinion.
The following table  sums up his observations on the
fermentation of pure sugar with yeast:|             r<i;'  gL' t? 2's       i''
).                           2. _...
A    I00   20000  4626   3230       2 320      5 55       934
C    100   x6o  4604   4385    ne    -         -
D    I00   I0 000  2-3 13    486  I o80     3566 T     C253
E      0 Ioo    3700  2626    965     964      399  I  3C3
F    10oo 0   25 I        00 06    6  I 0     *331    1 1* 33
G     i6    3.I59  0*699   072    mi not deter-  -.
mined.
H      4  - I-474  0326   0 335      - 
I     20    1 878-  0*476  0o590    01Ij3      0o723     0247




FUNCTIONS OF YEAST.               15 1
We see that in the experiments A B C, in which the
weights of yeast in the form of paste is above 15 or 20
per cent. of the weight of sugar, after fermentation less
yeast is collected than is originally put in; these are,
therefore, precisely the conditions of Thenard's experiments, who employed and recommended 20 parts of
yeast in form of paste to Ioo of sugar.
If, on the contrary, the weight of yeast in the form
of paste is equal to or less than Io per cent. of the
sugar, more yeast is collected than had been first
employed (exper. D E F G H I).
And, in every case, by adding the weight of extractive
matter to that of the yeast remaining after fermentation
(having deducted the glycerin and succinic acid which
are in solution in the fermented liquor, and come from
the yeast), we find that the sum always exceeds very
sensibly the whole weight of the original yeast.
The increase amounts to about 12 or I'5 per cent. of
the weight of the sugar decomposed.
The result of this appears to be, that in Thenard's
experiments, the disappearance of yeast was only
apparent. Less was collected because much was used,
and because the soluble part which passes into the liquid
is greater than the weight of the newly-formed globules.
M. Duclaux (Theses de la Faculte des Sciences de Paris,
I865) arrived at results of the same kind.
In Ioo grammes of fermented sugar he foundDry yeast employed.    Dry yeast obtained.
17'32             14'02
8'66              8'5I
4'33              4'78
2'-6              2'58




15 2           ON FERMENTATION.
However, in other trials made by the same author,
the weight of yeast found exceeded that of the yeast
employed, even in the case in which the original proportion exceeded I6 per cent. of sugar. This difference
is attributed by the author to the use at first of a kind
of yeast poorer in extractive principles, and yielding
fewer solubler substances to the water.
The results obtained by Pasteur and Duclaux, in
fermentation conducted on a small scale, have been
confirmed by other experimentalists, such as those of
Mayer, Fitz.
From the whole results published, it seems that we
may conclude that, within certain limits, and with some
variations, the weight of the newly-formed yeast is in
proportion to the weight of the sugar decomposed, and
equal to from I'5 to 3 per cent. of the fermented sugar.
We ought, however, to add, that the information
afforded by commercial experience, by the manufacturers of compressed yeast, do not agree with these
conclusions.
According to Marcher and Schulze, we are able to
obtain 28-6 parts of dry yeast for Ioo of alcohol, or,
which comes to the same thing, I4'66 parts of dried
yeast for Ioo parts of decomposed sugar.
In the fermentation of the must of grapes, the formation of yeast seems much greater in proportion to the
fermented sugar than M. Pasteur's ratio would make it.
M. Pasteur, who carries on with so much success the
manufacture of pure yeast on a large scale, by the fermentation of the wort of beer in a close vessel, obtains
1'5 kilogrammes (3'3 lbs. avoir.) of expressed yeast, which
contains 30 per cent. of dry matter, say 450 grammes




FUNCTIONS OF YEAST.               I53
(99 lbs. avoir.) of dried yeast per hectolitre (22 gallons)
of beer; allowing that this beer contains 8 per cent. of
alcohol, fermentation would have acted on I5'5  kilogrammes of sugar, there would then be 2'9 grammes of
new yeast for Ioo of decomposed sugar; these numbers
do not differ so much as the first, from the results
obtained on a small scale; it is true, however, that in this
calculation, we have taken for the proportion of alcohol
a very high weight which ought, perhaps, to be reduced.
The apparent contradiction between the results of the
laboratory experiments and those of commercial operations, may be explained by admitting, that the development and multiplication of the cells of yeast are not so
much dependent on the quantity of sugar decomposed,
as the laboratory experiments seem to indicate, that
they may vary within very wide limits, according to the
composition of the medium in which the operation takes
place.
This explanation seems to be corroborated by the
following experiments of Mayer (Landwi, Versuchsz,
vol. I6, p. 304).
The author fermented two portions of the boiled
must of grapes of the capacity of I90 cubic centimetres
(I I-59 cub. in). To one, he added imperceptible traces of
wine-lees (Saccharormyces c/lips.), and sowed in the other
a little beer-yeast. When the sugar had disappeared,
which required several weeks, he foundALCOHOL  CORRESPONDING   YEAST
FORMED. SUGAR DECOMPOSED. FORMED.
gr.        gr.         gr.
I. Saccharomyces ellip.. I3'II    25'6       2'38
2.            cerevisiM  15'2       29'7       2'04




154            ON FERMENTATION.
Therefore for 00oo parts of sugar decomposed there were
formedNo. I. 9'2 of yeast.
No. 2. 68,,
numbers much higher than the results obtained in
fermentations under other conditions of the medium.
On the other hand, M. Pasteur found that in fermentations, in the presence of nitrogenous and mineral
nutritive matter, nearly i per cent. of the weight of
sugar was formed in yeast and soluble products; a little
less, therefore, than when we operate with yeast completely formed, and pure sweetened water.
Thus, to 9'899 grammes of pure sugar-candy, dissolved
in a sufficient quantity of water, were added 20 cubic
centimetres of boiled and filtered water with which fresh
yeast had been washed (containing 0o334 grammes of
albuminoid and mineral matter), it was then filled up to
Ioo cubic centimetres and a trace of fresh yeast added.
The sugar had disappeared at the end of eleven days;
o0152 grrammes of dry yeast were collected. The nitrogenous remainder left after the evaporation of the liquid,
when the glycerin and succinic acid had been removed,
weighed o0260 grammes.
Thus, o'334 grammes of nutritive matter were
employed, and o'I52 grammes of yeast + o026o grammes
of soluble nitromineral matter was found; in all, o'412
grammes. The difference is 0o78 grammes.
When pure sugar ferments with a limited quantity of
yeast, this becomes exhausted, and at last becomes unfit
to effect fresh decompositions of sugar, in the absence
of soluble nutritive materials. This ought, in fact, to
be the case.  The amount of the nutritive materials




FUNCTIONS OF YEAST.            I55
which are soluble, or become so during the fermentation,
and which are contained in the limited yeast, being
gradually utilized for the formation of fresh globules,
there ought to come a time when the yeast will be
destroyed, for want of nutriment. This phenomenon
of the nutrition of the yeast at its own expense, does
not exclude the presence of nitrogenous matter in the
no longer fermenting saccharine liquid, for it may
happen that a part of the nitrogenous principles eliminated by the yeast are unsuited to its nourishment, and
only represent excremental products.  Leucine and
tyrosine seem to belong to this order of products; in
fact, some direct experiments of Mayer have proved
that they are unsuited to development.
In a fermentation, in which I'I98 grammes of washed
yeast (the weight of dried matter, containing 9'77
grammes per cent. of nitrogen) had been employed to ferment Ioo grammes of sugar, I'745 grammes of yeast were
collected after fermentation; this contained'55 per cent. of
nitrogen; the extractive residuum of the fermented liquid,
when deduction had been made of the succinic acid and
glycerin, weighed o06 grammes and contained 3'8 per
cent. of nitrogen. Under these conditions, the yeast
was exhausted-that is to say, the soluble nitrogenous
principles contained in the liquid had become unfit for the
development of new cells, and represented excremental
products. We know that among the products left undetermined by M. Pasteur, we must place leucine and tyrosine.
This experiment shows us the cause of the diminution of nitrogen in the yeast after fermentation. On the
one hand, the total weight of yeast has increased by the
addition of principles (cellulose) furnished by the sugar,




I156           ON FERMENTATION.
and not nitrogenous; on the other hand, the original
yeast has yielded to the liquid soluble nitrogenous products which have become unfit for its nutrition. Thus
-the apparent disappearance or diminution of the nitrogen of the yeast during fermentation with pure sugar
is explained in a very natural manner, a phenomenon
which had so puzzled Thenard, and which Dbbereiner
had wrongly thought might be attributed to the formation of ammoniacal salts.  We have already seen by
Pasteur's very careful experiments, that the ammonia
of the liquid has a tendency rather to disappear than to
increase during the fermentation.
According to Dubrunfaut (Comp. Rend., July, I87I),
the wort of beer, which, under ordinary circumstances,
reproduces sevenfold the weight of the yeast employed,
is so rich in matter reproductive of ferment, that it is far
from being exhausted by this process. An addition of
sugar excites an increase proportional to the sugar
supplied. If it has not been employed in excess, the
normal condition of the ferment with respect to nitrogen
has not changed; if the contrary has been the case, the
amount of nitrogen lies between that of fertile and of
barren yeast. All kinds of yeast, the results of fermentation other than that of beer made from malt, are in
this condition; they give an amount of nitrogen varying
between o-Io and o05.
Beer-yeast, notwithstanding the loss of weight sustained in washing, preserves its proportion of nitrogen,
while its richness in salts decreases from *Io to'02.
However often it may be washed, we can never obtain
water quite free from albuminoid and saline matter; which
proves that the yeast continues to live, even in pure water,




FUNCTIONS OF YEAST.                  I57
and to exercise there its vital functions on its own
substance, as animals  do  which  are condemned to
starvation.
The ash derived from  water in which yeast has been
washed, is always alkaline; that of the washed yeast
is acid; which induces us to believe that there is an
ammonio-magnesian phosphate present in the yeast,
which, in consequence of its insolubility, remains in the
washed ferment.  M. Dubrunfaut has, like M. Pasteur,
ascertained the constant disappearance of a certain
portion of the ammonia of the medium, as a concomitant
of the reproduction of ferment; but at the same time,
the ash becomes more acid. The addition of ammoniacal salts to fermentation going on under unfavourable
conditions, has the effect of facilitating the fermentation,
and preserving the proportion of nitrogen, o0I of the
yeast.*
M. Dubrunfaut performed a series of experiments to
ascertain the part played by different mineral salts in
the fermentation of sugar, and in the development of
yeast. He made use for this purpose of solutions of
pure sugar at Io per cent., and added to them various
mineral salts and yeast in the form of paste, all taken
at a weight 0o05 of the weight of the sugar. The ferment
employed, therefore, only represented in dry matter o'oI
of the weight of sugar. He tried, I. potassium nitrate;
2. ammonium sulphate; 3. potassium sulphate; 4. calcium phosphate; 5. magnesium sulphate; 6. calcium
* These last experiments would have been more properly placed in the
chapter concerning the influence of salts and nitrogenous matter, on the development of yeast. We only give them here as a supplement, which the
reader will be kind enough to read for himself in its proper place.




158               ON FERMENTATION.
sulphate; 7. sodium sulphate; 8. a wort without mineral
salts, to serve for comparison:FRACTIONS OF SUGAR DECOMPOSED IN THE SAME TIME.
Sodium sulphate.          0'52.
Calcium,,.            0' o62.
Magnesium,,                        0.   0o73.
Calcium phosphate...  o.8o.
Potassium sulphate..'88.
Ammonium,,.. o'94.
Potassium nitrate.. I'o
for complete and perfect transformation without production of acid.
All these salts gave higher results than those of the
wort used for comparison, which only transformed o'50
of the sugar into alcohol.
The nitric acid, in this experiment, entirely disappeared. It appears, then, from Dubrunfaut's observations, that yeast forms no exception with respect to the
facility with which nitric acid is assimilated.




CHAPTER VI.
ACTION  OF  VARIOUS  CHEMICAL  AND  PHYSICAL
AGENTS ON ALCOHOLIC FERMENTATION.
IT has long been known that certain chemical compounds, especially those which coagulate albuminous
substances and disorganize the tissues, or which, by
their presence in sufficient quantities, are incompatible
with life, are opposed to fermentation; such are the
acids and alkalies in suitable proportions, silver nitrate,
chlorine, iodine, the soluble iron, copper and lead, salts,
tannin, phenol, creosote, chloroform, essence of mustard, alcohol when its strength is above 20 per cent.,
hydrocyanic and oxalic acids, even in very small quantities.
An excess of neutral alkaline salts or sugar acts in
the same manner, by diminishing in the interior of the
cell the minimum quantity of water which is necessary
for the manifestation of its vital activity.
The red mercury oxide, calomel, manganese peroxide,
the alkaline sulphites and sulphates, the essences of
turpentine and of lemon, &c., also interfere with, and
destroy, alcoholic fermentation.
Phosphoric and arsenious acids are, on the contrary,
inactive. We owe to M. Dumas (Ann. Chim. Phys.




I 60           ON FERMENTATION.
5th series, vol. 3, I874, p. 8I) a very elaborate work on
the question which we are now considering; we will
give here the principal results. T.his illustrious chemist
first proved, by many experiments, that the fermentation
of sugar under the influence of yeast can be studied
like any regular phenomenon, which, when subject to
determinate disturbing forces, would be able to show
their influence with precision.
Thus, at 24~ C. (about 76o F.) 20 grammes of yeast
decompose I gramme of sugar (glucose) in twenty-three
or twenty-four minutes, (the mean of several trials
agreeing with each other).
In another series of experiments it was found that
IOO grammes of yeast decompose, at the same temperature, I gramme of glucose in twenty-four minutes.
Thus, yeast acts on glucose with the same rapidity,
in the proportion of 20 grammes of yeast to one of
glucose, as of Ioo to I.
Experiments made on the same yeast, with cane-sugar
and glucose separately, showed that the destruction of
I gramme of cane-sugar by 40 grammes of yeast had a
maximum duration of thirty-four minutes, while one
gramme of glucose required but sixteen or seventeen
minutes.
If we mix some beer-yeast with water, and add to
similar portions of the liquid, containing, for example,
I50 cubic centimetres of water and Io grammes of
yeast, quantities of sugar represented by 0o5 grammes,
I gramme, 2 grammes, 4 grammes, we find that the
time necessary for the destruction of the sugar is
exactly proportional to its quantity; or, in other words,
that under identical circumstances the duration of the




ACTION OF CHEMICAL AND PIHYSICAL AGENTS. I6I
fermentation is proportional to the quantity of sugar, it
being understood that the yeast is in excess.
By taking, then, for the length of the co-ordinates
measured along the axis of' the abscisswe the quantities
of sugar, and for the co-ordinates measured along axis
of the ordinates, the number of minutes necessary for
the disappearance of the sugar, we obtain a straight
line.
M. Dumas estimates approximately, according to
his results, that to decompose I gramme (I5-4 grains)
of sugar in one hour, it requires 400 milliards
(400,000,000,ooo) cells, supposing thent all to be
active.
The fact discovered by Dumas, that beyond a certain
limit an excess of yeast does not promote the fermentation of  a given quantity of sugar, is extremely important.
In fact, if the decomposition of sugar is a biological
phenomenon which takes place in the interior of each
cell into which the sugar penetrates; if it ought to be
attributed to the nutrition of the globules of yeast, it
seems that the rapidity of the decomposition of the
same quantity of sugar ought to increase indefinitely
with the number of cells which are in action, in the
same manner as in a meal, the quantity of food consumed increases with the number of guests.
Teinperature.-We have to  determine three data
relating to alcoholic ferment, with respect to heat:I. What are the two limits, maximum and minimum,
compatible with the life of the ferment? 2. At what
temperature is alcoholic fermentation, that is to say, the
activity of the ferment, the greatest? The lower limit
8




I 62           ON FERMENTATION.
compatible with the life of the Saccharogl'ces is not yet
determined with precision.
It is only towards So or Io~ C. (about 460 to 50o F.)
that the- cell shows its chemical energy by sensible
reactions (fermentation, absorption of oxygen, &c.), but
it may be cooled with impunity towards zero (32c F.), and
even below, if we are careful to subject it, after congelation, only to very slow progressive variations, to
re-melt the frozen water in which it is contained. The
higher limit depends on whether we operate on yeast
previously dried or still damp.
Thus, yeast dried carefully may be heated to I000
(2120 F.) without losing its vitality.
Damp yeast would lose its activity (Mayer) towards
530 C. (about 1280 F.), or at least at a temperature less
than 60~ C. (I400 F.).
As to the most favourable conditions of temperature
for good fermentation, they appear to range between 25'
and 30" C. (770 and 860 F.), varying within narrow limits,
according to the nature of the medium and that of the
ferment; even near zero (320 F.) the production of
alcohol by yeast is not absolutely destroyed.
Electricity.-The sparks of a Holtz machine, or those
of an induction coil passing through water containing
yeast, modify neither its power of changing cane-sugar
into glucose, nor its activity, as an alcoholic ferment.
Gas-ligfht.-The process of fermentation is more slow
in the dark. M. Dumas placed beer-yeast of the consistence of thick broth in flasks respectively filled with
oxygen, hydrogen, nitrogen, carbon oxide, nitrogen protoxide, proto-carburetted hydrogen. At the end of three
days, this yeast, placed in contact with sugar, acted in




ACTION OF CHEMICAL AND PHYSICAL AGENTS. I63
the same way as yeast kept in the open air, and the
microscope showed no change.
Alcoholic fermentation is more slow in a vacuum.
Sulphzir.-The presence of flour of sulphur, even in
equal proportions to that of the dried yeast, did not
sensibly interfere with alcoholic fermentation, as had
been predicted; only the carbon dioxide which was
disengaged contained I or 2 per cent. of sulphuretted
hydrogen.
Actionz of Acids. - Yeast is always acid.  If we
neutralize the free acid which is found in it by limewater, we find that in about five minutes the acidity
reappears. We are therefore obliged, in order to maintain a continuous neutrality, to add for each gramme of
yeast under examination, a quantity of lime equivalent
to'00oo3 grammes of normal sulphuric acid. M. Dumas
wished to ascertain whether the acidity of yeast can
be increased, without its power being impaired, and
whether the specific nature of the acids exercises any
influence on the result.
Experimenting on sulphuric, sulphurous, nitric, phosphoric, arsenous, boracic, acetic, oxalic, and tartaric
acids, he added to a mixture of sugar, water, and yeast
(5 grammes yeast, Io grammes sugar, 25 grammes
water) quantities of each of these acids respectively,
first equivalent, then tenfold, and a hundredfold of the
acid power of the yeast.
The addition of one of these acids, even in feeble
proportions, neither hastened the commencement nor
the end of the fermentation. It often arrested the decomposition of the sugar. In general, when we add a
quantity of acid equal to Ioo times the weight of acid




I164           ON FERMENTATION.
contained in the yeast, fermentation does not take
place. It was necessary to add 200 equivalents of
hydrocloric or tartaric  acid to attain  this result,
although ten equivalents of these acids are sufficient to
render the fermentation slow and incomplete.
Actiorl of bases.-M. Dumas having set going at the
same time eight experiments of fermentation, differing
from each other only by increasing additions of ammonia
firom o' (quantities equivalent to o, I, 2, 3, 4, 8, I6, 24
times the acidof the yeast employed), observed that in
the first five vessels the fermentation was equal. It is
only in proportions equal to eight, or even sixteen times
the acid of the yeast, that it begins to proceed more
slowly; but at the end of some hours the yeast has
again become acid. In the flask containing a quantity
of ammonia equivalent to twenty-four times the acidity
of the yeast, no fermentation takes place.
Yeast seems to possess the power of producing or
exhaling an acid which neutralizes the bases in contact
with it; but this power is limited. In fact, by adding
slaked lime or calcined magnesia in quantities equal to
half the weight of the yeast there will be no fermentation. The oxides of zinc, of iron, or even litharge, have,
on the contrary, no influence. The alkalies tend to arrest
fermentation, but do not destroy it, except when the
proportion is rather large. Sodium carbonate has no
effect, unless we raise the proportion to 70 grammes for
Io grammes of yeast, and 200 cubic centimeitres of
sweetened water containing J-  of sugar. Magnesium
subcarbonate has no action.
Action of salts.-M. Dumas allowed yeast to remain
three days in saturated solutions of various salts, and




ACTION OF CHEMICAL AND PHYSICAL AGENTS. I65
then tried its action on the solution of sugar-candy;
his experiments led him to class salts in four categories,
viz:Ist. Those under whose influence the fermentation of
sugar is entire, and more or less rapidPotassium sulphate            Sodium phosphate,,   chloride,,  sulphate,,   phosphate,,  bisulphate,,   sulphovinate.   pyrophosphate,,   sulphomethylate,,  lactate,,   hyposulphate         Ammonium pihosphate
Calcium hyposulphate          Magnesium sulphate
Potassium formiate            Calcium chloride
tartrate,,   phosphate
itartrate,,  sulphate,,   sulplhocyanide       Strontium chloride,,   ferrocyanide         Alum,    ferricyanide         Zinc sulphate
Cupric sulphate at jj,
2nd. Those under whose influence there is partial
fermentation of the sugar, more or less retardedPotassium bisulphite          Borax,,    nitrate             White soap,,    butyrate            Ammoniumn nitrate,,   iodide,,    tartrate
arseniate            Seignette salt
Sodium sulphite               Barium chloride
h,,   yposulphite          Ferric sulphate.: 
Potassium hyposulphite        Magnesium sulphate u'r.
3rd. Those under whose influence there is greater or
less change of the sugar, without fermentation



i65             ON FERMENTATION.
Potassium nitrate         Marine salt,,   chromate         Sodium acetate
bichromate       Sal ammoniac
Sodium nitrate            Mercuric cyanide.
4th. Those under whose influence there is neither
change nor fermentation —
Potassium acetate         Sodium monosulphide.,,   cyanide
Thus among the salts studied, there are some which
favour fermentation, up to a certain point, or at least
allow it to run its entire course without interruption;
such as potassium  tartrate.  There are others which
retard fermentation and render it incomplete, the action
being arrested, when the liquor still contains much
changed sugar in contact with it.
There are some which do not allow it to be set up,
although the sugar has been partially changed.
There are also some, which not only do not allow
fermentation to be set up, but which even oppose the
change of the sugar.
Strychnine has no effect on the properties of yeast.




CHAPTER  VII.
CAN NOTHING BUT ALCOHOLIC YEAST EXCITE
ALCOHOLIC FERMENTATION?
WE must now enter on a question of great importance
ill the history of fermentation in general. Admitting
with M. Pasteur, and all those who have preceded and
followed him in this inquiry, that alcoholic fermentation,
and the other phenomena of the same order, such as
lactic, butyric, fermentations, &c., are palpable manifestations of certain physiological functions of ferments, or
organisms of an inferior order, we may ask, whether
the power of resolving glucose into alcohol and carbon
dioxide, or of changing it into lactic acid, and that
again into a mixture of hydrogen, carbon dioxide, and
butyric acid, belongs, for each special fermentation, only
to a single organism, to a single ferment, or, at least,
to species very nearly allied, as we have seen in the
species of the genus Sacc/aronzyccs; or whether these
reactions are the result of cell life in general, when
organic cells are placed under special conditions. On
this hypothesis, ferments would have no advantage over
other living cells, than that of showing these manifestations in a more energetic and intense degree.
It is easy to perceive, a' priori, what new horizons
would be opened for biological chemistry by such an




I68            ON FERMENTATION.
idea, if it were correct, and what a vast field of experiments would be offered to chemical physiology.
There is no doubt that this thought must have
presented itself involuntarily, as we may say, to the
minds of men of science accustomed to reflect on these
delicate questions of the reactions in living organisms;
but M. Pasteur has the honour of first clearly expressing
it, and supporting it by positive experiments. (Pasteur,
Comp. Rend. de l'Acad. des Sciences, vol. 75, p. 784.)
These experiments were undertaken in order to prove
that thle alcoholic fermentation of sugar may be excited
by other organisms than the cells of Saccharolzyccs,
and notably by the elementary cells of larger plants,
such as are found in fruits, leaves, &c. The researches
of M. Lechartier and M. Bellamy, on the alcoholic fermentation of fruits (Comp. Rend. de l'Acad., I869, and
I872, vol. 75, P. 1,203; I874, vol. 79, p. 949, and I,oo6),
have been directed to the same end, and lead, as we
shall see, to this important consequence, that the
elementary organs of plants in general are endowed,
though in a less degree than the cells of yeast, with the
property of exciting alcoholic fermentation.
Analogous facts had been observed by other experimentalists; thus, M. Berard informed us, as early as
I821, that when fruits are placed in air, or in oxygen
gas, a certain volume of this gas disappears, whilst at
the same time there is a formation of a nearly equal
volume of carbon dioxide. If these fruits are left, on
the contrary, in carbon dioxide or in any other inert
gas, there is still a formation of carbon dioxide in a
notable quantity, "as if by a sort of fermentation."
M. Fremy had also shown that when we leave barley




ALCOHOLIC FERMENTATION.             I 69
in a solution of sugar in water, an incontestable intracellular fermentation takes place in the interior of the
corn.  (Comp. Rend. de l'Acad. des Sciences, vol. 75,
p. 976 and I,o6o.)
The same investigator observed the production of
alcohol and carbon dioxide in the interior of fruits,
such as pears and cherries; but guided by theoretical
ideas of fermentation, which we will presently explain,
he did not give to his interesting experiments the
interpretation which they may receive.
M. Fremy admits that ferments are formed under the
influence of the living organism. "Like all organisms
in process of development," said he, " alcoholic ferment
may present itself under the most various forms; it
already exists, but in an imperceptible state, in the
expressed juice of the grape, which appears to be
transparent; it soon shows itself as very attenuated
microscopical corpuscles; then undergoing a fresh development, it is precipitated at the bottom of the liquid,
in the well-known form  of grains of yeast.   I have
frequently examined with the microscope the juice and
the parenchyma of fruits, before or after their fermentation, and I affirm that I have found an innumerable
quantity of corpuscles which have all the appearance
of organic ferments."
Thus we see, from this extract, that the intracellular
fermentation of the fruit is not an immediate consequence of the biological functions of the living cell;
there is an intermediate principle, the ferment, produced
by the organism, which excites'the decomposition.
HIe ought, in a historical point of view, to mention
that the experiments of M. Lechartier, and M. Bellamy,




I70             ON FERMENTATION.
preceded those of M. Pasteur; we will here mention
the results obtained by these observers.
First, HM. Lecharlier's Researczces. -This observer
placed fruits (pears, apples, lemons, cherries, chestnuts,
medlars, potatoes, grains of wheat, linseed, currants) in
test-jars, connected with smaller test-tubes, placed over
a mercurial trough. Under these conditions, the whole
of the oxygen of the confined air in which the fruits
are placed is absorbed. This absorption is accompanied
and followed by a considerable production of carbon
dioxide gas. The disengagement of carbon dioxide is
generally divided into two distinct periods; in the first,
after the absorption of the oxygen gas of the air which
remained in the test-glasses, the disengagement of
carbon dioxide proceeds, at first, in a regular and
uniform manner, then.it slackens, and stops for a
certain time, to be afterwards renewed with an increasing rapidity, greater than that observed during the
first period. This continues for several months. At
this time, if we have taken the precaution to experiment
on fruits, isolated from each other, and kept from contact with the sides of the vessel containing them; if,
besides, we have been careful to prevent any deposit of
liquid on the surface of the fruit, we may ascertain the
production of notable quantities of alcohol, easily
separated by distillation, after the fruit that has been
experimented on has been crushed into pulp; and
what is more, a careful microscopical examination of the
parenchyma reveals no trace of alcoholic ferment.
Thus, on November I2, two pears, one weighing I57
grammes, and the other 125, were suspended separately, each in a glass jar, well corked, and provided




ALCOHIOLIC FERMENTATION.            171
with a tube by which gas might escape. Calcium chloride had been previously placed at the bottom of the
jars, to maintain around the fruits an atmosphere unsaturated with the vapour of water.
The test-jars were opened on July 19: 1,762 cubic
centimetres of gas, and 2'62 grammes of alcohol were
collected. The pears had preserved their colour, their
skin was wrinkled, but not damp. Their consistence
and -smell were like those of mellow pears. They had
lost together I34 grammes of water, yet they still contained it in the proportion of 69 per cent. of their
weight.
Microscopical observations, made at different distances from the centre, could discover no alcoholic
ferment.  The disengagement of carbon dioxide did
not take place in a regular manner; from March 3 to
April 8, there were only 28 cubic centimetres evolved;
and from April 8 to July I9, there was absolutely none.
The existence of alcoholic ferment within the pear is
incompatible with all -cessation of activity during so
considerable an interval of time. This fact, therefore,
corroborates the negative results obtained by means of
microscopical examination.
In other experiments, arranged in the same manner,
but continued much longer, M. Lechartier and M. Bellamy saw the disengagement of carbon dioxide resumed
with increasing activity, after a shorter or longer period
of cessation. In these cases, we can clearly ascertain
the presence of ferment. We may suppose that the
spores and germs of ferment, which exist on the surface
of all fruits, as now seems incontestable, have been able
to penetrate into the inside of the fruit by means of




172            ON FERMENTATION.
artificial fissures, and meeting there an appropriate
medium, have budded and set up the alcoholic fermentation due to ferment.
The different fruits mentioned above all gave analogous results, and the experiment before described
may serve as the type of the rest, which were very
numerous, and were published by the authors; we will
abstain from further allusion to them here.
The cessation of the disengagement of carbon dioxide
may last a long time; thus, pears have been preserved
inert, at the season of the year when the heat is
greatest, during a time varying from 3I to 272 days;
and when an end was put to the experiments, there
was no reason to suppose that this state of things
would be modified.
After this internal action, the fruits had undergone
great modifications.
(I.) As soon as the fruits were exposed to contact
with the air, they become brown throughout their whole
mass, like fruits that have become "sleepy " or rotten.
The leaves assume the colour and aspect of dead
leaves.
(2.) The cellular tissue is either partly or completely
disaggregated. Thus "duchesse" pears at the end of
a year, were like a mass of syrup covered with a skin.
(3.) A fruit which has lost its activity does not regain
it, even after being placed again in contact with the
air.
(4.) The germ contained in the fruit participates in
the decay, and the seed loses its property of germination.
It appears then, as these writers say, that at the




ALCOHOLIC FERMENTATION.             I73
moment when fruits, seeds, and leaves are detached
from the plant which bears them, life is not extinct in
the cells of which they are composed. This life goes
on, sheltered from the air, consuming sugar and producing alcohol and carbon dioxide. The moment when
carbon dioxide ceases to be formed, is that at which
all the vitality of their cells is destroyed. Fruits, seeds,
and leaves may then remain for an indefinite period in
an' inert state, if no organic ferment is developed in the
interior.
As to beet-root and potatoes, an especial fact has
been observed which ought to be mentioned.
The whole of the phenomena are the same as with
apples and pears; when the period of cessation arrives,
no alcoholic ferment is observed, but in the acid liquid
which impregnates the mass of their softened or disaggregated tissues, we find bacteria of different sizes,
and yet we can detect no disengagement of carbon
dioxide.
Secondly, M. Pasteur's Experimenlts. - The experiments made by M. Pasteur, while they verify the preceding conclusions, were especially intended to establish
particular views respecting fermentations, and a general
theory of these biological manifestations. The moment
has now arrived to allude to them, and discuss them.
We cannot do better than quote the exact words of the
author (Pasteur, Comp. Rend. de l'Acad. des Sciences,
vol. 75, p. 784, et seq.).
"I have been inclined for a long time to consider
fermentations, properly so called, as chemical phenomena, co-relative with physiological actions of a peculiar nature. Not only have I shown that their ferments




I74             ON FERMENTATION.
are not dead, albuminoid matter, but actual living
organisms; I have also excited the fermentation of
sugar, lactic acid, tartaric acid, glycerine, and, in more
general terms, of all fermentable matters, ins viiedia
exclusively mineral, an incontestable proof that the
decomposition of fermentable matter is correlative with
the life of the ferment, that it is one of its essential
aliments; for instance, under the conditions to which I
refer, it is impossible, that in the constitution of the
ferments which arise, there can be a single atom  of
carbon which is not taken from the fermentable matter.
"That which distinguishes the chemical phenomena
of fermentations from a number of others, and particularly from the functions of ordinary life, is the fact
of the decomposition of a weight of fermentable matter
much greater than that of the weight of ferment in
action.
"I have long suspected that this peculiar character
may be allied to that of nutrition without contact of
free oxygen. Ferments must be living organisms, but
of a peculiar nature, in this sense, that they have the
property of exercising all the functions of their life, not
excepting their multiplication,* without necessarily employing the oxygen of the atmospheric air.
" Let us call to mind those curious infusoria which
cause butyric or tartaric fermentation, or certain kinds
of putrefaction, and which not only are able to live and
multiply without contact with oxygen, but which perish
and cease to excite fermentation if we dissolve this gas
in the medium in which they feed. This is not all.
By careful experiments, made with the yeast of beer,
Not that of sporulation. Note by M. SCI-iVTZENBERGER.




ALCOHOLIC FERMENTATION.              I75
I have shown, that if the life of this ferment were carried
on partially by the influence of free oxygen gas, this
little cellular plant lost, in proportion to the intensity
of this influence, a part of its fermenting character, that
is to say, that the weight of yeast, which is produced
under these conditions during the decompositionl of sugar,
ilicreases progressively, a nd  approaches the weight of
decomposed sulgar, in exact proportion as its life goes on
in the presence of increasing quantities of free oxygenl
gas.
"Guided by all these facts, I have been gradually led
to look upon fermentation as a necessary consequence
of the manifestation of life, when that life takes place
without the direct combustion due to free oxygen.
"We may partially see, as a consequence of this
theory,. that every being, every organ, every cell, which
lives, or continues its life, without making use of atmospheric air, or which uses it in a manner insufficient for
the whole of the phenomena of its own nutrition, must
possess the characteristics of a ferment with regard to
the substance which is the source of its total or complementary heat. It.appears that this substance must
necessarily be oxygenated and carbonated, since, as I
stated just now, it serves as food to the ferment.  I
have just brought to the support of this new theory,
which I have already several times brought forward,
though with some hesitation, since the year I86I, some
new facts which will now, I hope, cause it to be received."
In a saccharine liquid, suited to the nourishment of
ferments, contained in a vessel which allows an artificial
sowing, while it prevents the spontaneous sowing of




176              ON FERMENTATION.
aerial germs, M. Pasteur placed on the surface a trace
of pure Mycoderma vii.o*  On the following days the
growth covers by degrees all the liquid, under the form
of a continuous film. We can easily ascertain that,
under these conditions, the development of the Mycoderima gives rise to an absorption of atmospheric
oxygen, which is replaced by an almost equal volume
of carbon dioxide, and that, on the other hand, no
alcohol at all is formed.
M. Pasteur having  already pointed  out that the
lfycoder-za viizi has the property, when it grows on the
surface of an alcoholic liquid, of burning the alcohol
already formed, yielding not acetic acid or alde oyde,
but water and carbon dioxide, it appears to us that we
cannot conclude from the absence of observed alcohol
in the experiment, that none has been formed. It is
quite possible that it may have undergone a complete
combustion at the very moment of its elimination. The
volume of carbon dioxide disengaged in the experiments
was nearly equal to the volume of oxygen consumed;
therefore, unless we suppose that the combustion only
acted on the hydrocarbons, or on carbon itself, we
cannot explain this equality; it might be due to the
disengagement of carbon dioxide under the influence
of an alcoholic fermentation, one of the characteristics
of which, alcohol, was burnt as fast as it was produced.
However this may be, let us return to M. Pasteur's
experiment. Let it be repeated exactly under the same
* In a publication previous to this, M. Pasteur had announced that the
sedimentary yeast was produced by the smycoderma vi'i, deprived of contact
with oxygen; M. Pasteur has reconsidered this opinion, which he no longer
thinks thoroughly well founded.




ALCOHOLIC FERMENTATION.            177
conditions, with this difference only, that when the film
is complete, the vessel be shaken, so as to break up
this film, and sink it as far as possible, for the falty
mnatler which accompanies it hinders its being entirely
wetted.
On the next day, and often after only a few hours,
when the experiment is made at a temperature of from
25~ to 300 C. (770 to 86~ F.), we see small bubbles of gas
continually rising from the bottom of the vessel, which
show that fermentation has commenced in the saccharine liquid. It continues during the following days,
although very feebly, and the presence of a sensible
quantity of alcohol in the liquid is readily ascertained.
A careful examination of the cells, or joints of the
submerged Mycoderma, by means of the microscope,
shows that these joints do not reproduce.
This latter fact seems to disagree with the opinion
so many times advanced elsewhere by the writer, that
alcoholic fermentation is always accompanied by the
development and multiplication of ferment, and that
it is a consequence of the formation of buds.
The experiment which we have just described in
detail, quoting almost literally the words of the author,
allows us to conclude with certainty that the Mycoderma
vini may play the part of alcoholic ferment, when it
is placed in contact with a saccharine medium; but we
cannot admit the other consequences that M:. Pasteur
tries to draw from it, as proofs of the theory before
cited.
We may say the same thing of the experiments on
fruits placed entirely, as M. Pasteur has done, in an
atmosphere of carbon dioxide, or of those of M. Le



178           ON FERMENTATION.
chartier, in which this condition was rapidly obtained
by the absorption of the oxygen of the surrounding
air.
They prove irrefragably that vegetable cells can produce alcohol and carbon dioxide at the expense of
sugar, and can act like the yeast of beer, but much less
energetically; but nothing in the description given of
their experiments by M. Pasteur and M. Lechartier
can induce the reader to admit, that this production
of alcohol, this cellular fermentation, only commences
from the very moment when the cell is either partially
or entirely removed from contact with oxygen.,
Even admitting, which the writers quoted do not
assert, that they have directly ascertained the absence
of alcohol in the fruit, as long as it was not kept from
the influence of oxygen, I still have the idea that the
alcohol formed may have been burned, and may have
disappeared under the form of water and carbon
dioxide. Besides this the yeast of beer, the Saccharomyces cerevisiw shows us in the most striking and
incontestable manner, an instance of a cell which
excites alcoholic fermentation, even in the presence of
free oxygen dissolved in the medium; and now that it
is no longer recognized as the sole producer of alcoholic
fermentation, I do not see why we should wish to
preserve for it so exceptional a character as that of
being the only exciter of it in the presence of oxygen.
As to this latter fact, we have only to quote M. Pasteur
himself, in order to prove its reality (Bullet., Soc. Chimique, I86i, p. 62I). "Yeast already formed may bud
and develop itself in a saccharine and albuminous liquid
in the entire absence of air or of oxygen. Little fer



ALCOHOLIC FERMENTATION.            I79
ment is formed, in this case, and a comparatively large
quantity of sugar disappears, sixty or eighty parts for
one of the ferment formed. Fermentation is very slow
under these conditions. If the experiment be made in
contact with air, and over a large surface, the fernentation, is rapid. For the same quantity of sugar that
disappears, much more yeast is produced.  This is
developed under these conditions with energy, but its
character as a ferment tends to disappear. We find, in
fact, that for one part of ferment formed, there will be
only from four to ten parts of sugar transformed. The
part of ferment played by this yeast, nevertheless,
remains, and shows itself very energetic, if we cause it
to act on sugar when not under the influence of free
oxygen gas.
M. Pasteur explains the fact of a tumultuous activity
at the commencement of the fermentation, by the
influence of the oxygen of the air which is dissolved in
the liquids when the action commences.
There seems to be some contradiction between the
conclusions and the facts. On one hand, it is ascertained
that fermentation in the presence of free oxygen is
very rapid and tumultuous; on the other hand, it is
concluded that yeast loses its character as a ferment in
the presence of oxygen. In reality, this contradiction
disappears if we measure, as M. Pasteur has done, the
power of the ferment, not by the quantity of sugar
decomposed by the unit of weight of yeast, in the unit
of time, which would lead us to consequences the reverse of those of this observer, but by the ratio between
the weight of the sugar decomposed, and that of the
ferment formed by budding. According to M. Pasteur,




180            ON FERMENTATION.
fermentation is a consequence of budding, and one of
the phenomena may form the measure of the other.
Thus, by ascertaining that in fermentation without
oxygen, the ratio between the sugar decomposed, and
the yeast formed, is from 60o or o80 to I, while in fermentation, in presence of oxygen, it is only 4 or Io to
I, M. Pasteur thinks himself justified in concluding
that, although in this latter case the fermentation is
more active, the character as a ferment of the yeast has
been lowered.
We may first remark that, in order to measure the
energy of a ferment, M. Pasteur makes use, on this
occasion, of an hypothesis which admits of discussion.
Is it not more natural to measure the energy of a
ferment by the effect which it produces, and, in the
present case, by the quantity of sugar decomposed in
the unit of time by the unit of weight?
That which does come out clearly from the interesting facts observed by M. Pasteur, is, that yeast is able
to multiply in a suitable albuminous saccharine medium,
with or without the presence of free oxygen; the
mutiplication is greatly favoured by oxygen, which
explains why the ratio between the sugar split up and
the yeast formed passes from 80.I, to 4., but at the
same time the activity of the ferment is notably increased, and the fermentation is rapid and tumultuous.
The following experiment is also, in my opinion,
opposed to M. Pasteur's theory. I have carefully
studied, by means of a rapid and exact method of
quantitative analyses of the dissolved oxygen, which
has been described elsewhere, the respiratory phenomena of yeast. I have thus been able to ascertain that




ALCOHOLIC FERMENTATION.             81
in a liquid medium containing oxygen in solution, the
quantity of oxygen absorbed by one gramme of yeast,
in the unit of time, and at the same temperature, was
constant, whether the liquid be saturated or supersaturated with oxygen, or contain a quantity of oxygen
less than is required for saturation. Thus, by diffusing
one or two grammes of yeast in a litre of aerated water,
we notice a sensible decrease of the respiratory power,
only when the proportion of oxygen is less than one
cubic centimetre per litre. On the other hand, these
respiratory phenomena follow the same law in a saccharine and oxygenated medium containing nutritive
albuminoid matter, but the intensity of the respiration
is increased; that is to say, that the proportion of
oxygen absorbed in the unit of time, by the unit of
weight, is more considerable at the same temperature
than in pure water. Finally, I have measured the
alcohol produced, in a given time, in two fermentations
placed identically under the same conditions, and with
this difference only, that in the one oxygen was continually kept dissolved.  The proportion of alcohol
was found, in this case, to be sensibly greater, which
agrees with Pasteur's experiments; during the whole
duration of the fermentation, we have been able to
detect a rapid absorption of oxygen, the intensity of
which corresponded with the general results obtained in
many experiments.
If, as M. Pasteur asserts, the decomposition of sugar
were the result of a respiration of the cells of yeast at
the expense of the combined oxygen, recruiting the
free oxygen, it seems evident that fermentation ought
not to take place, or at least ought to be sensibly




I 82           ON FERMENTATION.
lessened, in the presence of oxygen; but the reverse
of this is the case. The respiratory power of yeast is
independent of the quantity of oxygen contained in the
medium in which it lives; it only varies with the temperature, and the more or less favourable conditions of
nutrition, as well as with the more or less perfect state
of health of the cell.
Yeast placed in a saccharine and oxygenated medium
breathes as actively, or even more so, than in pure
water; it therefore fully satisfies its respiratory power
at the expense of free oxygen, and yet it excites the
fermentation of the sugar as actively, or even more so,
than in a medium which is not oxygenated. The conclusion to be drawn from this is plain; the respiratory
power, and the fermenting power, are two qualities
inherent in the cell of Saccaromzyces, but independent
of each other,; in this sense, that one of these phenomena is not the consequence of the other. The
energy of these two powers depends on conditions more
or less favourable to the nutrition and development of
the yeast; it is not, therefore, surprising to see them
increase or grow less together. In other words, they
are not, as M. Pasteur supposes, the two variable terms
of a constant sum, of which the one vanishes when the
other attains its maximum value; on the contrary, all
the facts tend to prove that these two values grow
wveak, are destroyed, or attain their maxima at the same
time, under the influence of the same causes.  By
weakening the vitality of the yeast, we diminish at the
same time its respiratory power, and its power as a
ferment; on the contrary, by placing the yeast under
the best physical and chemical conditions of nutrition,




ALCOHOLIC FERMENTATION.             183
we see the two factors increase side by side, and attain
their maximum together. Let us, however, hasten to
add, that these two phenomena are necessarily dependent on each other; and we cannot deny this without
contradicting that which we have just said.
In fact, sugar is a pabulum necessary to the development of the cells of the Sacccharoonyces; its presence is,
therefore, indispensable to the nutrition of these cells.
To deprive the yeast of sugar, or which amounts to the
same thing, to place it so that it is impossible for it
to act as a ferment, is to diminish its vitality, and to
place it in a state comparable to starvation; therefore
it is not surprising to see its respiratory power diminish,
all other things being equal.  Reciprocally, to prevent
yeast from breathing free oxygen, is to deprive it
of one of its most important functions, and again to
place it under pathological conditions; and its activity
as a fermnent will be lessened, as M. Pasteur himself
observes.
The following experiments all support this view.
We know, by the researches of M. Bechamp, that the
yeast of beer placed under conditions of starvation,
that is to say, kept from contact with a saccharine
liquid, undergoes a great modification, which has
nothing in common with putrefaction. It grows soft,
and converts its protoplasm, and a part of its contents,
into soluble principles, among which M. Bechamp distinguished leucine and tyrosine, a soluble albuminous
substance coagulable by heat, a ferment producing
change, a gummy substance, phosphates, and acetic
acid; these phenomena are further accompanied by
the production of alcohol and carbon dioxide, as well




84            ON FERMENTATION.
as by a disengagement of pure nitrogen. I have myself
closely examined this singular reaction, and have been
able to show, in the extract of softened yeast, the presence of xanthine, hypoxanthine, carnine, and guanine.
After these first researches, I thought it would be
interesting to study the modifications which took place
in the yeast when thus changed as to its respiratory
functions. For this purpose I made use of the methods
described elsewhere, when treating of the respiratory
intensity of yeast. A known weight of yeast is diffused
in a known volume of aerated water, the original oxymetrical degree of which is known; this degree is again
measured after a quarter of an hour, or half an hour's
respiration. The following facts were observed.
Four hundred grammes of fresh yeast, containing
28'94 per cent. of solid fixed matter, were diffused in
distilled water, so as to formar 2 litres (about 3V  pints).
This homogeneous liquid was divided into ten equal
parts, and introduced into bottles nearly filled, These
bottles were left to themselves in a stove heated to
20~ C. (68~ F.).
I did not consider it to be neces.sary to -employ in
these experiments the creosote water recommended by
M. Bechamp, as I was unacquainted with the nature of
its influence on oxymetrical analyses or on the respiratory power. I therefore ran the risk of seeing some
vegetable productions make their appearance, which
were foreign to the yeast, but the small proportion of
whllich, when compared with the yeast, could not disturb
the results by any very large error.
At the end of three days without nourishment, the
first and second vessels were emptied on a filter which




ALCOHOLIC FERMENTATION.            185
had been previously weighed; that which remained on
the filter, was, in this case, well washed with water at
300 C. (860 F.), until it was exhausted, then dried and
weighed. The weight of the filter being deducted,
there were found, for the 40 grammes of original yeast
5'37 grammes, or I3'4 per cent. of solid fixed matter.
The contents of the second bottle were boiled in water
before filtration; the fixed residuum found was, 5'73
grammes, or I4 per cent.
After digestion for six days, followed by boiling, the
third bottle gave a dry fixed residue of 4'I6 grammes,
or I:0'4 per cent.
After a longer time, the diminution in weight of the
solid insoluble materials of the yeast grew less by
degrees and became insensible. Fresh yeast, washed
with lukewarm water, loses about 9'5 grammes per cent.
of soluble matter. Thus, in five days, the yeast which
had been kept without nourishment, transformed into
soluble products, I8'5 —9 5=9~ grammes of insoluble
matter, which resisted the first washing.
When it has reached this limit, and has been well
washed with warm water till it runs off pure, yeast
shows itself completely inactive with regard to oxygen.
Thus fresh yeast, at 20~ C. (68~ F.), respired, per
gramme and per hour, I'4 cubic centimetre, whilst after
exhaustion, by washing the absorption of oxygen, under
the same conditions, did not exceed'23 cubic centimetre. Yeast digested, without nourishment, for fortyeight hours and well washed, being placed in a pure
saccharine medium, only excited a very slow and
scarcely perceptible fermentation, as M. BSchamp had
before observed.
9




I 86           ON FERMENTATION.
Thus, digestion without nourishment, when sufficiently
prolonged, and followed by thorough washing, affords
us a very simple method of removing, almost entirely and
simultaneously, the two most characteristic biological
manifestations of the yeast-cell. Yet its vitality is not
destroyed, if the experiment has not been carried too far.
If we add to aerated water, containing yeast which
has been digested and washed, and which only acts on
oxygen in an almost imperceptible manner, some water
with which digested yeast has been washed, we shall see
the respiratory phenomenon, at first very feeble, grow
more and more decided, and resume an activity equal,
and even superior, to that of fresh yeast simply diffused
in pure water. The fresh yeast itself respires more
energetically if, instead  of pure water, wue employ
aerated water containing soluble products, derived from
the digestion of yeast.
On the other hand, if we add the soluble products of
yeast to the mixture of digested and washed yeast and
saccharine water, a mixture which gives rise only to an
insensible fermentation, we see, in a very short time, the
decomposition of the sugar manifest itself by a more
abundant disengagement of carbon dioxide.
It follows, from the whole of the facts noticed in this
chapter, that we can no longer regard alcoholic fermentation as the simple result of the biological intervention of a special living organism, characterized by its
form and by the conditions of its development, which
alone has the power to split up some one particular
organic compound in some one particular way.
The various kinds of fermentation appear to us, more
and more, as particular cases of the chemical activity of




ALCOHOLIC FERMENTATION.              18/
living cells, and the ideas enunciated at the beginning of
this work on fermentation find a complete confirmation
in the new facts studied in this chapter.
M. A. Fitz (Soc. Chim. de Berlin, January and February, 1873), has published a long memoir on alcoholic
fermentation produced by the MJucor racrnosuso
The ratio of the alcohol to the carbon dioxide was
found, as the mean of many determinations, to be equal
to  Ioo while in the alcoholic fermentation set up by
yeast of beer, it is _oo.
Distilled alcohol contains a little aldehyde; the
residuum contains succinic acid; while the presence of
glycerine is doubtful.  Dextrin, inulin, and sugar of
milk do not ferment, alcoholically, in the presence of
MifuZcor racem1os0s.
With altered sugar, the fermentation set up by Mizucor
racemosus stops after a short time, so that half of the
sugar remains intact.
The cause of this rapid check in the process of fermentation is merely the sensitiveness of this ferment to
the influence of alcohol.
M. Fitz has, in fact, shown that fermentation ceases
when the proportion of alcohol in the liquid reaches
3'5 to 4 per cent. by weight.  In all other respects the
Mucor racemoslzs behaves like yeast; it alters sugar,
assimilates the nitrogen of ammoniacal salts, and reproduces at the expense of sugar (13 per cent. of the
sugar decomposed).
M. Traube (Soc. Chim. de Berlin, vol. vii. p. 887,
I874) has studied the action of media containing no
free oxygen oni alcoholic'ferments.  His experiments
lead to the following conclusions:



I88            ON FERMENTATION.
The cell of the ferment is not developed in the
absence of free oxygen, even in its most favourable
medium, the must of grapes.
The ferment, in process of development, continues
to increase in suitable media, even in the absence of all
trace of oxygen, as M. Pasteur had already shown.
The contrary assertions of Brefeld are erroneous.
M. Pasteur's theory, according to which the yeast,
in the absence of air, takes from the sugar the oxygen
necessary for its development, is not well founded; in
fact, this development stops long before the greater
part of the sugar is decomposed. Is it from the albuminoid matter that the ferment takes this oxygen in
the absence of air?
Yeast sets up alcoholic fermentation in a solution of
pure sugar in the absence of all trace of oxygen, but
without developing. This is contrary to the affirmation
of AM. Pasteur that fermentation is bound up with the
organization of the yeast, or is a phenomenon correlative to the vital activity of the cells,




CHAPTER VIII.
VISCOUS OR MANNITIC FERMENTATION OF SUGAR.
LIKE all the phenomena which are spontaneously
produced under the ordinary conditions of humanlife,
viscous fermentation has been known for a long time,
for it is developed in certain wines and natural saccharine
juices, as the juice of beetroot, carrots, and onions, as
well as in certain juleps and potions containing sugar
and nitrogenous matter, and it becomes apparent by the
viscosity of the liquid.
This reaction was studied by Braconnot (Ann. de
Chim. (I), lxxxvi. p. 97, 1813), Desfosse (Journ. de
Pharm., xv. p. 604), Franjois (Ann. de Chim. et de Phys.
(2), vol. xlvi. p. 212, I829), Pelouze and Jules Gay-Lussac
(Ann. de Chim. et de Phys. (2), vol. lii. p. 4Io, I833),
Kircher (Annalen. Ch. Pharm., xxxi. p. 337), Tilley and
Maclagan (Philos. Mag., xxviii. p. 12), Boutron-Charlard
and Fremy (Comp. Rend., xii. p. 708, I84I), and also
by Pasteur (Bullet., Soc. Chim., p. 30, Paris, i86i).
Viscous fermentatation is excited, according to
Pasteur, by a special ferment acting on glucose or canesugar, previously altered, transforming them into a.kind
of gum or dextrin, mannite, and carbon dioxide. Gum,
mannite, and carbon dioxide are the only constant products of this reaction; the lactic acid, butyric acid, and




Io90             ON FERMENTATION.
hydrogen, which we see appear at the same time, are
the products of other fermentations due to foreign ferments.
M. Peligot (Traite de Chimie de Dumas, vol. vi.
P. 335, I843) was the first to notice a special ferment,
capable of producing viscous fermentation in saccharine
solutions to which it is added.  Pasteur observed, afterwards, that this ferment is formed of small globules
united as in a necklace, whose diameter varies from
oo002 millimetres ('000047 in.) to'ooI4 ('000055 in.) (fig.
20). These globules, sown in a saccharine liquid containFIG. 20.-ViscoLs Ferment of Wine.
ing nutritive nitrogenous matter and mineral substances,
always give rise to viscous fermentation: Ioo parts of
sugar give about 5 I'09 of mannite and 45'5 of gum;
besides  these, carbon dioxide is disengaged.  These
results may be shown by the following equation:No. I. 2               25 (H2 O) = T2 (Cl2 H+O~ 0~) +
Cane-Sugar.      Water.         Gum.
24 (C6 H14 06) + 12 (CO2) + 12 (H2 0)
Manniteo    Carb. diox.    W'ater.
This gives, for Ioo parts of sugar,Mannite..             5109
Gum...   45'48
Carbon dioxide.   e.    6'18




MIANNITIC FERMENTATION  OF SUGAR.               r91
M. Monoyer (These pour le Doctorat en Medicine,
Strasbourg, I862) proposes to  represent the production of mannite and of gum by two separate equations,
applying  to  two distinct phenomena, independent of
each other.
No. 2. 13 (C12 H24 012)* + i2 H2 0 = 24 (C6 H14 06) + 12 CO2
No. 3. 12 (C1'2 H'2 01)* = I2 (C12 H20 010) +24 H2 0;
which, added together, reproduce Equation No. I (plus
12 molecules of water).  The  respective proportions
of gum and mannite, corresponding to the preceding
results, are obtained generally and with constancy when
we sow the ferment before described.
In certain viscous fermentations, however, the proportion of gum  is greater than that of mannite.  In
this case, according to M. Pasteur, the presence in the
liquid of larger globules of a different kind is perceived.
The same writer adds, that it may be possible that this
second ferment transforms the sugar into gum only,
without there being any fermentation of mannite.  To
prove the accuracy of this view, it would be necessary
to be able to isolate the second ferment from the first,
and to cause it to act separately: hitherto this separation has not been effected. However this may be, this
variability in the proportion of the gum  tells in favour
of M. Monoyer's opinion.
The liquids which are most apt to produce viscous
fermentation t can also undergo lactic and butyric fer* Perhaps better written, 13 (C6 H12 06 + Co H12 0') and I2 (C6 H12 0o +
C' H12 06) as these formulae represent the " changed" or " altered" canesugar, i.e., cane-sugar that has taken up one molecule of water and split up
into a mixture of dextrose and levulose.
t Decoction of the yeast of beer filtered, and with the addition of sugar;




192                ON FERMENTATION.
mentation; but, in this case, the organized forms of life
which are developed in the liquid are of a different
nature.
The gum  which is formed under these conditions is
more allied, by its characters, to dextrine than to gum
arabic. Nitric acid converts it into oxalic acid without
the production of mucic acid. The conditions of action
necessary to these gummy and mannitic ferments are
the same as those which suit alcoholic ferment. The
most favourable temperature is 30~ C. (86~ F.).
water of flour, barley, and rice with sugar added. White wines are more subject to this change, called ropiness, than red wines: as this difference appears
to be connected with the absence of tannin in the former, M. Fran9ois proposes the addition of tannin to white wine, as a remedy for the ropy
affection,




CHAPTER IX.
LACTIC FERMENTATION.
THE lengthened discussion into which we have entered
with respect to the history of alcoholic fermentation
allows us to be much more brief when speaking of the
other phenomena of this order, which have been generally attributed, since the time of Pasteur, to the intervention of lower organisms.  Except the chemical
reaction, and the form of the ferment, the general
characters of these fermentations are the same, for the
very simple reason that the conditions of nutrition and
of development of organic ferments do not vary within
very wide limits.
By lactic fermentation is understood the transformation of certain sugars, as sugar of milk and grape-sugar
into a sirupy acid, soluble in water, (lactic acid). By
comparing the formulae of the sugar which ferments,
and of the lactic acid, the only product of its fermentation, we see that we have here a simple molecular
transformation, or rather the splitting up of one molecule
into two more simple equivalent ones. We have, in
fact,C6 H12 06  2 C3 H6 03
Sugpar.   Lactic acid.




194             ON FERMENTATION.
This transformation has been observed, long since,
under many circumstances.
*M. Boutron and M. Fremy were the first to consider
it the result of a special fermentation, independent of
the viscous fermentation with which it was confounded.
(Ann. de Chim. et de Phys. (3), vol. ii. p. 257, 27I.)
When milk turns sour spontaneously, the sugar of
milk which it contains is converted into lactic acid, and
it was from  sour skimmed  milk that Scheele first
extracted lactic acid, as early as I7So. Braconnot
found the same acid in rice left under water in fermentation, in the juice of beetroot, which, after having
passed through the viscous fermentation, and a slight
alcoholic fermentation, becomes sour, and produces
lactic acid and mannite; he also found it in the water
of fermentation of peas and boiled French beans, and
in the sour water of baker's yeast. The same product
is found in the sour water of starch manufacturers, and
in sauerkraut.  M. Fremy and M. Boutron, as well as
M. Pelouze and M. Gelis, ascertained the best conditions for effecting the rapid transformation of sugar
into lactic acid. According to their researches; lactic
fermentation requires the presence of nitrogenous albuminoid matter in process of decomposition, and can
only continue if the degree of acidity of the liquor is
kept from  exceeding certain limits. The end is best
attained if wee saturate it from time to time with sodium
carbonate, or add previously a quantity of chalk, sufficient to neutralize all the acid which can be formed at
the expense of the sugar.
We will now give, in a few words, the best process, as
published by the writers who have studied this subject.




LACTIC FERMENTATION.               I95
Three or four litres of milk are taken, to which from
200 to 300 grammes of sugar of milk is added, the
liquor is left exposed to the air, in an open vessel, for
several days, at a temperature between I5~ and 20~ C.
(50o~-68~ F.).  After this time the liquor is found to
be acid; it is then saturated by sodium  carbonate, and
these saturations are carried on successively, at intervals of twenty-four hours, till all the sugar of milk is
transformed. It is now boiled, filtered, and cautiously
evaporated to a sirup. The remainder is treated with
alcohol at 38~, which dissolves the sodium lactate.  This
solution is precipitated by a suitable quantity of sulphuric acid, which precipitates the soda; it is then
filtered and evaporated.  The remainder, saturated with
lime, will give botryoidal crystals of pure calcium lactate.
(Boutron and Fremy.)
Bensch's process is also to be recommended. It consists
in dissolving three kilogrammes of cane-sugar (about
6'6 lbs. av.) and 15 grammes ('o83lbs. av.) of tartaric
acid in I3 kilogrammes of boiling water. The solution
is then left to itself for some days; after which 60
grammes (' 321bs. av.) of old decayed cheese, diffused
in 4 kilogrammes, (881lbs. av.) of curdled and skimmed
milk, I 2 kilogrammes of washed chalk (3'31bs. av.) are
added. The whole is put in a warm place 30~-350 C.
(86~-95~ F.), and stirred from time to time. After
eight or ten days, the liquid forms into a mass of calcium lactate. Ten kilogrammes of boiling water (221bs.
av.) and I5 grammes ('o331bs. av.) of slaked lime are
then added; it is boiled, and passed through a cloth;
then evaporated and crystallized. The crystals are
pressed and purified by new crystallizations. The cal



196                 ON FERMENTATION.
cium lactate is then dissolved in water precipitated by
a suitable quantity of sulphuric acid (2Io   grammes for
I kilogramme of lactate); the calcium sulphate is separated, and the liquid is neutralized by zinc carbonate;
we thus obtain zinc lactate, which can be easily purified
by crystallization, and is readily converted by sulphuretted hydrogen into lactic acid and insoluble zinc sulphide. Nine kilogrammes of sugar thus yield easily
I.of kilogrammes of calcium  lactate.
The juice of fermented beetroot (Pelouze and J. GayLussac), and the water in which sauerkraut has been
washed (Liebig), will also serve for the extraction of
lactic acid, by analogous methods, and by the use of
chalk, or zinc carbonate. The terms of the reaction,
and the conditions favourable to its production, were,
therefore, well known; but before the researches of
Pasteur, only vague and ill-founded ideas were held
respecting the cause which produced this phenomenon.
The apparent absence of an organized ferment, which
had  not been  found,*  gave  a  powerful support  to
Liebig's theory.
Casein or albuminous matter seemed to act upon
sugar only by its own decomposition, and it might be
supposed that the molecular movement, the consequence
* The true lactic ferment had, however, been already surmised by Dr. Remak, (Remak Canstatt's Jahersbericht, i. I841), and partly described by
Blondeau (x848), "Journ. de Pharm. " (3), vol. xii, 244 and 336. Remak had
observed that the globules of beer-yeast are often mixed with smaller ones,
which give rise to a special fermentation, and can, in no case, be transformed
into globules of the yeast of beer. Blondeau points out in beer-yeast two
kinds of cells, the ordinary ones of. oI millimetre in diameter, setting up
alcoholic fermentation, and cells about four times smaller, which give rise to
lactic fermentation. Blondeau attributes to them both butyric fermentation
and the ammoniacal fermentation of urea; but in this he was mistaken.




LACTIC FERMENTATION.              I97
of this spontaneous decomposition, was capable of
transmitting itself to the saccharine principles contained
in the liquor.
M. Pasteur, as early as 1857, guided by well-founded
ideas as to the causes of alcoholic fermentation, and
fermentation in general, sought for the lactic ferment;
that is to say, for the organism which transforms sugar
into lactic acid by the effect of its nutrition and
development, and succeeded in finding and studying it,
although it did not show itself so distinctly as alcoholic
ferment.
"If we examine," said he (Ann. de Chim. et de Phys.
(3), vol. iii. p. 407, Memoire sur la Fermentation appele
Lactique), " with attention, ordinary lactic fermentation,
we find cases in which we can recognize upon the
deposit of chalk and nitrogeneous matter, spots of a
grey substance sometimes forming a zone on the surface
of the deposit.  This substance is sometimes found
sticking to the upper parts of the vessel, whither it has
been carried by the movement of the gas. Its examination with the microscope scarcely enables us to
distinguish it, unless we are prepared for it beforehand,
from caseum, disaggregated gluten, &c.; so that nothing
shows that it is a special substance, nor that it has been
formed during the fermentation. Its apparent weight
is always very trifling compared with that of the nitrogeneous matter primarily necessary for the production
of the phenomenon.  Indeed, it is so very often mixed
up with the mass of caseum  and chalk, that we should
have no reason to suspect its existence.  It is this,
nevertheless, which plays the most important part."
To prove this, M. Pasteur thoroughly washed fresh




198            ON FERMENTATION.
yeast with fifteen or twenty times its weight of boiling
water. To this liquid, filtered with care, and containing
the most appropriate nitrogeneous and mineral aliment
for the nutrition of organic ferments, was added from
50 to Ioo grammes of sugar per litre, and chalk. A
trace of the grey matter mentioned above was sowed
in it, carbon dioxide introduced instead of air, and the
whole was kept at 30~ or 35~ C. (86~ to 95~ F.). In a
short time, all the symptoms of lactic fermentation
were developed, accompanied by those of a weak butyric
fermentation.
When the chalk has disappeared, the concentrated
liquid furnishes an abundant crystallization of calcium
lactate, while the mother liquor retains calcium butyrate. Sometimes the liquor becomes very viscid. In
a word, we have before our eyes a most characteristic
example of lactic fermentation, with all the accidents
and the usual complication of this phenomenon. At
the same time that the reactions that have been described take place, the liquid, limpid at first, becomes
troubled; a deposit appears which increases continually,
as fast as the chalk is dissolved. We may substitute
FiG. 2I.-Lactic Ferment,
in this experiment any plastic nitrogeneous substance,
either fresh or decomposing, for the decoction of yeast.
The substance which is deposited, and whose production is correlative to the reactions known by the name




LACTIC FERMENTATION.              199
of lactic fermentation, exactly resembles, when taken
in mass, ordinary yeast, which has been drained of
water and pressed. It is rather viscid, and of a grey
colour. Under the microscope, it appears as if formed
of small globules or short joints, either isolated or in
a mass, forming flakes resembling those of certain
amorphous precipitates (Fig. 2I). The globules, much
smaller than those of yeast of beer, are violently
agitated by the Brownian movement, when they are
isolated. When washed with much water by decantation, and then diffused in pure solution of sugar, it
begins to render it acid at once, but progressively,
and very slowly, because acidity greatly interferes with
its action. If we add chalk, so as to maintain the
neutrality of the medium, the transformation of the
sugar is sensibly accelerated; at the end of an hour,
gas begins to be disengaged; and the liquor becomes
charged with calcium lactate and butyrate in variable
quantities.
When the solution of sugar contains nitrogeneous
matter, and salts suited to the nutrition of ferments,
the lactic ferment is developed, and we obtain quantities which have no limit except the weight of the sugar
and the albuminous substances employed.  But little
of this ferment is necessary to transform a considerable
wveight of sugar. Its activity is only weakened when
we dry it, or boil it in water. This fermentation ought
to be set up without access of air, in order that we
may not be annoyed by vegetation and infusoria from
without.
Lactic fermentation, under the conditions indicated
by M. Pasteur, wheazn  te lactic fcrilzcnt is dcvelopcd




200            ON FERMENTATION.
spontanzcozs/y, is often more rapid than alcoholic fermentation, and shorter than in the ordinary process.
(Boutron and Fremy, Bensch, &c.)
The most favourable temperature seems to be about
350 C. (95~ F.).
If, without making any change in the conditions of
the saccharine medium, which when sowed with- the
lactic ferment, produces lactic fermentation, we sow
this medium with globules of yeast, we observe the
appearance of alcoholic fermentation and the development of alcoholic ferment.
Finally, to complete the analogy between the two
kinds of fermentation, M. Pasteur showed that lactic
ferment can, like alcoholic ferment, be produced as if
spontaneously in saccharine liquids of a composition
adapted to its development. Thus in alcoholic fermentations in an artificial medium, sowed with traces
of alcoholic ferment, M. Pasteur almost always saw
lactic ferment and fermentation appear side by side
with alcoholic ferment and fermentation.
In these cases of spontaneous production of lactic, in
the presence of alcoholic ferment, the predominance
of one or the other organism, and consequently of its
effects, depends on the composition of the medium,
being more or less suited to one or the other ferment,
and notably so, on the neutral state of the liquid.
Thus, if we add magnesia to a mixture of solution of
sugar and beer-yeast, there will be at the same time
alcoholic and lactic fermentation with precipitation of
magnesium lactate, and we shall see, mixed with the
globules of yeast of beer, a considerable quantity of
the small globules of lactic ferment.  A  medium,




LACTIC FERMENTATION.             20 I
slightly alkaline, is more suited to the development
of the new ferment; it is the same with respect to
alcoholic ferment.
The concomitant development of alcoholic and lactic
ferment explains why frequently the presence of lactic
acid has often been observed as one of the products of
the decomposition of sugar under the influence of beeryeast.
Most careful and frequent experiments proved to
M. Pasteur, that not even the smallest quantity of
lactic acid is formed in normal alcoholic fermentation.
When it does appear, which is very rarely, unless we
choose favourable conditions, we may be sure that the
beer-yeast is mixed with lactic ferment, which can
easily be recognized by its form, its size, and its
movement.
To be certain of the presence or absence of lactic
acid, we operate as has been before explained (page 30)
when treating of succinic acid and glycerine. The
ethereal alcoholic solution of the extract of the fermented liquid is evaporated; the remainder is saturated
with lime-water; the liquid is again evaporated, and
the remainder is taken up by ethereal alcohol. The
residuum is finally heated with boiling alcohol at 95 per
cent., which dissolves the calcium lactate.
The following substances are susceptible of undergoing
lactic fermentation, the glucoses and those bodies which
are convertible into glucoses. The saccharoses which
most easily undergo alcoholic fermentation, are those
which are with the greatest difficulty transformed into
lactic acid, and vice versa. Sugar of milk does not readily
produce alcohol, while it is much more easily resolved




202             ON FERMENTATION.
into lactic acid, although according to Lubolt (Journ.
J. Prakt. Chim. vol. 77, p. 282) and Proust (Ann. de
Chim. et Phys. (2), vol. Io, 29) the action is very slow,
sugar of milk being found in the liquor up to the end
of the fermentation.
Sorbin, inosite, mannite, and dulcite do not undergo alcoholic fermentation, but are affected by lactic
ferment.
Calcium malate, and, in general, all substances whose
fermentation produces butyric acid, seem to have the
power of yielding lactic acid.
In the fermentation of calcium malate, carbon dioxide
is disengaged.
C4 H6 05    C3 H6 03 +  C 02.
Malic Acid.    Lactic Acid. Carbon dioxide.




CHAPTER X.
AMMONIACAL FERMENTATION.
UREA (C HO NM O) is, as is well known, one of the
most important constituents of urine. It appears in it
as the principal form under which nitrogen is eliminated
from the animal organism.
This body, of simple composition, crystallizable in
long prisms, only differs from ammonium carbonate by
the elements of water.
We have, in fact,(CH4 N2 O) +  H20  =   CO2  +  2 N  H3a.
Urea.     VWater.  Carbon dioxide.  Ammonia.
By boiling in alkaline, or even in pure water, this
phenomenon of hydratation is produced more or less
quickly. We even observe it with the more complex
compounds, known under the name of ureids, and in
which we must admit that the molecule of urea is
associated with other organic groups, such as uric acid,
alloxane, creatine, &c.
A solution of pure urea in water may be preserved
for a long time without change. It is not so with the
natural solutions of urea, such as urine, which contains
~ Or better, CH4 N2 0 + 2 2 0 — = (NH4)2 C 03.
Urea.    Water.  Ammonium carbona.




204             ON FERMENTATION.
salts and other nitrogeneous principles more similar to
albuminous substances.
It is well known that, after a longer or shorter time,
according to the conditions of temperature, and the
state of health of the individual who has excreted
the urine, this liquid, after its emission, becomes alkaline, instead of acid as it was before; at the same time,
it exhales a very decided odour of ammonia; at this
moment the urea has disappeared, or is on the point of
disappearing entirely; we find in its place an equivalent
quality of ammonium carbonate.
In certain cases, the urine is already alkaline and
ammoniacal, when in the bladder.
The name of ammoniacal fermentation has been
given to this spontaneous transformation.  (Dumas,
Traite de Chimie, vol. 6, p. 380.)
According to the observations of Miller (I86o, Journ.
ffir Prakt. Chem., 8I, p. 467) and of Pasteur (Comp.
Rend., vol. 50, p. 869, May, I86o), the transformation of
urea into ammonia and carbon dioxide is due to the
intervention of a special organic ferment, produced by
one of the torzlacei and formed of chaplets of globules
very similar to those of beer-yeast, but much smaller;
their diameter is about 5io~ of a millimetre (,ooooo78 in.).
M. Van Tieghem has very thoroughly studied this ferment. It is found in the white deposit left at the bottom
of urinals. M. Jaquemart (Ann. Chim. Phys. (3), vol. 7,
p. 149, I843) had already noticed that this deposit was
very apt to excite this transformation.
Long study of the organic products which are developed in urine exposed to air, has convinced M. Van
Tieghem of the constant presence of a (torulaceous)




AMMONIACAL FERMENTATION.            205
ferment, whenever urea ferments; and of the intimate
connection which exists between its easy or difficult
development, and the rapid or slow transformation of
the urea.
" In the case, seldom realized, in which this torulaceous
growth is developed alone, the liquid remains limpid,
the fermentation is rapid, and the deposit which forms
at the bottom of the vessel is composed exclusively of
chaplets and masses of globules mixed with crystals of
urates and of ammonium-magnesium phosphate. If the
torulaceous growth is only accompanied by infusoria,
as is usually the case, the fermentation, though somewhat slower, is still easy; but if there appear, besides
the infusoria, vegetable productions in the liquid and on
the surface, the torulaceous growth is developed with
difficulty, and the transformation is very slow; the
liquid may remain acid or neutral for months together.
" If, instead of leaving the urine to the variable chances
which the sequence of the presence of the germs in the
air involve, we place it in a stove in a corked bottle,
having added to it a trace of the deposit of a good fermnentation, all accidental variations disappear, and the
phenomenon takes place always in the same manner;
one or two days are sufficient for the urea to disappear,
and at the same time the torulaceous growth alone is
developed.
"The transformation of urea in urine is therefore
correlative to the life and development of an organic
vegetable ferment. This ferment is developed within
the liquid itself, and especially at the bottom of the
vessel, where, by its accumulation, it forms a whitish
deposit, and is composed of chaplets, or small masses of




206            ON FERMENTATION.
spherical globules, without granulations, without any
distinct envelope of the contents, and which appear to
develop by budding; their diameter is about'ooi5
millimetres ('oooo59 in.)."
M. Van Teighem considers that he has proved by
direct experiment, that the splitting up of hippuric acid
by hydratation, into benzoic acid and glycocol, which is
observed in the urine of the herbivora after emission, is
due to a fermentation analogous to that which splits up
the urea. The active ferment must be identical with
the ammoniacal ferment. Thus, ammonium hippurate
dissolved, either in yeast-water or in solution of sugar
containing phosphates, is always split up in consequence
of the development of a microscopic vegetable organism identical with the torulaceous growth described
above. (Comp. Rend. de l'Acad. des Sci., vol. 58, p. 533.)
According to MUller, the activity of the phenomenon
is proportionate to the number of globules. When toa mixture of. sugar and urea, dissolved in water, we add
beer-yeast, we always see the small globules of ammoniacal ferment make their appearance, as soon as the
liquid shows an alkaline reaction (Pasteur). The yeast
of beer, by itself, decomposes the sugar without exciting
the decomposition- of the urea.  The most suitable
temperature is that of the human body, 37~ C. (99 F~).
Finally, as is the case with the must of grapes and
the.wort of beer, the ferment does not pre-exist in the
urine; it must be brought from without, in the form
of germs.
The necessity for ammoniacal ferment, in order to
transform the urea found in urine, under ordinary conditions of temperature, raises a medical question of




AMMONIACAL FERMENTATION.           207
great importance. In fact, many cases of disease have
been observed in which the urine was more or less ammoniacal in the bladder. This change usually accompanies either vesical or renal injuries, or serious general
diseases, such as typhoid fever. When the urine is ammoniacal after the catheter has been introduced, it is to
be feared that the instrument has served as a vehicle,
and carried into the bladder the germs of the torulaceous growth, thus setting up infection. Ammoniacal
urine, from a patient at la Charite, was examined by
M. Gayon, at the very instant of emission; he found in
it innumerable organisms: this examination took place
some days after the patient had been relieved by the
catheter. However, in the opinion of surgeons, the
urine often shows itself to be ammoniacal without any
previous operation of this kind; for example, in cases
of retention, although the long continuance of the
liquid in the bladder may not be the only condition of
this phenomenon. In fact, M.. Verneuil did not find
the urine ammoniacal in the case of an hysterical young
girl, in whose case the catheter had been passed, after
several days' retention.
As M. Pasteur has observed, the canal of the urethra,
with relation to the infinitely small germs which act as
ferments, may be compared to the Thames tunnel,
through which they can pass and repass easily. If then,
we should feel astonished at anything, it is iather at
finding the ammoniacal infection occur so infrequently.
It is, perhaps, because the urine is usually acid, and
that this acid is unfavourable to the development of the
germs of fermentation.
However this mav be, M. Pasteur, without being




208            ON FERMEN TATION.
able to give any positive proof, is inclined to believe
that the decomposition of urea is not possible, without
the presence of a- peculiar ferment, and that there is
no purely chemical reaction in the human body, capable
of giving rise to ammonium carbonate in the urine.
This conviction has been the result of long study of
ferments, and of his great experience.   He owns,
however, that the definite solution has not yet been
obtained. (See on this subject the discussion on ammoniacal urine at the Academie de Medicine, Moniteur
Scientifique (3), vol. 9, p. I43.)
In conclusion, the transformation of the urea in urine,
at the ordinary temperature, may certainly be caused
by a special ferment. It is possible, but as yet not certainly proved, that the presence of this ferment may be
indispensable.




CHAPTER XI.
BUTYRIC FERMENTATION AND  PUTREFACTION.
A GREAT number of chemical compounds are susceptible of fermenting butyrically, that is to say, yielding butyric acid as a product of their transformation,
when they are placed under suitable conditions.
Such are lactic acid, and all substances capable of
undergoing  lactic  fermentation-sugars, amylaceous
matter, tartaric, citric, malic, mucic acids, and albuminoid substances.
It is easy to represent by equations the production
of butyric acid at the expense of the greater part of
these bodies, which are well defined and of well-known
composition.
Thus sugar and lactic acid would giveC6 H12 06 = 2 C3 HI 03 = C4 H8 02' +   2 CO2  +  H4
Glucose.   Lactic acid.   Butyric acid.  Carb. dioxide. Hydrogen.
For malic acid we shall have2 C4 HI O  2 C3 H6 03 + 2 C02 =  C4 HS 02
Malic acid.    Lactic acid. Carbon dioxide. Butyric acid.
4 CO2   +    H4
Carb. diox.    Hydrogen.
Tartaric acid would be resolved into butyric acid,
carbon dioxide, and water;
10




210               ON FERMENTATION.
2 C4 H6 06 =  C4 Hs 02  +    4 C02    + 2 H2 0
Tartaric acid.   Butyric acid.   Carbon dioxide.   Water.
Perhaps lactic acid is formed first:
3 C4 HI 06 =  2 C3 H6 03 +   6 CO2 +   H6
Tartaric acid.    Lactic acid.   Carb. diox. Hydrogen.
According to M. Personne, citric acid would be transformed previously into lactic and acetic acid:
4 C6 HS 07 + 2 H~ O =  3 C2 H4 02 + 4 C3 H6 03 + 6 CO 2
Citric acid.  Water.    Acetic acid.  Lactic acid.  Carb. diox.
Mucic acid, which differs from citric acid only by
containing more water, is resolved, like it, into acetic
and butyric acids, carbon dioxide, and hydrogen:
3 C6 H10 08 =  3 C2 H4 02- +C4 H8 02 +  8 C02 + H10
Mucic acid.    Acetic acid.   Butyric acid.  Carb. diox. Hydrogen.
In other analogous phenomena, which go on at the
expense of the same substances, we find acids homologous to butyric acid; in the same way as, in alcoholic
fermentation, alcohols are formed with higher equivalents than that of ethyl-alcohol.
Thus, under the conditions of lactic fermentation, we
see the formation of propionic acid, acetic acid, and
valerianic acid at the expense of sugar or of starch.
Glycerine, placed in contact for a long time with
beer-yeast, also furnishes propionic acid, mixed  with
formic and acetic acids. M. Monoyer represents in
a general manner the production of fatty acids at the
expense of sugar by the equationn - I (C6 H12 66 ) — [CCn H2n 02  + (n - 2) CH2 02
Fatty'acid.   Formic acid.




BUTYRIC FERMENTATION AND i'UTREFACTION. 211
Formic acid itself would be decomposed into carbon
dioxide and hydrogen.
Crude calcium tartrate, mixed with organic matter,
and left under water, in summer, ferments and produces
anll acid which was at first identified with propionic acid,
but which, according to Limprecht and Von Uslar, must
be isomerous with ordinary propionic acid.
Crude tartar, without the addition of lime, gives
nothing but acetic acid. We have2 C4 Hi 06 = C3 H6 02 + 5 CO2 + H6
Tartaric acid. Propionic acid. Carb. diox. Hydrogen.
C4 H6 06    C2 H4 02 + 2 C02 + H'2
Acetic acid.
In the fermentation of mucic acid, of which we have
spoken above, the butyric acid, which only appears
slowly, and in small quantity, ought to be considered as
a secondary product. The principal phenomenon may
be summed up in a chemical point of view by the
equationC6 I-10 08 = 2 C2 H4 02 - 2 C02 + H2
All the compounds of the malic group, such as malic,
fumaric, aconitic, aspartic acids, and asparagine, undergo,
under the form  of salts of lime, and in the presence
of animal matter in process of decomposition, a transformation into different products, among which we may
consider succinic acid as the principal term.
With malic acid, we should have2 C4 HG 05 = C4 H6 04 + C2 H4 02 + 2 CO2 + H2
Malic acid.   Succinic acid. Acetic acid. Carb. diox. Hydrogen.
The sum of the last three terms of the second member




212             ON FERMENTATION.
gives the composition of tartaric acid; we may, therefore, consider succinic fermentation as the result of two
reactions. In the first, two molecules of malic acid are
converted into one molecule of tartaric acid, and one of
succinic acid:
2 C4 H6 05 = C4 H6 04 + C4 H6 06
Malic acid.   Succinic acid. Tartaric acid.
The second reaction would be the fermentation of tartaric acid formulated above.
The production of valerianic acid in the succinic
fermentation of calcium malate is explained by the
equation2 (C4 H6' 04) = C5 Hl 02 + 3 CO2 + H2
Succinic acid. Valerianic acid.
As maleic, fumaric, and aconitic acids only differ from
malic acid by less water, their transformation into
succinic acid is explained in the same manner.
As aspartic acid may be considered as amido-succinic acid, and asparagine as amido-succinamic acid,
the production of succinic acid at their expense is not
surprising.
We have just passed in review a great number of
reactions; they all belong to the class of fermentation,
because they are excited by certain bodies which act
only by their presence.
They have, besides, one thing in common —the formation of acids of the fatty series.
We must consider it as yet vague and uncertain
whether the cause of these fermentations is to be attributed to the presence of a special ferment for each, as
we have seen in lactic and alcoholic fermentations.




IBUTYRIC FERMENTATION AND PUTREFACTION. 213
In fact, this cause has only been thoroughly studied
and determined by Pasteur, for the butyric fermentation
of sugar and calcium lactate.
In the fermentation of ammonium tartrate, M. Pasteur found a ferment similar to lactic ferment, which,
acting on ammonium paratartrate, causes the decomposition of the levo-tartrate; which affords a new
example of elective fermentation, comparable to that
which we have observed in levulose and glucose.
In M. Pasteur's opinion, the butyric decomposition
of calcium lactate is due to the presence in the liquid
of a special ferment-fermcultztm buztyricuzm.
"Butyric ferment is composed of little cylindrical
rods, rounded at the extremities, usually straight, either
isolated or united in a chain of two, three, or four joints,
and even of more. The diameter of these small rods
is generally rfjo  of a millimetre, and the length of the
isolated portions from xo  to   o  mm. ('oooo687 to
~ooo687 in.). These organisms move forward by sliding.
During this movement their body remains rigid or
undulates slightly; they spin round, they balance themselves on end, and agitate their extremities: they are often
bent. These singular organisms are reproduced by fission.
"The butyric ferment is, therefore, an infusorium of
the genus vibrio." (Pasteur, Comp. Rend., 52, p. 344,
February, i86I).
The same writer has ascertained that this ferment,
placed in a solution of sugar containing phosphates and
ammoniacal salts, reproduces, and causes butyric fermentation.
The conditions of its development are similar to those
of lactic fermentation.




214             ON FERMENTATION.
The most favourable temperature is 40~ C. (IO4~ F.):
the medium should be neutral, or slightly alkaline. Al
acid medium  is opposed to the development of the
germs of butyric fermentation. However, when once
formed, they can live and excite the decomposition
of sugar or lactic acid in an acid medium, provided
there be no excess of acidity.
M. Pasteur at first asserted that butyric vibrios
not only lived without free oxygen,* but that oxygen
kills them.  The respiratory theory of fermentation,
proposed by this observer, does not agree with this fact.
If fermentation is the result of such a need of oxygen,
that the ferment takes it up from organic compounds,
exciting their decomposition by a rupture of equilibrium,
we cannot understand how oxygen can act as a poison
to the ferment. I do not know whether M. Pasteur
has since maintained the opinion that oxygen kills
butyric vibrios.
The conditions of nutrition of butyric ferment are,
according to Pasteur, the same as those of ferments in
general.  However, considering its tardy appearance,
as compared with lactic ferment, in mixtures which
undergo lacto-butyric decomposition, we may admit
that it requires albuminoid substances in a process of
more advanced change for its nourishment.
In sugar, we generally see viscous, lactic, and butyric
fermentation appear in succession. However, it is not
proved that butyric fermentation cannot appear before
lactic fermentation, and thus act on the sugar itself.
Pzutrefactioun-Putrid Fermez tatioz. —Albuminoid substances, and bodies allied to them, which enter into the
* The alcoholic and lactic ferments are in the same category.




BUTYRIC FERMENTATION AND PUTREFACTION. 215
composition of living organisms, have, for a long time,
enjoyed a special reputation for instability, which varies,
however, according to the nature of the substance.
Before Pasteur's researches, it was generally admitted,
that as soon as the influence of life is withdrawn (the
vital force which alone was able to maintain them in
their integrity), these products begin to be transformed,
to change, and to be decomposed into several principles,
among which are found compounds with a strong and
putrid odour.
The remarkable researches of Appert on the preservation of animal substances, and those of Gay-Lussac on
the fermentation of the must of grapes, had given the
idea that the momentary intervention of oxygen is
necessary to excite the first step in this decomposition.
This initial impulse once given, the phenomenon of
decomposition goes on spontaneously; and the organic
matter in process of transformation is even susceptible
of transmitting the molecular movement with which it
is imbued to more stable bodies, such as sugar, which
of themselves undergo no modification. This is, as we
have already seen, the theory of fermentation borrowed
by Liebig from Stahl and Willis, with certain modifications of form.
However, Schwann (Ann. de Poggend., 41, p. I84),
Ure (Journ. Prakt. Chem., I9, I86), and Helmholtz (J. F.
Prakt. Chem., 3 I, p. 429), had shown that the greater part
of bodies subject to decomposition, when heated in a
retort with water, so as to drive out all the air by boiling,
are no longer decomposed, if instead of allowing ordinary air to enter the retort as it grows cold, we are
careful only to admit air previously subjected to a red




216            ON FERMENTATION.
heat. Under these conditions, putrefaction does not
make its appearance, and we no longer observe the
development of infusoria and mildews.
The abundant presence of infusoria and mildews in
putrefaction had been long known, but it was not thought
that these microscopic beings were the true causes
which determined the decomposition.  They are developed, it was said, owing to germs brought by the
air, or already contained in the decomposing bodies, or
by spontaneous generation, and because they find a soil
favourable to their nutrition. The bond between their
appearance and putrefaction was only one of concomitance.
Schwann, and the other authors quoted above, thought,
on the contrary, that the germs of infusoria and of mildews set up putrefaction by their development; and, as
a proof of this, they brought forward their experiments,
in which putrefaction was no longer produced, when the
pre-existent germs were destroyed, and their introduction
by means of the air was prevented.
As the calcination of the air gave rise to some objections, especially to that of a possible change in the
constituent principles of this gaseous mixture, Schroeder
and Th. V. Dusch (Ann. der Chem. und Pharm., vol. 89,
p. 232), repeated Schwann's experiments, with this
difference, that instead of allowing calcined air to reenter the retort in which the organic substance had been
boiled, they simply filtered the air through a sufficiently
thick layer of cotton wool; they thus succeeded in
mechanically arresting the germs and solid matters held
in suspension, but without influencing in any way the
properties and the composition of the air. The wort of




BUTYRIC FERMENTATION AND PUTREFACTION. 2I7
beer, broth, meats recently boiled in water, are then
preserved very well, even during the heat of summer.
Some contradictory facts, however, lent support to
the arguments of the adversaries of the theory of putrefaction under the influence of infusoria. Thus, the
authors of the before-mentioned experiments had themselves ascertained that milk recently boiled coagulates,
grows sour, and putrefies, just as well in air that has
been strained as in ordinary air; meat not steeped in
water, but simply heated in a water-bath, is also not
preserved in strained air: in these two cases we observe
neither infusoria nor mildew, and yet decomposition
is produced.
It is then evident, it was said (Gerhardt, Chimie
Organ., vol. 4, pp. 545), that it is indeed the air which
brings and deposits in matters in a state of putrefaction the germs of organisms, but it is not less certain
that these are not the first cause of decomposition, since
it can be produced without their intervention. If the
calcined or strained air is less active, in many experiments, than ordinary air, it is because not only are the
germs of infusoria removed by these operations, but
also the remains of decomposed matter which are suspended in it; that is to say, ferments whose activity
would be added to that of oxygen.
The question was in this state when Pasteur resumed
the study of putrefaction, by looking upon it in the same
light as had guided him in his researches on fermentation. Sustained by the idea that all these phenomena
can be explained by the presence, the development,
and the multiplication of microscopical plants or
animals, he sought to prove that some of these exist




218            ON FERMENTATION.
also in the putrefaction of animal nitrogenous substances.
These experiments were conducted by two methods
which lead to the same end, and confirm each other.
On the one hand, they tend to show that putrefaction
is always accompanied by the presence, the development, and the multiplication of infinitely small, organized, living beings; on the other hand, they prove,
that whenever we place ourselves under conditions
calculated to avoid the presence of the germs of organisms, at the commencement of the experiment, decomposition does not take place, even in piroduzcts the most
liable to it.
By their precision and their extent, they are calculated to remove the objections raised by the partial
decompositions noticed by his predecessors, Schroeder
and V. Dusch, and which we have mentioned before.
We will speak of the second series of experiments
when we treat of the origin of ferments, only saying
here, that the many trials made by Pasteur lead to a
positive solution of the question. By preventing contact of the germs with animal matter, we prevent, at
the same time, every trace of fermentation from showing
itself.
M. Pasteur distinguishes two orders of phenomena
in putrefaction; some are produced under the: influence
of organic ferments which live without the aid of oxygen,
like butyric ferment; in others, on the contrary, the
oxygen takes part, as an essential element, promoting
combustion; oxidation is also excited by organisms.
We shall treat of slow combustions in the chapter
on acetic- fermentation; this will give us a simple and




B3UTYRIC FERMENTATION AND PUTREFACTION. 219
clear example of the reactions of this group, and may
be considered as the type of slow comnbustions. We
have nothing to add here to what will be said on this
subject, and there only remains for us to speak of putrefactions without oxygen, orputridfereneztation.
When, in a putrescible liquid, containing albuminoid
organic matter, the dissolved oxygen has been absorbed,
and has completely disappeared under the influence of
the first infusoria developed, such as the Mo/nas crepusczuhzim, and the Bacteriumz termo, "the vibrio ferments,
which do not require this gas to sustain their life, begin
to show themselves, and putrefaction is immediately set
up. It is accelerated by degrees, following the progressive increase of the vibrios.  As to the putridity, it
becomes so intense, that the examination of a single
drop of the liquid, under the microscope, is a very painful
task."
" It follows, from what has been said, that contact of air
is by no means necessary for the development of putrefaction. On the contrary, if the oxygen dissolved in a
putrescible liquid was not at once removed by the action
of special organisms, putrefaction would not take place;
the oxygen would destroy the vibrios which would try
to develop at first."
When the putrescible liquid is exposed to the air, we
notice the two kinds of reactions simultaneously; there
forms on the surface a complete film, composed of
bacteria, mucors, and mucidines, which excludes the
oxygen, and prevents its penetrating into the liquid.
The vibrios which multiply there, under.shelter of this
rampart, transform by fermentation the albuminoid
matter into more simple products, while the bacteria




220            ON FERMENTATION.
and mucors excite the combustion of these products,
and bring them back to the state of the least complex
chemical combinations. Such is the representation of
the whole of the phenomena of putrefaction as drawn
by M. Pasteur (Comp. Rend., June, I863).
We have already made some reservations relative to
this singular property of vibrios-that of not being able
to endure the presence of oxygen.
The opinion held by Schwann, Ure, Helmholtz,
Schroeder, and V. Dusch, and finally by Pasteur, relative to the cause of putrefaction, is corroborated by the
very process which is employed to preserve perishable
bodies. The conditions of preservation are precisely
such as oppose the development of organisms.
Such are the employment of cold, zero C. (320 F.), and
below; also of a sufficiently high temperature. Cooked
albuminous matter resists putrefaction much longer,
because the germs which were there are destroyed; but
decomposition will, nevertheless, show itself if we do
not carefully guard against effects from without. Appert's process, which consists in cooking meat, or other
perishable substances, in iron boxes hermetically sealed,
realizes these conditions. The germs are killed, and
there is no possibility of fresh ones entering. As, at the
same time, the small quantity of air contained in the
box loses its oxygen, it has been thought that the
preservation depended on this complete elimination of
the oxygen at I00oo C. (2 I2 F.).
The total absence of water very efficaciously opposes
the development of living organisms. Thus we can
preserve, as we may say, indefinitely, dried meat and
vegetables.




BUTYRIC FERMENTATION AND PUTREFACTION. 22I
All substances known as antiseptics are also enemies
to ferments. Thus common marine salt, alcohol, creosote, phenol, salicylic acid, sulphurous acid, the sulphates, potassium acetate (Sacc.), carbon dioxide, tannin,
the acids, many metallic salts, as'those of copper, mercury, iron, aluminum; potassium chromates, arsenious
acid, prussic acid, lime water, the antiseptic properties
of which are well known and have been frequently tried,
all these are also poisons for ferments of various kinds
in the quantities in which they are active.
T'he preservative action of oil, grease, ashes, fine sand,
bran, sawdust, coatings of paraffin or gelatine, is explained
by these porous or impermeable bodies preventing the
approach and access of germs brought by the air, like
the cotton wool in Schroeder's experiment.
Products of Putrefaction.-The products of putrefaction are very numerous. This may be easily understood, first, because the putrid change of an organ or
liquid directly taken from the animal or vegetable
economy is the resultant of the decomposition of the
various constituents which are found in it. The special
study of the products of putrefaction of each particular
albuminoid substance has only been attempted in a very
few cases.
In the second place, the compounds, definite in appearance, which undergo putrid fermentation, are so complex
in their constitution, that we ought to expect to meet
with a great number of derivatives formed by putrid
decomposition.
The most constant products which make their appearance in putrefactions screened from the air are leucine,
and probably some of it homologues, tyrosine, the vola



222            ON FERMENTATION.
tile fatty acids of the series Cn H2n O0 (formic, acetic,
propionic, butyric, valerianic, caproic, &c.), ammonia, and
some compound ammonias (ethylamine, propylamine,
amylamine, trimethylamine), carbon dioxide, sulphuretted hydrogen, hydrogen, and nitrogen.
If we refer to what will be presently said with reference to the decomposition of albuminoid substances
under the influence of barium hydrate, we shall be able
easily to account for the appearance of these various
products. On one hand, the albuminoids contain the
elements of urea, and ought to be considered as coinpound ureids. This fact alone explains the appearance
of carbon dioxide, and of a part of the ammonia. (See
ammoniacal fermentation.)
The albuminoids are decomposed by hydratation
under the influence of baryta, furnishing leucine and
some of its homologues, tyrosine and a sulphide. These
first products may, probably, undergo the ulterior action
of ferments, and yield ammonia and volatile fatty acids.
We know, in fact, that in presence of putrefied fibrin,
leucine is resolved into ammonia and valerianic acid.
C6 H13 N 02 + 2 H2 0 = C5 H10 02 + N H3 + CO2 + H4
Everything leads us to believe that putrefaction is a
complex phenomenon —that it is only a successive series
of fermentations exerted on more and more simple
products.
Thus, for example, when we leave fibrin to spontaneous decomposition, without access of air, it is resolved
into two principles, as under the influence of seasalt. One of these principles is albumin, which, on
account of its greater resistance to the action of fer



BUTYRIC FERMENTATION AND PUTREFACTION. 223
ments, will be found for a long time in the putrid liquid.
The second product of this decomposition, undergoing
somewhat quickly a more thorough change, yields acetic,
butyric, valerianic, and capric acids, as well as ammonia
(Brendecke), which are evidently derived from the
amido-acids homologous with leucine.'The chemical reactions which accompany the putrefaction of the albuminoids are, then, for the most part,
phenomena of hydratation, which may be reproduced
identically by chemical forces alone, independently of
vital action. Thus we shall see that phenomena of this
kind may be excited by the action of soluble ferments,
whether diastasic or indirect; and we are induced to
suppose that a part, at least, of the transformations
undergone by proteids, and their more immediate derivatives, are the consequences of phenomena of this order
(indirect fermentation).
Nothing resembles putrid fermentation, with reference
to the derived products, more nearly than the change
which takes place in the constituent parts of yeast,
when left to itself without nourishment, deprived of sugar
and oxygen.
We see, in fact, the appearance of leucine, tyrosine,
sarcine, &c. This is the first step; the action stops
there, and goes no farther; the yeast, or the special
soluble ferment which it secretes, is unfit to attack
these bodies again; but if we wait for the development
of vibrios, we shall find the production of ammonia,
carbon dioxide, and volatile fatty acids, at the same
time that the leucine partly disappears.
M. Ulysse Gayon has published quite recently, as
a thesis for the "Doctorat es Sciences" (Paris, I875,




224            ON FERMENTATION.
Faculte des Sciences de Paris, No. 362), the result of
many experiments on the spontaneous decomposition of
eggs. The question was important, and very interesting
to the adversaries of the theory of spontaneous generation. Besides, the facts observed by M. Donne and
M. Bechamp on this subject seemed contrary to the
ideas of M. Pasteur on the general cause of putrefaction.  M. Gayon, a pupil and demonstrator of
M. Pasteur's, endeavoured to bring the spontaneous
decomposition of eggs and their putrefaction under the
general law enunciated by his teacher.
M. Donne (Experiences sur l'Alteration spontanee
des CEufs, Comp. Rend. de l'Ac. 57, p. 450, I863) had
said: " If we take eggs in their natural state, not shaken,
and leave them to themselves, they remain for weeks
and months, even during the great heat of summer,
without undergoing any putrid decomposition. The
egg has no unpleasant smell, and nothing, either possessing animal or vegetable life, is produced, either on
the surface of the membrane or in the inside; there
are no traces of infusoria or microscopical vegetation.
" If, on the contrary, we destroy the physical structure of the interior of the egg by shaking; if, that is
to say, we break up the texture, and the cells of the
albuminous substance, and thus mix together the yolk
and the white, then, even without access of the external
air, and even guarding against this intervention by extra
precautions, such as a coating of collodion spread over
the surface of the egg, we find all the phenomena of
decomposition make their appearance, after a longer or
shorter time, according to the temperature, but always
in less than a month; all the phenomena of decomposi



BUTYRIC FERMENTATION AND PUTREFACTION. 225
tion, with the exception, however, of the production of
living organisms, either vegetable or animal; for, whatever may be the degree of rottenness to which we allow
the egg to proceed, we can never discover the slightest
trace of animalcule, or of microscopic vegetable life;
the matter of the egg grows troubled, and of a livid
colour; it exhales a fetid odour directly we break the
shell, but nothing, absolutely nothing, stirs in its substance; nothing lives, and the most careful and frequently repeated examination by means of the microscope does not enable us to discover the least trace of
an organized or living being."
We may add, that M. Bechamp also found no organisms in rotten eggs.
M. U. Gayon's experiments, into the details of which
we cannot enter, led him to the following conclusions:"Putrefaction in eggs, whether in the presence or the
absence of air, is correlative to the development and
multiplication of microscopical organisms of the family
of vibriones.
"In other terms, contrary to the result found by M.
Donne and M. Bechamp, eggs make no exception to
the great law of correlation which M. Pasteur has
demonstrated for all the phenomena of fermentation,
properly so called."
We are thus, on the subject of eggs, confronted by
two distinct affirmations, as much opposed as black
and white.  M. Donne found them; M. Gayon did
not. Web have no balance wherewith to estimate and
compare the skill of the two observers. It appears to
us certain that M. Gayon saw what he described;
but we cannot affirm that M. Donne was absolutely




226                  ON FERMENTATION.
mistaken, and  that the  eggs, in  the  conditions under
which he placed them, contained vibrios which he did
not find.
In the absence of any other criterion, we bring forward  a very important fact, mentioned by  M. Gayon
himself.
This skilful microscopist observed that some of the
eggs experimented upon at the temperature of about
250 C. (770 F.), whether shaken or no, underwent a
special modification, distinct from ordinary putridity
and  from  acid fermentation.The decomposed mass is of a dirty yellow colour,
it has an odour of dried animal matter, and is very fluid;
we see in it, also, a great number of needle-like as well
as botryoidal crystals, formed of tyrosine. It contains
much  greater quantities of tyrosine and  leucine  than
are found in ordinary putrefaction.   M. Gayon was not
able to  discover, under these circumstances, any trace
of microscopic organisms, either in the  inside, on the
surface, or in  the substance of the membranes.  IHowever, tyrosine and leucine are evident and unquestionable symptoms of the decomposition of albuminoid
matter.
Between the production of these substances and the:- In certain cases, M. Gayon saw the contents of eggs, especially of those
which had been shaken, transformed into a homogeneous mass, of the consistence of butter, and of a bright yellow colour, with a sour smell, and a
strongly acid reaction. M. 1Bchamp observed a similar change in an ostrich
egg, and was able to distinguish the presence of alcohol, acetic acid, and
sulphuretted hydrogen. M. Gayon attributes this change, to which he gives
the name of acid fermentation, to the presence of special organisms. These
are small immovable rods, with pale outlines and homogeneous tints, either
isolated or articulated two and two together, from T1 to 1, of a millimntre
in length, and smi, to i'a0 of a millimPtre in thickness.
iBk~d65 t




3UTYRIC FERMENTATION AND PUTREFACTION. 227.
phenomena called putrefaction there is, chemically
speaking, no very clear distinction to be drawn. They
are reactions of the same order, decompositions, more
or less extensive, of the proteid molecule; the traces
of sulphuretted hydrogen, and other fetid products
which communicate such a repulsive odour to putrefaction, cannot serve to establish an absolute and philosophical line of demarcation between the decomposition
without organisms observed by M. Gayon, and what is
wrongly termed putrefaction properly so called.
The result of this seems to be, that albuminoid matters are able to undergo certain decompositions, certain
changes, without the intervention of living organisms.
By means of a very simple and ingenious apparatus,
M. Gayon succeeded in extracting the gas contained
in large ostrich eggs in a state of putrefaction.
One of these eggs, in a state of thorough putrefaction,
yielded I50 cubic centimetres of gas, containing per
cent.:Sulphuretted hydrogen            Traces.
Carbon dioxide.....30'5
Hydrogen.                        402
Nitrogen......  29'3
I00'0
The presence of nitrogen might be due to the accumulation of a certain quantity of air in the air-bubble before
putrefaction.
Among the solid and liquid products of the putrefaction of eggs,the presence of small quantitiesof leucine and
tyrosine, alcoholic products, and volatile acids (butyric
acid) were recognized. The sugar had disappeared.




228            ON FERMENTATION.
CHAPTER XII.
FERMENTATION BY OXIDATION.
ACETIC fermentation and the reactions which we shall
class with it under the generic name of fermentation by
oxidation, have a special character, which we have
not met with in any of the phenomena which we have
hitherto studied.
Not only are the fermentable matter and the ferment
concerned in the reaction, the one furnishing the constituent parts of the new bodies which are formed, and
the other acting as a cause, but we find that a third
factor, the oxygen of the air, becomes necessary.
In other words, under the name of fermentation by
oxidation, we shall speak of combustions set up by
living organisms, which serve as media between the
oxygen of the air and the combustible body or fermentable matter. A~ s to the results of this combustion, they
may vary, according to the nature of the body which
is burnt, and may even be resolved into the simple products of the most complete combustion (water and
carbon dioxide).
We will begin with the acetic fermzetatiool of alcohol.
It has been long known that the alcohol contained in
fermented liquids, such as wine, beer, &c., will disappear
under certain circumstances, and give place to vinegar




FERMENTATION BY OXIDATION.           229
or acetic acid, and that the air, or rather its oxygen,
plays a part in this reaction. The progress of chemistry,
and the exact determination of the respective composition of alcohol and acetic acid, give a simple and clear
account of the reaction; it may be formulated thus:C2 H6 0 + 02 = H2 0 + C2 I14 02
Alcohol.           Acetic acid,
The oxidation may take place by two reactions, with
the production of an intermediate product, aldehyde:C2 HG 0 + 0   H= I2 O + C2 H4 0
Alcohol.            Aldehyde.
Cs H4 0 + 0 = C2 H4 02
Aldehyde.    Acetic acid.
In a purely chemical point of view, we have here a
very simple phenomenon, on which we need not dwell.
The determining causes of this oxidation will have
greater claim  on our attention.  D6bereiner having
shown by an experiment, which has become classic, that
the vapour of alcohol mixed with the oxygen of the
air becomes acid, being transformed into acetic acid,
thought that he had ascertained by this means the true
theory of the phenomenon, the essential conditions of
which became the simultaneous action of alcohol and
oxygen, in presence of a porous body, such as finely
divided platinum, charcoal, wood shavings, &c., capable
of favouring by a catalytic action, that of contact, the
oxidation of the alcohol.
It was on this idea that the process of rapid acetification, called the. German process, was founded; this was
first tried by SchUitzenbach in 1823. Long before this,
Boerhaw had already introduced into practice an analo



230             ON FERMENTATION.
gous commercial process.  He employed vats three
metres high and one and a half metres in diameter, with
a double bottom pierced with holes, placed at 30 centimetres (about a foot) from  the bottom.  Bunches of
grapes, from which the juice had been expressed, were
placed on this false bottom, so as to fill the vat. One of
the vats was entirely filled with wine, the other was only
half filled; after twenty-four hours, liquor was drawn from
the full vat and poured into the other so as to fill it; this
operation was alternately repeated till the acidification
was complete. It is seen that, in the vat which is half
full, the alcoholic liquor which soaks the grape stalks
and skins is exposed to the action of air on a large
surface, and the oxidation is singularly favoured by this
fact.
In Schiitzenbach's process, use is made of a large oak
vat, from two to three metres in height by one in diameter, furnished with a false bottom pierced with holes,
and  placed about 30 centimetres from  the bottom.
Some centimetres higher, the circumference of the vat is
regularly pierced with a series of holes passing entirely
round it.  These orifices are inclined from  without
inwards, so as to prevent the liquor from escaping.
At the upper part, at the distance of 30 centimetres
firom the lid, another false bottom is placed, pierced with
many small holes and several large ones; the latter
are usually closed by plugs, and are intended for the
purpose of renewing the air, when it is deoxygenated.
The whole; is closed by a cover furnished with an opening carrying a funnel which can be closed, and serves
for the introduction of the liquor to be oxidated. The
interval between the two partitions is filled with beech



FERMENRTATION BY OXIDATION.          231
shavings. A thermometer placed in the interior of the
vat gives an idea of the intensity of the reactions.
All being thus arranged, hot vinegar is first poured
into the vat; this filters through the shavings, impregnates them, and serves afterwards to facilitate, or rather
to set up, the oxidation of the alcohol. In order to be
transformed into vinegar, suitable mixtures of alcohol
and vinegar are employed, similar to those which are
used in the Orleans process. The temperature of the
surrounding air is maintained at 2I~ C. (about 7~0 F.).
The alcoholic liquor employed is heated to between 260
and 27~ C. (790 and 8I~ F.); the temperature rises spontaneously to 380 to 420 C. (IOI~ or Io8~ F.) in the vats.
The alcoholic liquor is never completely oxidized by
its first passage through the vat; the operation is once
or twice repeated, either in the same vat, or in others by
the side of it. (For further details see special works on
chemical technology.) In good manufactories, a result
is obtained which does not differ by more than six per
cent. from that indicated by theory; and even this loss
may be lessened by certain suitable accessory arrangements, to which we need not now allude.
The French method of acidification of wine, called
the Orleans method, is very different. We will say a
few words about it before we enter on the results of
Pasteur's researches, and the commercial consequences
which have been derived from them.
The oxidation takes place in barrels placed side by
side, on wooden frames, supported by stone pillars. At
the upper front part of each vat two orifices of unequal
size are made; the larger serves for the purpose of
introducing and drawing off liquor, the other for the




232             ON FERMENTATION.
admission of air. These vats, which contain from two
to four hundred litres (44 to 88 gallons) called mnothers,
are at first one-third filled with strong boiling vinegar;
from II to 12 litres (20 to 2I pints) of wine are then
added, and the vats are then left undisturbed; at the
end of a week's acetification, another quantity of wine
is added, and so on from time to time till the vessel is
half full of vinegar. A third part of the contents of
the " mother" is then drawn out by means of a siphon,
and wine is again added from time to time in portions
of I I to  12 litres (20 or 2I pints).  A  regular and
continued process is thus carried on. It is often found
that, without any apparent cause, a certain "mother"
refuses to acidify the wine, or only gives rise to a very
slow and tedious oxidation; in this case, the employment of a much stronger wine, or a much higher temperature, sometimes restores the action to its normal activity.
These anomalies have only been explained by Pasteur's
experiments. The operation in one barrel is considered
to be terminated when a stick plunged into the liquid is
covered with a thick white froth (flour of vinegar); as
long as the froth is red, the addition of wine is continued.
The most favourable temperature is between 24~ and
27~ C. (760 to 82~ F.).
In this special manufacture of vinegar, the influence
of porous bodies in determining the oxidation cannot
be brought to bear, as in the experiment with platinum
or in the German method of manufacture. The vinegarmakers, from their special experience, attribute the
acidification to the action of a deposit which forms
in the barrels, and to which they give the name of,, mother of vinegar."




FERMENTATION BY OXIDATION.         233
According to the opinions of Liebig, which have so
long prevailed in scientific matters, either dead or living
organic matter, when in contact with alcohol in wine,
possess the property, after having absorbed oxygen, of
oxidizing, at the ordinary temperature, organic and inorganic substances. This property would therefore exist
in solid organic substances which are in a state of decomposition or putrefaction. To support this view, Liebig
(Ann. de Chim. (4), vol. 23, p. I78,) recalls de Saussure's
experiment, who found that soil placed in a mixture of
hydrogen and oxygen gave rise to the formation of
water and the disappearance of hydrogen, and of an
equivalent quantity of oxygen. We may state in a few
words, and without entering into too many details on a
question which seems determined, that Liebig explains
the production of acetic acid by a certain movement
communicated by principles in process of decomposition
-a movement which in this case excites oxidation.
In fact, his ideas are wanting in clearness. Sometimes
he seeks for the key to the phenomenon in a catalytic
action of porous bodies; sometimes, with reference to
his general theory of fermentation, he attributes it to the
influence of a ferment (organic matter in process of
decomposition).
Such was the state of the question when Pasteur
undertook his researches on the causes of the spontaneous acidification of wine and alcoholic liquors.
According to him, the oxidation of alcohol is the consequence of the action of a cryptogam of the genus
Mycodermna. We will give a summary of the experiments on which his opinion was founded.
If on the surface of any organic liquid, necessarily
11




234            ON FERMENTATION.
containing phosphates and nitrogenous organic matter,
we allow any species of mycoderma to develop itself,
until the whole surface of the liquid is covered with it;
if then we carefully remove the nutritive liquid by means
of a siphon, without suffering any portion of the membranes to break up; then, if we substitute for this liquid
an equal volume of water, containing Io per cent. of
alcohol, we immediately see the plant which is placed
under these abnormal circumstances of nutrition, set up
a reaction between the oxygen of the air and the alcohol
of the liquid. The acetification begins immediately,
and goes on with great activity. After a certain time,
the action, impeded by the great acidity of the liquid,
proceeds more slowly; but we can restore to it all its
activity by substituting alcoholized water for the acid
liquid. A time, however, comes when the plant, becoming partly decomposed itself, communicates to the liquid,
in consequence of the organic and mineral elements of
its dead tissues, properties which serve as nutriment for
the various species of mycoderma. The action then takes
on a different phase; the acetic acid and alcohol disappear with great rapidity, and the liquid becomes completely neutralized; because as soon as the plant finds
in the subjacent medium the nutritive principles suited
to its development, it sets up much more intense oxidizing action, and burns, not only the alcohol, but also
the acetic acid, converting it into water and carbon
dioxide.
This complete combustion is noticed whenever we
cause the mycodermata to be developed on alcoholic
liquids containing food fit for the nourishment of the
plant, such as wine, beer, or fermented organic liquids;




FERMENTATION BY OXIDATION.          235
unless, however, we place the mycoderma, whether intentionally or unintentionally, under the conditions of
incomplete or tardy development, that is to say, in a
sickly state.
To sum up, we may say that the mycodermata, developing on the surface of an alcoholic liquid which contains suitable nutritious principles, burn the alcohol, and
bring it to the same state as oxygen at red heat, that is,
complete combustion. If, on the contrary, we diminish
the vital activity of the mycoderma, whether by depriving it of its nourishment or by any other means, the
oxidizing action which it may be able to set up will not
go so far, and the alcohol may change into acetic acid.
M. Pasteur's experiments will also show that the
influence attributed to ordinary porous organized bodies,
in the German manufacture of vinegar, is the result of
imperfect observation.
The beech-shavings act, not on account of their
porosity, but because their surface is coveredwith thin
pellicles of mycoderma; the many points of contact
with the air favour the action, but are not the determining causes of it.  To prove this, M. Pasteur
caused some alcohol diluted with water to trickle down
a cord. The drops which fell from the end of the cord
did not contain the smallest quantity of acetic acid.
The experiment lasted for more than a month, the
liquid trickling extremely slowly, only one drop in two
or three minutes. If we repeat this experiment, having
previously steeped the cord in a liquid on the surface of
which there is a pellicle of mycoderma, a portion of
which clings to the cord as it is withdrawn, the alcohol,
which passes slowly; down this cord, will be charged with




236              ON FERMENTATION.
acetic acid, and this acetification may be continued for
several weeks.
M. Mayer made some similar experiments, which
entirely confirmed Pasteur's results. Thus filter paper,
previously boiled with hydrochloric acid at 50 per cent.,
then with caustic soda at 50 per cent., and finally washed
with distilled water, was laid on the surface of an alcoholic liquid, without giving rise, even after the expiration
of a month, to the least trace of acetic acid. The result
was also as negative when a funnel was filled with fragments of glass and of the same paper, and alcohol at
9 per cent. was allowed to run slowly through this filtering mass.
M. Pasteur did not go more deeply into this question, and did not ascertain by what means the mycodermata could thus give rise to more or less energetic
oxidations.
In one of his latest writings on this question (Reponse
aux Critiques de M. Liebig, Ann. Chim. Phys. (4),
vol. 25, p. I48, I872), he thus expresses himself:"This little microscopical plant (Mycoderma aceti)
possesses the power of condensing the oxygen of the
air in the same manner as spongy platinum  or bloodglobules,* and of conveying this oxygen to  matter
beneath."
It seems, from these few words, that M. Pasteur
compares the action of his organic ferment, the Mycodermtna aceti, to that of spongy platinum: his theory of
acetic fermentation really differs very slightly from that
The oxygen of the globules of blood is fixed in the hanmaglobin, and its
oxidizing power is scarcely more energetic than that of ordinary dissolved
oxygen.




FERMENTATION BY OXIDATION.           237
of Liebig. The latter admits that inanimate porous
substances participate in the properties of spongy platinum; Pasteur, on the contrary, attributes this quality
only to living organisms.
Mayer's experiments tend to prove that oxidation by
mycodermata is a special biological action, which cannot
be attributed solely to the physical condition of the plant
which acts as ferment. In fact, we have only to heat an
alcoholic liquid, covered with its pellicle of acetic mycoderma in full process of acidification, and we shall arrest
all oxidation; yet it is difficult to believe that under
these conditions the physical state has been sensibly
modified.
M. Mayer also noticed striking differences between
the mode of action of spongy platinum and that of the
/Mycorderza aceti. For the first, an elevation of temperature above 35~ C. (95~ F.) favours the oxidation, by
augmenting the tension of the vapour of alcohol, while
the maximium activity of the Afycodermca aceti is placed
between 200 and 300 C. (68~ and 86~ F.); its oxidizing
power is destroyed under IO~ C. (5o0 F.) and above
350 C. (950 F.). We can oxidize very concentrated
alcohol with platinum; the concentration is even a
favourable factor; while for physiological acetification,
the alcohol employed ought scarcely to contain more
than io per cent. of ethyl hydrate.
The immediate and practical consequence of the
results obtained by Pasteur is, that in order to act
efficaciously, the mycoderma ought to be in contact,
at the same time, with  the air and the alcoholic
medium. The new commercial method for the acetification of fermented liquids, now employed to some




238            ON FERMENTATION.
extent at Orleans, is founded on this principle. The
following is the process:The I7fycordcra anceti is first sowed on the surface
of an aqueous liquid containing 2 per cent. of alcohol,
I per cent. of vinegar, and traces of alkaline and
alkalin-earthy  phosphates.  When  the  surface  is
covered with the membrane, the alcohol begins to
acidify. This action being fully set up, some alcohol,
wine, or beer mixed with alcohol, is added every day to
the liquid, in small quantities; this is continued till the
oxidation becomes slower; the acetification is then
allowed to terminate, and the vinegar is drawn off.
The membrane is collected, washed, and employed for
a new operation. It is better always to give the plant
sufficient alcohol, so that its activity should not be
exerted on the acetic acid. Nor ought it to remain too
long out of the liquid, or it would lose its active force;
finally, it is better to moderate its development, to prevent burning oxidation.
A vat, one square metre in section, and containing
from 50 to ioo litres (from  iI to 22 gallons), can furnish per day from 5 to 6 litres (9 to Io' pints) of
vinegar.  The successive phases of the operation are
ascertained by means of a thermometer divided into
tenths of a degree, the bulb of which is plunged into
the liquid.
When we act upon diluted alcoh6l, it is better to add
to the liquid -    of a mixture of magnesium phosphate, and potassium and ammonium phosphates.
Special arrangements allow the introduction of the
liquid, without there being any necessity for disturbing
the superficial film of mycoderma.  The vessels are




FERMENTATION BY OXIDATION.             239
cylindrical, or of a prismatic form, of one square metre
in section, and -} metre in depth, closed by a cover furnished with orifices for the admission of the air.
FIG;. 22.-M-ycoderna aceti.
It only remains for us to say a few words about the
botanical character of the fM9corderaza a eti.
The  continuous  membranes, either wrinkled  or
smooth, which are found on the surface of liquids
while in acetic fermentation, are generally formed
(Fig. 22) of very minute elongated cells, whose greater
diameter varies from I, 5, to 3 thousandths of a millimetre (0oooo59 to  oooI I8 in.); these cells are united
in chains, or in the form  of curved rods.  Multiplication seems to be effected by the transverse division of
the fully developed cells. This division is preceded by
a median constriction, which has been considered by
some authors as a morphological characteristic of
the cell.
It appears, from this description, that the mycoderma of vinegar belongs to the family of bacteria.
The general conditions of the nutrition of acetic
bacteria have been discovered by Pasteur, and closely
resemble, up to a certain point, those of beer-yeast.
Thus mineral salts, alkaline and alkalin-earthy phosphates, proteid nitrogenous substances, or ammoniacal
salts, are elements necessary for the development of




240             ON FERMENTATION.
these organisms. Diluted alcohol (io per cent. at most)
seems, in this case, to take the place of hydrocarbon
matter; it may be supplemented by the acetic acid;
for, according to M. Pasteur, the progressive weakening
which vinegar undergoes, when the acetification is left
too long to itself, is due to a subsequent burning of the
acetic acid.
M. Blondeau (Comp. Rend., vol. 57, p. 953) has even
observed that sugar can acidify without passing through
the condition of alcohol, under the influence of the
mother of vinegar.
We must, however, notice that the activity of the
ferment is increased by the presence in the liquid of a
certain quantity of acetic acid.
Antiseptic agents, in general, which by their presence
delay and arrest the development of the yeast of beer,
and consequently alcoholic fermentation, act in the
same manner with respect to the Mycoderi.za accti.
Sulphurous acid is especially active in this manner;
and it is partly in order to avoid the acetification of
wine, that care is taken to put in the tuns intended to
receive it, sulphurous acid, or to burn sulphur matches
in them.
The AIycoderina vizni, of which we have spoken
above (p. 59), resembles in many respects the acetic
ferment.  Like it, it is developed on the surface of
fermented alcoholic liquids, in the form of smooth or
wrrinlkled films or membranes; but these latter, however, are thicker and more compact.  It acts also as
the means for conveying the Oxygen of the air to
the alcohol of the medium, and to the other comr
bustible principles; but, under its influence, combustion




FERMENTATION BY OXIDATION.         241
is complete, and accompanied by the production of
carbon dioxide and water; it is to this action that we
must attribute the rapid weakening of wines covered
with mycoderma. We have elsewhere seen that, when
immersed in the midst of a saccharine liquid, it can
act like the yeast of beer, and produce alcoholic fermentation.
Its nutritive principles are the same as those of the
mother of vinegar (alcohol, salts, nitrogenous compounds)-; besides, it appears also capable of utilizing
for nutrition certain secondary products of alcoholic
fermentation, such as succinic acid and glycerin. The
forms of the cells of this mycoderma-forms that we
know to be variable, seem.to depend in a great degree
on the conditions of nutrition.
Its activity of development appears included between
i6~ and 30~ C. (6I~ and 860 F.).
Slow Combitstioll. —The Mycodermzata of which we
have just spoken are not the only organized ferments
capable of exciting the slow combustion of carburetted
materials.
I- t has long been known that organic matter, of vegetable or animal origin, left in contact with air, undergoes
progressive and complex transformations, known under
the name of putrefaction, of slow combustion, and of
eremocausis, whose effect is to transform them into
principles more and more simple, by means of decomposition and oxidation; so that, in the end, the
carbon is restored to the air in the form of carbon
dioxide, the hydrogen under the form of water, the
nitrogen either as free nitrogen or ammonia. M. Pasteur
has distinguished, among the complicated facts of putrid




242            ON FERMENTATION.
fermentation, two orders of distinct phenomena, although
each is connected with the reactions set up by living
organisms. The first includes the putrefaction, which
takes place without the assistance of oxygen in the air,
which is caused by the presence of vibrios. We have
spoken of this in the chapter on butyric fermentation,
with which these phenomena are connected.
The second, slow combustion, is due to bacteria,
mucors, and mucidines, that is to say, to vegetable
ferments, which, like the Mycoderlza vizzi, and others,
possesses the remarkable property of exciting the
oxidation of a great number of organic principles,
such as sugars, alcohols, organic acids, albuminoid
nitrogenous matter, &c., at the expense of the oxygen
of the air.
After having proved, by careful experiments, to which
we will return when we treat on the origin of ferments,
that spontaneous slow combustion of animal or vegetable
substances depends necessarily on the, development of
organisms in the interior, or on the surface, of the
substances which are in process of decomposition, and
that without organisms there is neither combustion nor
absorption of oxygen, M. Pasteur traces the following
picture of putrid decomposition in contact with air
(Comp. Rend., June, I863):" Even the most easily decomposed animal matter, as,
for instance, blood or urine, may be preserved for an
indefinite length of time in air which has been calcined
or deprived of its germs; under these conditions, the
absorption of oxygen is but trifling, and putrefaction
does not take place; and, at the same time, no infusoria
are produced. If, on the contrary, this same substance




FERMENTATION BY OXIDATION.            243
remains exposed to the ordinary air, it is oxidized,
putrefies, and infusoria are developed.
"It is commonly known that putrefaction takes a
certain time to declare itself, a period varying according
to the circumstances of temperature-of the neutral,
acid, or alkaline character of the liquid. Under the
most favourable circumstances, at least twenty-four
hours are- required before the phenomenon begins to
manifest itself by external signs.  During the first
period, an internal movement takes place in the liquid,
the effect of which is to withdraw entirely the oxygen of
the air which is in solution, and to substitute for it carbon
dioxide gas. The total disappearance of the oxygen,
when the medium is neutral or slightly alkaline, is generally due to the development of the smallest kinds of
infusoria, especially the J/Ionas crepusculum, and the
Bacterizm ternzo. A very slight troubling then  takes
place, because these little beings pass about in all directions. If the vessel containing the putrescible liquid
has a large opening to the air, the bacteria perish only
in the liquid mass, after the removal of the oxygen,
while they continue, on the contrary, to propagate, ad
infinitum, on the surface, because it is in contact with
the air. There they cause a thin film to form, which
goes on thickening by degrees, until it falls to the bottom
of the vessel; then another forms, and so on continually.
This film, to which different mucors and mucidines are
attached, prevents the solution of oxygen gas in the
liquid, and consequently allows the development of
vibrios. With respect to these latter organisms, the
vessel is as if it were closed against the introduction
of air.




244            ON FERMENTATION.
The putrescible liquid thus gives rise to two very
distinct kinds of chemical action, which have reference
to the two sorts of organisms which are nourished in it.
On one hand, the vibrios, living by the co-operation
of the oxygen of the air, set up in the interior of the
liquid acts of fermentation-that is to say, they transform
the nitrogenous matter into more simple, but still complex, products.
The bacteria (or the mucors), on the other hand, consume these same products, and bring them to the state of
the most simple ordinary combinations, water, ammonia,
and carbon dioxide.
The compounds which longest resist slow combustion
are the fixed fatty acids, forming the adipocire of the
old chemists, cellulose or its derivatives formed by
dishydratation, such as ulmic acids, vegetable mould,
peat, &c.
The oleic acid, on the contrary, disappears altogether.
But little is known of the details of these- various
phenomena of slow combustion.
Can the same organism set up the combustion of
different organic principles, differing from one another
in their constitution? Is the action of the oxygen progressive, or is it complete -from the very commencement? These and many other secondary questions,
more or less interesting, present themselves; but their
solution requires long and minute researches, similar to
those which relate to alcoholic fermentation. However,
the cause of the phenomena is known, and the way for
new investigations is open.




CHAPTER  XIII.
APPLICATIONS OF THE RESEARCHES AND IDEAS OF
M. PASTEUR.
WE have already seen, when treating of acetic fermentation, what consequences M. Pasteur has drawn from
his observations, in order to regulate and facilitate the
transformation of fermented liquors into vinegar.
The manufacture of beer may also, according to this
investigator, derive advantage from the careful study of
fermentation and of ferments.
If we add to the wort of beer ordinary yeast, principally composed of cells of Saccharolnyces cerevisiw,
with very few foreign organisms, under the most favourable conditions for the development of yeast, it will
reproduce almost alone, setting up direct alcoholic fermentation. As the quantity of original yeast is found,
after the fermentation, to be six or seven times greater
than at first, supposing that no foreign germs have been
introduced from without, we can understand that the new
yeast will be purer than the first. By continuing with
this, new fermentations of the wort of beer, we shall
procure, by a kind of selection similar to that described
by M. Raulin, with reference to Asspergill/zs zziger, a very
pure yeast, free from the mixture of other organisms.




246            ON FERMENTATION.
When this result has once been obtained, it will be sufficient to maintain the integrity and purity of ferment, by
excluding from contact with air the fermenting vats of
beer, and by carrying on the process in closed vessels,
instead of open vats. The principle of the new process
patented by M. Pasteur, into the details of which we
cannot enter, depends then on the employment of pure
yeast, and on fermentation with the exclusion of air, in
order to avoid the introduction of foreign organisms,
which, by their ulterior development, might produce
changes of another order, lactic fermentation, &c.
For this purpose, the wort, after it is prepared, is
drawn off while boiling into vessels made of wood or
metal, and cooled in a current of carbon dioxide,
or of air purified from ferments, and then yeast is
added.
The beer, after the first fermentation, is drawn off
into casks, in which it ripens and grows clear. The
wort may be carried to a great distance, and the beer
has, according to M. Pasteur, superior qualities both as
to taste and preservation.
Preservation of Wilie.- M. Pasteur has made a long
and very attentive study of the changes which may take
place in wines in the various phases of their preservation.
His observations have been preserved in a very excellent
work published on this subject. We can only give here
the more general results obtained by this author. He
attributes the changes which take place in wines to the
development of special living ferments.
A special organism corresponds to each order of
change, i.e., to each disease of the wine. The germs of
these ferments are found in the must of fermented grapes,




RESEARCHES OF M. PASTEUR.                                  247
pi /
_ a,, ii:I     l  1111111
FIG. 23. —Giret and Vinas' apparatus for warming wines.




248            ON FERMENTATION.
and may develop themselves whenever the conditions
become favourable.
In order to prevent their development, and to allow
the wine to be kept for an indefinite term, without any
change, we have only to raise the temperature for a
moment to 60~ C. (I40~ F.). The germs are killed at this
comparatively low temperature, because of the presence
of alcohol (from 8 to Io per cent.).
The warming of wines for the purposes of preserving
them had already been proposed by M. Vergnette de la
Motte, but the rational explanation of this process is
due to M. Pasteur. He also determined with greater
precision the conditions of the operation.
This warming may be effected while the wine is in
bottle or in casks.
Fig. 23 represents the apparatus of Giret and Vinas
for the continuous warming of wine. It is composed of
a stove surmounted by upright tubes communicating with
the chimney. A water-bath completely surrounds these
tubes. The cylinder of the water-bath is fixed to the
stove by means of two rims, between which is a slip of
cloth saturated with paste. The two rims are compressed
by iron clamps, so that this cylinder can be easily removed. The wine circulates in a cavity formed by two
concentric cylinders, secured above and below by two
annular collars. The refrigerator is formed of a cylinder
containing an interior cavity similar in every respect
to the preceding one. The cover of the refrigerator
is movable, and is fastened by an arrangement like
that which serves to connect the cylinder with the
stove.
The surfaces in contact with the wine are tinned. The




RESEARCHES OF M. PASTEUR.             249
water-bath contains water, the refrigerator, as well as the
enclosure around it, only holds wine.
The arrows show the direction of the circulation; a
thermometer placed within the bulb of the tube which
connects the water-bath with the refrigerator gives the
maximum  temperature.  (For further details on this
subject see " La Chimie Technolique de Wagner," French
translation.)*
Application of M. Pasteur's ideas to Pat/lology.-Some
years since I wrote in the following terms (La Chimie
appliquee a la physiologie animale a la pathologie et au
diagnostic medical, par P. Schiitzenberger, I864):"All diseases, contagious either by inoculation or by
more or less direct contact, whether epidemic or endemic,
are evidently produced by the introduction of foreign
poisonous substances into the living organism, producing
true poisoning. When in an affection of this kind, as for
example, in cholera, yellow fever, or malignant ulcers,
the general symptoms and the mode of evolution have
a well-marked constancy of character, notwithstanding
differences of race, species, and individuals, one is compelled to admit the specific nature of the poison which
gives rise to certain pathological manifestations.
"These conclusions, drawn from many facts, observed
in every part of the world, are so simple and natural that
no one denies them; but when it is required to state the
precise nature of the morbific influence; as we enter the
domain of hypothesis, the most various and contradictory opinions have been given, and may be supported
with a greater or less appearance of probability.
"Infectious diseases were, for a long time, attempted to
There is an English translation, by W. Crookes, London.




250             ON FERMENTATION.
be explained by intra-organic fermentations, set up by
foreign bodies which everything induced us to consider
as of an organized nature; but at a time when the ideas
of fermentation, properly so called, were vague and illdefined,, it was difficult to maintain with certainty such
a doctrine.
" It has long appeared to us that Pasteur's researches
on this subject have not only succeeded in determining
with greater precision than before facts up to that
titne partly known, but that they are destined in the
future to throw a bright light on etiology, and on the
pathological history of contagious diseases, whether
epidemic or endemic. Yet we should not have ventured
to discuss here a personal conviction, which, though
shared by many physicians and observers, only rested
on analogy, were it not that a recent discovery, due to
the skilful investigation of M. Davaine, had corroborated
this opinion by a positive fact scientifically determined,
a fact which will certainly not remain unsupported by
others.
" We set out with the hypothesis that the greater part
of infectious diseases have for their immediate cause the
penetration into the organism, and the development of
the living germs of ferments, or living ferments already
formed, of either animal or vegetable nature, and we will
make use of the knowledge already acquired in order to
support this opinion with a certain amount of probability. We must premise, however, that it will be
necessary, before we decide this question definitively,
to wait for a direct and experimental proof, such as Al.
Davaine has given with reference to the " blood of the
spleen" or malignant boils. We think that researches on




RESEARCHES OF M. PASTEUR.          25 I
fermentation and putrefaction have been carried far
enough to allow important trials to be made in this
direction with some hope of success."
During the ten years since this page was written,
skilful experimentalists, guided by the same ideas, among
whom I may mention Pasteur himself (researches during
the cholera epidemic), have studied this subject with
great care, and yet I must admit that there has been no
result from these inquiries; the question of the etiology
of infectious diseases has made no important advance;
the observation made by M. Davaine remains without
any additional support.
Are we then to conclude that these attractive predictions are erroneous, and must be rejected, or that these
observations are impossible, even with the increased
microscopical power which we possess? It is difficult
to decide one way or the other, and every question requires for its decision positive facts; negative results
can only serve as a means of checking our observed
facts.








BOOK II.
ALBUMINOID SUBSTANCES-SOLUBLE OR INDIRECT FERMENTS —ORIGIN OF FERMENTS.
CHAPTER I.
ALBUMINOID SUBSTANCES, OR PROTEIDS.
ALBUMINOID substances, or proteids, play so important
a part in biological phenomena in general, and in the
nutrition of ferments in particular, that it would be
impossible for us not to devote a few pages to the
study of these bodies.
If these substances represented chemical species well
defined and correctly classed among organic compounds-in other words, if their constitution were
thoroughly known, we should merely refer the reader to
works on pure chemistry, as we have already done in
the case of fermentable sugars. But unfortunately, in
spite of many important investigations, it must be said




254            ON FERMENTATION.
that the history of proteids is still one of the most
obscure subjects of organic chemistry, one of the most
urgent desiderata of biological science.
So long as the question of the constitution of the
immediate principles of animal tissues has not been
determined, it will be in vain for physiological chemistry
to investigate by the most careful direct analysis the
different elements of an organ, or of a liquid, whether
in a normal or pathological condition; we shall always
be stopped by the unknown in the interpretation of
the results which have been obtained. The many and
various reactions which take place in the organism
may be regarded as true fermentations, in which the
fermentable bodies are partly represented by proteids.
It is not necessary to go into this at great length to
make it understood how much these reactions, as observed
in the different active phases of an organ, will gain
in scientific value, when we are able to formulate them
by means of an equation, as we give the formula of
alcoholic fermentation.
We shall study albuminoid substances, by taking
this general view of them, and by bringing out more
prominently, among the results obtained, those which
throw some light on the constitution and mode of
decomposition of these bodies.
The existence of a certain number of albuminoid
substances, considered as distinct species, has been
admitted, being founded on more or less important
differences in their physical or chemical properties, or
in their elementary composition.  This division and
classification may be made by two methods. If we
notice only very decided differences of character and




ALBUMINOID SUBSTANCES, OR PROTEIDS.  255
composition, we may form out of albuminoid substances
different groups or families united by common bonds,
but sufficiently distinct for confusion or error to be
rendered impossible.
Among these groups many closely allied species may
be placed, which owe their existence and individuality
only to certain subtile divergencies of properties which
present to the mind nothing very distinct, such as
the manner of forming precipitates with different
reagents.
Without entering into these details, we will take as
types of each group the most important principle from
a biological point of view. It cannot be said that
substances to which a special name has thus been given,
as the albumin of the egg, casein, and fibrin, are immediate well-defined principles; for here especially we
have no criterion by means of which we are able to
establish a chemical species. We may, indeed, suppose,
with a certain amount of probability, that they are
only mixtures, in variable proportions, of bodies very
nearly allied and almost identical, the separation of
which from each other would be very difficult, if not
impossible.  The natural fatty bodies give us instances of this complicated combination of products
very similar in their composition and character, and
which direct analysis has scarcely any power to
separate.
This hypothesis has also derived some support from
exact observations.  Thus, albumin, which was long
considered as an immediate principle, is, in fact, only
compounded of many albumins, having very Izearly the
same composition, and which can only be distinguished




256             ON FERMENTATION.
from each other by their rotatory power, and by the
temperature at which they coagulate.
This question can only be settled by a careful study
of the products of the analysis and decomposition of
albuminoid substances; as the nature of fatty bodies
has only been understood by examining the products of
their saponification, which are more easily separated
than are the original bodies.
These views, which are verified more and more, were
developed by M. Bouchardat (Thes pour le concours
d'agregation, I872), and by M. Berthelot, after a communication made to the " Societe Chimique" by the
author of this book. They are very reasonable, and
capable of wide application.
The nitrogenous principles which enter essentially
into the constitution of the organs and liquids of the
animal and vegetable economy are naturally divided
into various families.
The first includes albuminoid substances, properly so
called; that is to say, bodies the most nearly allied, by
their chemical composition and by the whole of their
properties, to the albumin of egg.
The second is formed of more remote products,
usually less rich in carbon and containing more nitrogen.  They enter into the composition of less vital
tissues, that is to say, those in which the phenomena
of nutrition and the changes are more restricted;
of those which do no work, that is, do not develop
much force, such as bony, cartilaginous, elastic, fibrous,
cellular, horny, or epidermic tissues. In this family we
find horny tissue, or keratrin, ossein, epidermose, elasticin, mucin or the gelatines, and the fibroin of silk.




ALBUMINOID SUBSTANCES, OR PROTEIDS. 257
The soluble ferments, the peptones, are products of
the decomposition of albuminoid substances, and should
be classed separately.
We find in the organism, besides these, certain mixed
principles placed between hydrocarbons and proteids,
true complex nitrogenous glucosides, decomposing into
glucose and other: nitrogenous principles, such as chondrin and chitin.4
Albmininoid Szubstances, propcrly  so called.-Into this
family we generally admit the following compounds:I. Albumin of the egg, the albumin of serum and
(serine), vegetable albumin; these substances are soluble in water.
2. Casein, vegetable casein, paralbumin, syntonin,
myosin, legumin, amandin, the proteids or albuminates
of the German chemists (this name cannot be adopted
in  French, since it gives an  idea that the body in
question is a salt), paraglobin, metaglobin or fibrinoplastic or fibrinogenous substances.
3. Blood-fibrin, coagulated albumin, amyloid matter,
and vegetable fibrin.
With regard to their centesimal composition, albuminoid substances are very similar to each other; yet the
analyses do not agree sufficiently to allow us to conclude that their composition is identical, or that they
are isomeric. We subjoin in a tabular form the principal analytical results:* According to my latest researches, albuminoid substances, even those of
the first family, contain, in small proportions, cellulose amides as an integral
part of their molecule. The distinction established between the albuminoids
and chitin is therefore not absolute. The latter contains, it is true, a greater
proportion of cellulose amide.
12




253S                   ON FERMENTATION.
|  2 d       a a    ha        Authors.
U                0
Egg albumin, not coagulated.              533   7I  5'8I8    23'6    I'8  Dumas and Cahours.
Egg albumin, not coagulated...  54'3   7   I57    22               -    Scheerer.
Egg albumin, purified    52'9  7'2  156    -     -   Wurtz.
Egg albumin, coagulated  52'9  7'2    58    -    -   Wurtz.
Blood fibrin.....  528   7'0    8    234          Dumas and Cabhours.
53,7  71  I5'8   23-4    -    Scheerer.
Casein.....'   53'5  71 7I5'8   236    -   Dumas and Cahours.
e,..'....    54.0  7'2  I5'7  231    -   Scheerer.
Gluten, or  vegetable
fibrin......      53    6'8  I50    -        -      ou singault.
Legumin...       537   72  157    234    -   Scheerer.
*...... 50'5  6'9  I8'2          -    Dumas and Cahours.
Amyloid matter...,    53'6  7'0  I5.3
Syntonin.........  4    702  16             1'I
On  the  one  hand, it is difficult to  attach  too  great
importance  to  small differences, observed  in  analytical
results obtained with  bodies so difficult to purify, amorphous, and often  containing mineral substances; on the
other hand, nothing authorizes us to admit the complete
identity of their composition.
In fact, these bodies have certainly a very high
molecular weight.    Thus for albumin, by two methods,
(analyses of the potassic and platino-hydrocyanic combinations)  nearly  the  same  number,  I612, has  been
arrived at for the  molecular weight; so  that if we wish
to translate the results of the elementary analysis into a
chemical formula, as is usually done, we are led to very
high numbers in the expression as in that proposed by
Lieberkuhn, C72 H "12 N18 S 022.
But it is evident to any one who is in the habit of
calculating analyses, that a difference of one atom of
carbon, hydrogen, or oxygen, in a formula with such




ALBUMINOID SUBSTANCES, OR PROTEIDS.  259
high numbers, gives variations which are within the
limits of the errors of analysis. Elemefitary analysis,
as we are at present capable of performing it, cannot
solve the question of isomerism or non-isomerism.
Are we farther advanced in that which relates to the
constitution of these bodies? At present, we can foresee
that, though the question is still in suspense, it will not
be long before it is determined.
In order to ascertain this constitution, it is necessary
to know exactly the whole of the constituent parts resulting from the destruction of proteids, under certain determinate conditions. But in the various reactions which
cause their splitting up and transformation into more
simple principles, we have been able to recognize the
formation of certain well-defined compounds; but these
bodies are more frequently accompanied by a relatively
considerable number of uncrystallizable bodies, which
have not yet been studied, and which render any
attempt to determine an equation of the reaction
illusory.
On this account, we cannot form any definite idea of
the manner in which the 72 atoms of carbon, the 112
atoms of hydrogen, &c., of the albumin are united.
I will give a short summary of the principal results
obtained with respect to these reactions.
As long since as I820, Braconnot observed the production of the sugar of gelatin or glycocoll, C2 H5 NO2
(amido-acetic acid) by boiling gelatin in sulphuric acid
moderately diluted; by substituting muscle for gelatin
he obtained, under the same conditions, the leucine,
C6 H"3 NO' (amido-caproic acid).
Liebig afterwards showed that another crystallizable




260              ON FERMENTATION.
product is formed at the same time, tyrosine, C9 H" NO3
(oxyphenyl-amido-proprionic acid).
Erlenmeyer and Schaeffer, by extending these researches (the action of boiling diluted sulphuric acid),
to most of the albuminoid  substances, observed the
constant formation of leucine and tyrosine.
They obtained for Ioo parts of dried matterLeucine.   Tyrosine.
Fibrin....          I4'8
Albumin  Io..                    I'
Syntonin....  8.. I'0
Casein gives leucine and tyrosine, and a sirupy residuum.
Ritthausen obtained by the same kind of reactions,
from products formed by the action of boiling dilute
sulphuric acid on vegetable nitrogenous substances, such
as gluten, two well defined crystallizable acids.
Coaglutin yielded an acid of the formula C5 H9 NO4
(glucamic acid), homologous to aspartic acid.
Legumin gives, under the same circumstances, legumic
acid, Cs H14 N2 06.*
According to Hlasiwetz and Haberman, the greater
part of animal and vegetable proteids are capable of producing these nitrogenous acids (aspartic, glutamic), when
boiled with dilute sulphuric and hydrochloric acids.
I have lately been led to study, with great care and
in all its details, a reaction which allows the albuminoids
to be almost entirely resolved into crystallizable principles. SMy first experiments were made in order to ascer* Ritthausen says that he has since found that legumic acid is only a mixture of glutamic and aspartic acids. I do not think that this opinion is
correct.




ALBUMINOID SUBSTANCES, OR PROTEIDS.  261
tain whether a part of the nitrogen of proteinic compounds was not found in the state of urea, and whether
this class of bodies does not represent complex ureids.
After having vainly sought for urea among the products of the physiological decomposition of proteids,
while ferment was kept without nourishment, I boiled
albumin, casein, &c., with barium hydrate, in an apparatus so arranged as to allow the ebullition to continue
during several successive days, without any diminution
of the water. I could easily accomplish this, by heating
it in a retort, attached to a condenser so placed as to
allow the water to return; the latter was connected with
two Wolfe's bottles, containing a known volume of
normal sulphuric acid, intended to retain the ammonia
disengaged.
Under these conditions at Ioo~ C. (212~ F.), it was
found that during the first few hours there was an abundant disengagement of ammonia, which diminished by
degrees, and at last became imperceptible.
Nasse had previously observed this fact, and had
for this reason, divided the nitrogen of the albuminoid
substances into two portions; one, the least important
in quantity would be found, as he stated, feebly fixed
or combined (Losegebtidener Stickstoff).
The barium experiment gives us the key to this
difference. In fact, at the same time that the ammonia
is set free, we see a granular precipitate formed in the
liquid which was originally clear; this precipitate increases progressively to a certain limit, and then ceases
to be formed.
It is almost entirely composed of barium carbonate,
mixed with a small quantity of barium sulphite and




262            ON FERMENTATION.
oxalate, with silica derived from the glass attacked
by the alkaline liquid, and with alkalino-earthy phosphates contained in the albuminoid matter used in the
experiment.
By measuring the amimonia and barium carbonate
formed, after boiling for I2o hours, there were found in
IOO parts by weight of dry albuminGram.
Ammonia.              I 7
Barium carbonate.. I II
By calculating the weight of carbon dioxide which
corresponds to the barium carbonate, we find that the
quantities of carbon dioxide and ammonia set free, are
almost exactly in the proportion which urea would yield,
by resolution into ammonium carbonate:CH4 N2 0 +- H2 0 = C02 + 2 NH3
CO2    44
ratio of,= - = I'29.
2 N Ha   34
We have, then, reason to admit the presence of the same
grouping of elements as in urea in albuminous substances.
Boiling at I0oo  C. (212~ F.), even prolonged for eight
days, does not entirely destroy this grouping; but albumin is resolved, under the influence of barium hydrate,
into several more simple combinations, possessing various
degrees of stability.
When, however, the temperature is raised to I40~ or
I500 C. (284~ or 3020 F.), the decomposition is complete
in a few hours (I2 to 24), and the quantities of ammonia
and carbon dioxide remain the same, even if we prolong
the operation, and raise the temperature of the mixture




ALBUMINOID SUBSTANCES, OR PROTEIDS. 263
to 2000 C. (about 39I~ F.). These experiments must
necessarily be made in a close vessel.
Under these conditions, we find in Ioo parts of dry
albuminAmmonia disengaged     4-2   A       4'5
Barium carbonate.. 25    e  e 29
These numbers give also the ratio of I'29 between CO2
and NH3.
Albumin must therefore contain, in i8 atoms of
nitrogen, about 4 belonging to the urea group.
M. Bechamp has noticed, among the products of
the oxidation of albumin by potassium permanganate,
the presence of a small quantity of urea. This fact,
which was denied by the German chemists, has been
confirmed by Ritter's experiments. However this may
be, the urea found existed in but small quantity in the
products of the reaction; and it is possible that its
production is the result of decomposition, making that
which is already formed sensible rather than the result
of an oxidation.
The barytic liquid, separated from the ammonia and
from the barium carbonate, and freed from the baryta in
excess by a current of carbon dioxide, retains but little
colour; by concentration and crystallization, followed by
treatment of the mother-liquor with alcohol, we succeed,
at last, in bringing it entirely into a crystalline form, that
is to say, to the state of a definite body. We may, therefore, by making a direct analysis of the crystals obtained,
arrive at a complete notion of the composition of albuminoid substances. The reaction of the barium hydrate,
at from 1500 to 2000 C. (3020 to 394~ F.) gives us an oppor



264            ON FERMENTATION.
tunity of trying every analytical method on the products
resulting from the splitting up of albuminoids. These
are not merely two or three defined and crystallized
bodies, which are separated- from a mass of sirupy matter
of considerable quantity, which still remains undetermined; on the contrary, we get all the compounds that
are formed, and we thus find ourselves in a position to
construct an equation of the composition of albumin
and allied substances.
Although my researches on this subject are not
entirely completed, I may already state, from the
results obtained,I. That barium hydrate decomposes albuminoid substances by simple hydratation, the experiment being
made in the absence of oxygen.
2. That the principal products of this reaction aretzhe elements of urea (ammonia and carbon dioxide, in
the proportion of I'29); traces of sulphurous acid, of
sulphuretted hydrogen, and of oxalic and acetic acids;
tyrosine C9 H" NO' (oxyphenyl, amido-propionic acid),
in very small quantity, two or three per cent. of the
albumin at the most.
We also find,30 The amiido acids of the series Cn H2n+' NO2, corresponding to the fatty acids Cn H2n 02 from amidocenanthylic acid C7 H'5 N02 to amido-propionic acid;
leucine C6 H13 NO2; butalanine C5 H"` NO2; and amidobutyric acid C4 H' NO2 abound in this mixture.
4. One or two acids nearly allied to aspartic and
glutamic acids C5 H" NO4 and C4 H7 NO4; one or two
acids analogous, and very nearly similar, to the legumic
acid found by Ritthausen C8 H14 N2 0'.




ALBUMINOID SUBSTANCES, OR PROTEIDS.  265
A smatll quantity of a substance analogous to dextrin,
which, by being boiled in acids, is converted into a body
which energetically reduces Fehling's liquid; nitric acid
changes it into oxalic acid.
With the exception of these bodies, nothing of importance is found either as a chemical compound or as
an amorphous mass.
Most of the definite compounds found by this reaction
are similar to those which have already been mentioned
among the products of the splitting up of albuminoid
substances, under the influence of acids; but I repeat
that the importance of the preceding reaction consists
especially in the demonstration that these compounds
alone constitute the albuminoid molecule; and in the
proof that the elements of urea or carbamide form an
integral part of this molecule.-(See the Memoir of the
author on Albuminoid Substances, Bulletin de la Soc.
Chim. de Paris, February I5th, March 5th, and March
i5th, I875.)
The albumin is not split up all at once in this manner
into comparatively simple products; by stopping the
reaction at different periods of its development, or by
using occasionally less elevated temperatures, we meet
with intermediate compounds, uncrystallizable, or crystallizable with difficulty, the thorough study of which will
be very interesting, in relation to the phenomena of
nutrition and of biological reactions.
The products which we obtain under the influence of
the alkalies melted in their water of crystallization
(ammonia, hydrogen, ammoniacal compounds, methylamin, saniline, picoline, petinine, leucine, tyrosine, glycocoll, carbon-dioxide, formic, valerianic, butyric, and oxalic




266            ON FERMENTATION.
acids) are derived from a more energetic action, exerted
on the first series of compounds formed by the action
of baryta. It is the same for the compounds which originate under the influence of oxidizing agents, such as the
mixture of dilute sulphuric acid and potassium bichromate
(formic, acetic, butyric, valerianic, caproic, and propionic
acids, with their corresponding aldehydes, benzoic acid
and benzoil hydride, hydrocyanic acid and butyl cyanide).
It does not enter into our programme to write the
complete history of albuminoid substances. (The reader
may consult with advantage on this subject the Dictionary
of Chemistry, by M. Wurtz; l'Histoire Generale des
Matieres Albuminoides, par G. Bouchardat; These
pour i'Agregation, Paris, I872.) We shall merely give a
summary of the theories proposed, in order to explain
the composition of these complex bodies.
All albuminoid substances, heated for some time with
alkalies, dissolve and give up sulphur to the alkali, under
the form of sulphide and hyposulphite; the neutralization of the liquid by an acid causes a voluminous floccose,
white precipitate, soluble in dilute lye-water; this precipitate has the same composition whatever may be the
albuminoid matter employed. Mulder, relying on these
facts, considered this precipitate as the base of albuminoid
substances, and gave it the name of.protein. The German chemist thought at first that his protein no longer
contained sulphur; later experiments have shown that in
reality it still contains some, but that this sulphur cannot
be removed by alkalies under the form of sulphides.
According to Mulder's theory, all albuminoid substances
are combinations of protein with variable quantities of
sulphur, phosphorus, and mineral matter.




ALBUMINOID SUBSTANCES, OR PROTEIDS. 267
This opinion, admitted very generally at first, was by
degrees abandoned, on account of the great number of
contradictory observations.
Liebig considered albuminoid substances as having
the same elementary composition, i.e., as isomeric compounds. This view, which admits of discussion, throws
no more light than that of Mulder on what is now called
the chemical constitution of these compounds, considering that protein is almost as complex in its character as
the original products. Sterry-Hunt gave an explanation
which at first sight was very seductive; he considered
albuminoid substances as amides or nitrites of cellulose,
dextrin, gum, or sugar.
The objection to this simple idea is, that it does not
agree with known facts; tyrosine, leucine, and aspartic
acid are not considered as derived from sugar, from
cellulose, or from their nitrites. These latter bodies
(nitrites of hydrocarbon substances) have been also little
studied, and are little understood.
Berthelot, in his treatise on elementary chemistry
(1872), considered, from the whole of the facts then
known, albuminoid substances to be complex amides,
formed by the union  of amido acids of the series
C" H2n1~ NO'(such as glycocoll and leucine), of tyrosine,
with certain oxygenated principles, some of which belong
to the acetic series, and others to the benzoic series.
He thought that the nature of the amides, and of
the oxygenized bodies which generated them, as well as
their relative proportions, was the cause of the differences
which exist between the various albuminoid bodies; and
that chitin and chondrin contained, in addition, the
elements of glucose.




268             ON FERMENTATION.
The results which I have obtained by means of the
decomposition by baryta, show that the albuminoids are
formed by the association, in different proportions, of the
urea and amido-acid combi~nations, some belonging to
the series of leucine, Cn H 2n+l NO2, the others, which
are more highly oxygenized, belonging to the series
Cn H"'-' N04 (aspartic and  glutamic acids);  the
more complex acids, such as the legumic, may be
considered as the products of an incomplete decomposition. Tyrosine (C9 H1 NO3) represents the aromatic
series; it is from this that benzoic and.paroxy-benzoic
acids and bromanil are derived, being obtained under
different conditions.
Besides this, it is easy to see that, with the amidoacids Cn H2n+' NO2, it is impossible, even when without
the urea, to arrive at the composition of the albuminoids; whatever combination of these bodies we may
choose, when we deduct the elements of water in sufficient proportion to obtain the value of the oxygen of
the albumin, there remains a very notable excess of
hydrogen. The intervention of the amido-acids of the
aspartic series Cn H'n-1 NO4, in the grouping of
proteids, is therefore indispensable to explain the
constitution of these bodies.




CHAPTER II.
SOLUBLE FERMENTS AND INDIRECT FERMENTATION.
IN the chemical phenomena which we have hitherto
studied, we have found the cause of the reaction connected in such an intimate manner with the presence of
a cellular organism, that all efforts made with the view
of separating this cause from this form of life, even
for an' instant, have failed, or at least have produced no
precise result definitely adopted by the scientific world.
The farther progress of science will, we hope, allow us
to penetrate more deeply into the very essence of the
phenomena.
If the remarkable and important labours of Pasteur
have taught us that the transformations of sugar into
alcohol, lactic, and butyric acid, gum  and mannite,
those of the albuminoid substances into various putrid
principles, as well as the conversion of alcohol into
acetic acid, depend on the presence of inferior organisms,
and that the germs of these organisms come from without, it is no less true that we have as yet no certain
idea of the mode of action of organic ferments. There
have been, it is true, hypotheses enough to bring about
the solution of the problem. Liebig, obliged to allow
more than he had hitherto done to the presence of
living organisms, said in his last memoir (Ann. Chim.




270                 ON FERMENTATION.
Phys. (4), vol. 23, p. 6), that, in a chemical point of
view, the only one which he will not give up, a " vital
act," is a phenomenon of motion, and that in this
sense M. Pasteur's opinion is not contradictory to his
own, and is not a refutation of it.*
We have already discussed elsewhere the theory of
M. Pasteur, propounded  as early  as  I86I, and lately
resumed with greater confidence.
Fermentation is a consequence of the life of ferments
without oxygen. These simple organisms need oxygen
so much, that when they are in a medium which is
deprived of it, they take it even from sugar and other
analogous bodies. Fermentation is therefore a consequence of the disturbance of equilibrium resulting from
this respiration.
This manner of considering the question is completely at variance with the fact that the production of
alcohol at the expense of sugar, under the influence of
a ferment, is not interfered with by the presence of
oxygen; according to Mayer, it is neither increased nor
lessened. Pasteur considers that it is rather increased
by oxygen.
It is certain that a ferment is able to live and develop
in a saccharine, nitrogenous and mineralized medium,
without the intervention of oxygen. These same evolu* M. Liebig aims here at the following sentence of M. Pasteur: —"The
chemical act of fermentation is essentially a phenomenon correlative to a vital
act, commencing and ceasing with it. There is never any alcoholic fermentation without there being, simultaneously with it, organization, development,
multiplication of globules, or the continuous consecutive life of globules already
formed." According to the German chemist, fermentation is a movement
communicated by instable bodies in process of chemical transformation; it
signifies but little whether or no these transformations take place in a liRing
organism.




SOLUBLE FERMENTS.               271
tions are more active under the influence of air. The
only inference that we can draw from these wellestablished facts is, that up to a certain point, the
sugar, which the ferment always resolves into alcohol
and carbon dioxide, as long as it can meet with it, and
unless it is exhausted, is able to supply to the ferment
the living forces required for its development. But we
know not whether alcoholic fermentation is a consequence of this borrowing of living force, or whether it
is a previous action.
The chemical cause of alcoholic fermentation set up
by yeast has been sought for from another point of
view, which must now be the object of our studies.
We know that cane-sugar, before it undergoes alcoholic fermentation, is hydrated, and thus split up, as
often occurs under the influence of acids, into two
opposite glucoses; the ordinary glucose, or grape-sugar,
which causes the plane of polarization to turn to the
right and lavulose, or uncrystallizable sugar.  This
change was at first attributed to the acidity of the ferment. M. Berthelot, and after him M. Bechamp,
showed that the active agent is a soluble, neutral, nitrogenous principle, excreted by the ferment, which is
found in greater or less abundance in the filtered water
in which yeast has been washed (the zymase of Bechamp, the alterative ferment (ferrmnet inZIversive) of
Berthelot).
This soluble principle, to which we can attribute no
organization, but which is directly derived from a living
organism, possesses the remarkable power of altering
cane-sugar in a few moments.
When this fact was established concerning the altera



272             ON FERMENTATION.
tive, soluble, inorganic ferment, analogous compounds
had been long known, also soluble, nitrogenous, and inorganic, which were especially characterized by specific
acts which they could exercise chemically on various
principles.
It was thus that M. Payen and M. Persoz obtained
from barley that had germinated, and had been afterwards ground, by treating it with water, a soluble
substance capable of turning starch into sugar. The
presence of emulsin had also been ascertained in
almonds, and this transforms amygdalin into essence
of bitter almonds.
That which especially distinguishes these chemical
reactions, set up by these various soluble, inorganic
principles, is the greatness of the effect compared with
the trifling quantity of the active agent. This same
character is found also in direct fermentation, due to
the immediate intervention of living organisms.
We will give the name of indirect fermentation to the
reactions of which we have just spoken, and whose cause
is derived from an organism, but can act without it.
It was natural to seek for the cause of direct fermentation in the intervention of active products,
whether soluble or not, elaborated by organic ferments.
Thus two orders of phenomena, which are certainly
not without relation to each other, were connected
together.
This theory, which would cause the direct or true
fermentation of M. Pasteur to be confounded, as to
its essence, with indirect fermentation, or fermentation caused by inorganic soluble ferments, has not
found sufficient support from characteristic and well



SOLUBLE FERMENTS.               273
observed facts, to be established as a truth that has
been demonstrated; but it has not yet been absolutely
contradicted. It is, however, easily to be seen that
such an explanation becomes useless. If the decomposing force is derived from the cell, what would be
the utility of this intermediate soluble or insoluble
solid substance which transmits the force?
Gzneral Characters of Soluble Fermazents.-Soluble ferments are all derived directly from living organisms in
the midst of which they originate. Up to the present
time, the specific characters, of which we shall presently
speak, have not been communicated to any artificial
organic substance.  We are, therefore, compelled to
believe that this specific character is a consequence of
the origin of soluble ferments. Their composition resembles that of albunilnoid substances; in fact, they contain carbon, nitrogen, hydrogen, and oxygen. But the
analogy will go no farther. When we have eliminated,
by proper processes which we are about to describe, the
albuminoid substances which always accompany soluble
ferments in their first solutions, we find that the product,
though it preserves all its chemical activity, no longer
manifests the general reactions of albuminoid substances. It no longer yields a precipitate with tannin
and corrosive sublimate; iodine and nitric acid no
longer colour it.
Elementary analyses have also revealed sensible differences; but as there was no proof that the products
analyzed were pure, we can draw no conclusion from
this concerning the chemical nature of soluble ferments.
It is probable that they are derived from the physiological splitting up of proteids.




274             ON FERMENTATION.
The soluble indiirect ferments, or  2symasrs, present
themselves, when in a dry state, as amorphous, colourless, pulverulent matter; they are usually precipitated
from their aqueous solutions by alcohol, corrosive sublimate, neutral or basic lead-acetate. These precipitates,
when decomposed by sulphuretted hydrogen, restore to
the water the soluble matter unchanged, and preserving
its specific properties; but it is still, in this case, accompanied by a greater or less quantity of albuminoid matter.
If the sublimate precipitates them from liquids extracted from the organism, it is rather by mechanical
action than by the fact of a chemical combination; for
we have just seen that soluble ferments, when deprived
of albuminous principles, are not precipitated by the
sublimate.
By taking advantage of the facility with which the
zymases are mechanically carried down by solid deposits
in process of formation in the midst of the liquid which
contains them, certain chemists have discovered means
of purifying them.  Thus Conheim  separates pure
ptyalin (salivary diastase) by acidulating the saliva
strongly with tribasic phosphoric acid; the phosphoric
acid is subsequently neutralized by lime-water, until an
alkaline reaction takes place. The precipitate of tricalcic phosphate carries down with it, as it is formed, the
ptyalin and albuminoid matter; the filtered liquid has
no action on starch. The deposit, washed with water,
yields the ptyalin to the solvent, and retains the proteinic matter; it is then only necessary to'precipitate it
by alcohol, and we obtain a white, light, flaky deposit,
which, dried in vacuo, presents itself in the form of an
almost colourless powder.




SOLUBLE FERMENTS.                275
Pure pepsin is extracted in a similar manner from
natural or artificial gastric juice (obtained by exposing
the mucous membrane of the stomach, separated from
the muscular membrane, and cut up, to a temperature
of 350 C. (950 F.), with water containing 5 per cent. of
phosphoric acid. This juice, which contains phosphoric
acid, is precipitated by lime-water. The tricalcic phosphate is transformed, by the addition of a proper quantity of phosphoric acid, into insoluble and crystallized
bicalcic phosphate, which it is only necessary to wash in
order to remove the pepsin adhering to it. We may also
dissolve the precipitate of calcium phosphate in dilute
hydrochloric acid; then pour into the liquid a solution
of cholesterin in a mixture of four parts of alcohol and
one of ether; shake up with the liquid the cholesterin
which is separated, and collect it on a filter; wash it
with water acidulated with acetic acid, then with pure
water.  The damp cholesterin, to which the pepsin
adheres, is treated with pure ether, which dissolves it,
while there remains in the lower part of the vessel a
solution of pure pepsin in water (Briicke). This pepsin
yields no precipitate except to platinum bichloride, and
neutral and basic acetates of lead. Nitric acid, tannin,
and corrosive sublimate are without effect. It only yields
a very slight colour with nitric acid and ammonia.
Danilewsky makes use of collodion to precipitate
soluble ferments. The precipitate, having been well
washed, is treated, after being dried, with a mixture of
ether, alcohol, and water, which dissolves the nitrated
cellulose, leaving in solution the active principle, free
from albuminoid matter.
Von Wittich (Arch. f. d. ges. Physio.,-vol. 3, p. 339),




276             ON FERMENTATION.
proposes the following method, applicable to the extraction of soluble ferments in general. The vegetable or
animal organ which contains them is rapidly cut in
pieces, cleared from blood, if necessary, by washing in
water, and then left under alcohol for twenty-four hours;
afterwards dried in the air, pulverized and sifted.
The powder is diffused in glycerin; and this glycerin
solution is precipitated by alcohol. By repeating this
operation several times (solution in glycerin and precipitation by alcohol) an active powder, free from albuminoid
matter, is obtained.
We have but little information respecting indirect
ferments, regarded in a chemical point of view. We
cannot say whether or no they have the same composition, and only differ by their specific activity. That
which gives considerable importance to these products
is the transforming power which they exercise over a
great number of organic compounds, and the important
part which they play in many physiological reactions.
The activity of an indirect ferment depends on the
temperature, like that of organic ferments. In general
terms, we may say that it increases with the temperature
up to a certain limit, beyond which it undergoes a rapid
depression, till it ceases altogether. This limit varies
with the nature of the ferment; it is always under
Ioo0  C. (2I2~ F.), and is found to be higher than that
of organic ferments.
The action of chemical agents on soluble ferments
is also not quite to be compared with that exerted on
organic ferments.
Thus M. P. Bert has observed that compressed
oxygen destroys the latter ferments after a longer or




SOLUBLE FERMENTS.               277
shorter interval, while soluble ferments are not modified in their activity.  This interesting experiment
establishes a very distinct line of demarcation between
the two kinds of-ferments, and may serve to classify
them, whenever microscopical indications leave any
doubt as to their character.
M. Bouchardat (Ann. Chim. Phys. (3), vol. I4, p. 6I),
who has carefully studied the influence of chemical
compounds on diastase, has observed that certain substances, which are antagonistic to alcoholic fermentation, have no influence on the effects of diastase; such
as prussic acid, the mercurial salts, alcohol, ether, chloroform, and certain essences (cloves, turpentine, lemon,
mustard, &c.)  Citric and tartaric acid, which only
slightly interfere with alcoholic fermentation, completely
destroy the activity of diastase.
M. Dumas (Comp. Rend., vol. 75, p. 295), made
special experiments on the action of borax on this
class of ferments. He found that a solution of borax
coagulates beer-yeast; the supernatant liquid has lost
the property of altering cane-sugar; it also neutralizes
the action of the water of yeast on saccharose. If we
place sweetened water and the water of yeast in one
tube, and sweetened water with yeast-water and a
solution of borax in another, the first will soon show
signs of alteration,- while the second will manifest none.
Analogous effects are observed with synaptase or emulsin,
diastase, and myrosin. All these ferments cease to act
on amygdalin, starches, and myronic acid, from the
moment that they are placed in contact with a solution
of borax. This salt appears, then, to have a specific
action in destroying the activity of all soluble ferments.




278             ON FERMENTATION.
We have seen, on the contrary, that yeast, placed in
contact for three days with a saturated solution of
borax, is able still to set up alcoholic fermentation.
Borax may then, like compressed oxygen, serve as a
differentiating character of soluble and organic ferments.
Bodies, or Chiemical Comzpounds, on which Soluble Ferments are able to Act, and the Kind of Reactions which
they Excite.-Organic ferments exert their activity on
a great number of organic compounds belonging to
different groups.
The glucoses, acids rich in oxygen, such as malic,
tartaric, citric or lactic acids, albuminoid substances,
urea, and alcohol, can be affected by organic living ferments. The reactions which these ferments cause them
to undergo are often very complex, and cannot be formulated by simple equations, provided that we wish to
represent correctly all the terms; in fact, in almost
every case, we have not been able to reproduce artificially the complex conditions of organic matter under
the influence of organic ferments.
Indirect or soluble ferments are able to act also on
various classes of organic compounds, but the mode
of action is generally the same. There is a more or
less simple splitting up accompanied by a hydratation. The nature of this splitting up is always conformed to the peculiar constitution of the compound,
and may be explained, in most cases, by chemical processes in which the direct or indirect intervention of a
living organism cannot be brought in.
Thus starch is' resolved by hydratation into glucose
and dextrin, and this in its turn is converted into
maltose, as well under the influence of diastase as




SOLUBLE FERMENTS.               279
by being boiled with a dilute acid (sulphuric acid).
The alterative ferment hydrates a molecule of saccharose, and converts, it into two molecules of glucose:
dilute acids behave in the same manner.
Certain special soluble ferments, such as synaptase
or emulsin contained in sweet and bitter almonds, and
the myrosin of white or black mustard, act on natural
glucosides; that is to say, on those special compounds
that are met with in plants in such great abundance,
and which we may consider as ethers composed of
glucoses, or of polyatomic alcohols. The result is a
splitting up, by hydratation, into glucose and another
principle.  Chemical means, such as boiling with an
acid or an alkali, lead to the same result.
The fatty bodies (ethers compounded with glycerin),
as is well known by Chevreul's beautiful experiments,
fix the elements of water when they are heated with
boiling alkalies, or with acids, and yield glycerin and a
fatty acid, the sum of the weights of which is equal to
the weights of fatty matter used, plus the water fixed
by the reaction.
But we know by the works of M. C. Bernard, that
the pancreatic gland secretes a soluble nitrogenous
substance, capable, like boiling alkalies, of saponifying
greasy substances.
It is very probable that the digestion of albuminous
matters, and their conversion into peptones, under the
influence of the gastric or pancreatic juice, or rather
under that of soluble ferments contained in the secretions, are only the results of a hydratation and a
splitting up, the conditions of which we are able to
realize without the agency of life.




280             ON FERMENTATION.
We may almost foresee that, in general, every phenomenon of the splitting up of an organic compound
into its proximate constituent parts, requiring only a
simple fixation of water, will find in the organism
itself its soluble ferment, that is to say, the agentcapable of producing this decomposition. It is thus
that ethers composed of mon-atomic or polyatomic
alcohols, including the glucosides and fatty bodies,
certain complex acids, such as the hippuric, glycocholic, taurocholic, &c., are resolved into two or more
simpler molecules.
Can the same soluble ferment act upon different
chemical compounds, decomposing them in the manner
indicated by the peculiar constitution of each? There
is not the shadow of a doubt as to the positive answer
to this question. Thus we see emulsin or synaptase,
contained in almonds, cause the decomposition of a
great number of crystallizable and definite principles
of the vegetable organism, such as amygdalin, salicin,
arbutin, helicin, phlorizin, esculin, daphnin.
It is true that these various bodies belong to the
same family, and have the same functions; they are
all glucosides, and we must not be more surprised to
see the same force transform them than to observe the
saponification of neutral fatty bodies under the same
influences.
In certain cases, we see the same organic liquid, the
same secretion, such as the pancreatic juice, exert its
active power upon very different principles, which do not
present, as in the previous example, an analogous constitution. The pancreatic juice transforms, modifies,
and digests albuminoid substances, turns starch into




SOLUBLE FERMENTS.              28I
sugar, and saponifies fatty substances. We may well
ask if this multiple specific power belongs to one and
the same principle, or whether it indicates the presence
of several distinct soluble ferments in the pancreatic
juice.
The experiments of Cohnheim and Danilewsky tend
to prove that the various physiological properties of the
pancreatic juice are due to special principles. These
chemists have, indeed, been able to separate by precipitation, by means of calcium phosphate or collodion, a
nitrogenous substance, which is not albuminoid, and
which shows all the characters of salivary diastase, and
which rapidly turns starch into sugar, without digesting
the proteids.
Thus, when we pour the ethereal alcoholic solution of
collodion into the extract of the gland, obtained by
pounding it with magnesium carbonate and water, and
afterwards filtering it, the ferment of albuminoid substances is precipitated with the collodion, while the
amylaceous ferment remains in the liquor, and may
be obtained by evaporation. Calcium phosphate precipitated from  an aqueous solution of the pancreatic
gland (by phosphoric acid and lime-water), carries down
with it the diastasic ferment; while, on the contrary,
the albuminoid ferment remains in solution. If, in the
experiment made with collodion, the precipitate be redissolved, washed in water, and dried in aqueous ether, we
obtain two layers; the lower and watery one contains
the albuminoid ferment in solution. As to the ferment
which saponifies the fatty matters, it has not yet been
obtained by itself, and independently of the two others.
L imits of the Activity of a Soluble Feriizent.- Certain
13




282             ON FERMENTATION,
experiments tend to prove that soluble ferments have
not an unlimited activity. It is known that, with a
determinate weight of one of these bodies, we can
modify an incomparably greater quantity of fermentable
matter; but does the ferment always lose its power after
a time, and become exhausted? The limit of activity
has been determined for only a small number of these
ferments, and the results published warrant but very
undecisive conclusions. Most of the experiments of this
kind have been made with impure ferments, still containing albuminoid substances.
Thus, according to Payen and Persoz, one part only
of diastase is sufficient to liquefy and turn to sugar 2,000
parts of starch; but, on the one hand, the diastase prepared by alcoholic precipitation, according to the process
employed by these observers, is a complex mixture, into
which the really active substance enters, perhaps, only in
a very small proportion. The ratio of I to 2,000 would
thus become much smaller. On- the other hand, it has
been ascertained that the presence of a certain quantity
of glucose interferes with the transformation of dextrin;
and in order to allow the action to resume its course,
it is only necessary to dilute the liquid with much water,
or to remove the glucose as it is formed, by causing it
to undergo an alcoholic fermentation by means of
yeast.
According to M. Berthelot, the alterative ferment of
yeast is able to alter from 50 to IOO times its weight of
sugar. We ought to make the same remark about these
numbers as we did respecting diastase. Nothing, therefore, proves decidedly that soluble ferments lose their
specific power as they exercise it. The opposite opinion




SOLUBLE FERMENTS.              283
finds some support in the infinitely small quantities of
the ferments necessary to produce a very important
and considerable effect. It is also evident that these
decompositions are effected rather with disengagement
than absorption of heat, since the same decomposition
by hydratation takes place by the action of sulphuric
acid in small quantities, and the same amount of acid
will act indefinitely. They do not, therefore, require the
employment and the consumption of living forces, and
the principle of the conservation of energy is not opposed to the idea of an indefinitely prolonged action.
This manner of considering the question does not
exclude the possible, and even probable fact, shown in
certain cases, of progressive weakening of the ferment,
after which it will have lost its specific power: a similar
chemical decay may accompany the manifestation of
the specific power, or be produced without it, in an
independent manner, and without being a consequence
of it.
Particular Studies of Solubie Ferments or Zymases.After these general considerations, we will notice what
speciality is presented by each indirect ferment. In this
particular study, we will not return to the physical
characters, the chemical composition and properties of
zymases, nor to the method of preparing them in greater
or less purity. These different questions have been fully
treated in the general study of these bodies, and we
should expose ourselves to needless repetitions by bringing forward again for each ferment that which applies
to the whole class.
The points which will claim our attention are the
chemical reaction, the different sources of the ferment




234              ON FERMENTATION.
which excite it, and the special conditions which
favour it.
I. Diazstases.-We shall give the general name of
diastases to ferments capable of turning starch into
sugar.
If we only regard the original and the final terms of
the reaction, we arrive at this conclusion, that starch,
under the influence of diastase, combines with itself the
elements of water, and is converted into glucose.
C6 H1~ 05 + H2 0 = CG H12 06
Amylaceous inatter. Water.  Glucose.
Yet, according to the researches of Musculus (Ann.
Phys. Chim. (3), vol. 55, p. 203;Bulletin de la Soc. Chim.,
Paris, vol. 23, p. 31, 1874); those of Schwarzer (Bull. Soc.
Chim., vol. 14, p. 400); those of Schultz and Maierker
(Bull. Soc. Chim., vol. I9, p. I7), and those of Payen, the
change does not take place so simply. By previously
warming the starch with water at 700 C. ( 580 F.), before
adding the diastase, the reaction by means of iodine
disappears at the moment that the saccharification has
affected a quarter of the liquid.*   But the action of
diastase does not stop there; if we add a larger quantity,
it continues till it has reached the half (5i per cent.,
according to Payen; from 5 I to 5 I'7 per cent., according
to Schultz and Maerker); that is to say, that the starch
is converted into a mixture of glucose and dextrin in
equal equivalents. From this moment the saccharification stops, notwithstanding the addition of greater
quantities of diastase.
With soluble starch the action is the same. The:= Not the third, as Musculus had previously announced.




SOLUBLE FERMENTS.                          285
reaction by iodine disappears as soon  as a quantity of
glucose, equivalent to a quarter of the starch, has been
formed; but with an excess of diastase, glucose is rapidly
and easily formed at the expense of half the starch.
According to these results, and without taking into
consideration the disappearance of the peculiar colour
produced by iodine, when the quantity of sugar is equal
to one-quarter, a fact which must be interpreted by fresh
experiments, we may suppose that the amylaceous
molecule is resolved by hydratation into two molecules,
one of glucose, the other of dextrin.
2 (C6 Hl~ 05) +  H2 0 =  C6 H0 05 +  C6 H12 06
Starch.       Water.      Dextrin.       Glucose.
M. Payen has shown long since, that by the intervention
of yeast we produce the entire transformation of starch
into glucose, and finally into alcohol and carbon dioxide.
This experiment would prove that the presence of
glucose is antagonistic to the action of diastase upon
dextrin,
Thus starch, soluble starch, dextrin, and, according to
the researches made by C1. Bernard, the glycogen matter
of animal tissues can be changed into sugar under the
influence of diastase.*
* A diastase of considerable activity may be obtained by a modification of
the process of MM. Payen and Persoz. One part of germinated barley in
powder is added to two parts of water. After an hour's maceration, the liquid
is pressed out, and an equal volume of alcohol at 800 C. (I760 F.) is added to
it. It is then filtered, and the first rather voluminous precipitate is rejected.
An equal volume of alcohol is again added to the filtered liquid. A very slight
precipitate is then formed, which is collected on a filter; this filter is dried
with the precipitate at a gentle heat. A diastase paper, which is very active, is
obtained in this manner, and.nmay be indefinitely kept without change. (MusculI1S.)




286             ON FERMENTATION.
Diastasic reactions play an important part in the
animal organism, and especially in the digestion of
starchy food. Thus we find diastasic ferment in the
animal economy as well as in the vegetable tissues. At
its first entrance into the digestive tube, the food is mixed
with the saliva, a liquid which contains, as shown by
M. Mialhe in 1845, ptyalin, a fermenting principle analogous, or rather identical, with the diastase of germinated
barley. MM. Bouchardat and Sandras proved nearly
at the same time the existence of an analogous agent in
the pancreatic juice.
The pancreatic juice acts infinitely more energetically
than saliva in the saccharification of starchy substances.
The action of an infusion of pancreatic juice upon starch
is excessively rapid-indeed, we may say, instantaneous;
it is, in fact, in the small intestine that the principal digestion of amylaceous substances is effected.
This same ferment is met with in other parts of the
organism; everywhere where starch, whether animal or
vegetable, must be rendered soluble. Thus, in the liver
there exists a kind of animal starch, glycogen, which is
turned  into sugar by contact with  blood, and  a
ferment, in order to be conveyed in this form into the
torrent of the circulation. At the period of animal
life when this change is to be accomplished, the ferment
appears, and the accumulated starch is destroyed. M.C1.
Bernard (Digestion comparee chez les Animaux et les
Vegetaux, Revue Scientifique, 1873, p. 5 5) draws a
very striking parallel between the chemical phenomena
of nutrition in animals and vegetables.
"The ferment appears in seeds from the commencement of germination; this talces place in the potato in




SOLUBLE FERMENTS.                  287
spring; then the fermentiferous agent shows itself in the
tubercle, as it did in the germinated barley; it liquefies
the starch, and puts it into a condition to be distributed
over the parts where it may minister to the nutrition,
that is to say, to the development of the life of the
plant.
"In most animals, the phase of the production of
glycogen, and that of its fermentation, are not so distinc
as in plants. The two phenomena are often continuous
and simultaneous. An exception must, however, be
made with respect to the earliest periods of life,
especially in those animals which undergo metamorphoses. For example, if we consider the larva of the
common fly, mzusca hZlcelia, the gentle-to call it by its
common name- we find that it contains an enormous
quantity of starch. It is truly a bag full of glycogen.
At this time we find nothing else but glycogen, and not
even a trace of sugar. The reason of this is that the
glycose ferment does not yet exist.  But soon the
chrysalis will succeed to the larva, and then in the new
phase of existence, during which the perfect animal is
in formation, the reserve of glycogen must be utilized.
The ferment appears, and the starch is liquefied.
"Something analogous to this is seen in beings of a
higher order, for eyample, among the mammalia, at
that period of the embryonic life when nutrition is
hastened, and the plastic and formative activity attains
its highest degree. The glycogen matter deposited at
various points of the fcetus, and of its envelopes, is set
in motion, dissolved, and transformed into sugar. *
i The experiments of C1l. Bernard, Hensen, Magendie, and Schiff prove that
in living blood there exists a soluble ferment, capable of transforming starch




288                 ON FERMENTATION-.
"The digestion of starches consists in their transformation into soluble and assimilable substances, soluble in order to be able to circulate from  one part of
the organism to another. Digestion is, therefore, the
prologue of the act of nutrition. Whenever these
starchy matters are to nourish an organism, we shall
find this previous preparation. But all organisms, in
the vegetable as well as in the animal kingdom, employ
starchy substances as food, all, therefore, digest these
substances, in the strict sense of the word."
The agents in these acts of digestion are the diastase
of germinated barley, saliva, pancreatic juice, &c.
The action of diastase on  starch is exerted  at the
ordinary  temperature, and attains its maximum  effect
at 75~ C. (I67~ F.). Boiling destroys the specific power
of this substance.
It is almost useless to mention, since the fact is so
well known, that the effects of diastase on starch  are
imitated by the intervention of boiling dilute sulphuric
acid.
Al4terative FerZeZt.-Cane, sugar, Qr saccharose, C"'
H122 01, is transformed, as is proved by the researches
of M. Dubrunfant (Ann. de Chim. et  de Phys. (3),
vol. 21, I69, I847; Comp. Rend. de l'Acad. vol. 29, p. 5I,
1849, and vol. 42, p. 90I, I856) by, becoming hydrated,
tunder the influence of dilute mineral and even organic
and glycogen into sugar. This ferment is also found in very fresh liver. It
may be eliminated from it by making a cold infusion of a liver, which, on
account of general disease, produces no more sugar, and contains no glycogen,
and by precipitating it subsequently by alcohol. The deposit, redissolved in
water, acts on starch. This special ferment exists in the blood of frogs in spring
and in summer. In fact, dextrin injected into their veins passes with the urine
in the form of sugar. In winter it is wanting, for dextrin appears in the urine
without alteration.




SOLUBLE FERMENTS.                289
acids, into sugar, which causes the plane of polarization
to turn to the left, and energetically reduces Fehling's
cupro-potassic reagent.  Saccharose, on the contrary,
turns the plane of polarization to the right, and has
no action on Fehling's liquid. The alteration is produced in the cold, but more rapidly in the warm process; it has been effected by simply boiling solutions
of cane-sugar in water, by their exposure to light, or
by the mechanical means. Thus the mere pulverizing
the sugar causes a small portion of altered sugar to be
formed. The product of the alteration is not a unique
and simple principle.
The same, chemist proved that saccharose, after
alteration, is found to be changed into two sugars of
the formula C' Hl2 06, one of which turns the plane of
polarized light to the right, and is identical with grapesugar, or ordinary glucose; the other, on the contrary,
causes the plane of polarization to deviate to the left;
this is the uncrystallizable sugar of acid  fruits, or
icezlvose.
As at I5~ C. (59~ F.), the specific rotatory power of
glucose is equal to + 57.80, whilst that of laevulose
is - Io6~, and as the two sugars are found in the mixture in equal proportions by weight, it can be understood that this mixture, called altered sugar, has a
laevo-rotatory action due to the excess of one of these
powers over the other.
Altered sugar has a lIvo-rotatory power of 25~ at
the temperature of I5~ C.
The formation of these two sugars (glucose and
lavulose) is easily formulated by the following equation:



290               ON FERMENTATION.
C12 H"1 0"l + H2 0 =  CO H12 66 + CO Hi2 0'.
Saccharose.   Water.    Glucose.    Laevulose.
This is confirmed by the fact, that under the influence
of heat, saccharose is.changed into glucose and laevulosan (levulose less water), which is itself changed into
levulose under the influence of acids.
The following is the manner in which M. Dubrunfant
brings about the separation of these two bodies.
Ten grammes of altered sugar are dissolved in Ioo
grammes of water,* and the whole heated with six
grammes of calcium hydrate. The mixture, which at
first is fluid, forms into a mass at the end of a short
time; this is the calcium laevulosate, which separates as
it crystallizes, while the glucosate remains in the motherliquor. The mass is pressed at once, and the solid
and liquid parts, after having been dissolved or diluted
with a suitable quantity of water, are decomposed
separately by carbon dioxide; the former furnishes a
solution of lavulose, the latter of glucose. We can also
effect a separation, or at least isolate  the laevulose
free from glucose, by stopping a fermentation of canesugar almost in the middle of its course.  The glucose
fermenting more easily than the Ikevulose, the latter is
found alone.
Without dwelling any further on all these proofs of a
true splitting up, which leave no doubt as to the nature
of the reaction, let us proceed to the ferment.
This alteration, this splitting up of the saccharose by
* According to the experiments made at the laboratory of the Sorbonne, the
proportion of water indicated by Dubrunfant is too great to allow of the crystallization of the calcium laevulosate. Better results were obtained by employing four parts of water to one part of altered sugar.




SOLUBLE FERMENTS.                        29T
hydratation is produced before any alcoholic fermentation takes place. Cane-sugar can not be resolved into
alcohol and carbon dioxide till after it has been altered,
but beer-yeast itself  performs  this  operation.   The
action was, at first, attributed to the acidity of the
yeast, and M. Pasteur thought that it was the result of
the presence of succinic acid.
M. Berthelot was the first to show  that the alteration
of cane-sugar by yeast is independent of the  conditions of acidity and neutrality; that it is due to the
intervention of a special soluble ferment contained in
the cells of yeast. This principle is found in the water
in which yeast has been washed, and acts energetically
upon sugar, even in a neutral liquid.
The properties of this soluble ferment, besides its
peculiar activity, cause it to resemble all the bodies of
this class. I have ascertained that it is easy to separate it entirely from the albuminoid substances which
accompany it, by following the method of precipitation
by phosphoric acid and lime-water mentioned above.*
Saccharose is not susceptible of being absorbed and
assimilated under its original form, any more than amylaceous matter, although it has the advantage over the
latter in its solubility. Like starch, it has to be digested,
and the products of this digestion are glucose and lkevulose.t   M. Bernard  has proved  the existence  in  the
* M. Bdchamp found, after M. Berthelot, the alterative ferment (zymase) in
the water of yeast, spontaneously weakened by want of nourishment. Under
these conditions, I have myself observed that a liquid is obtained, excessively
active, and which produces alteration almost instantaneously.
t Cane-sugar is, according to the expression of M. Bernard, like an inert or
indifferent matter which can circulate with impunity in the blood or the sap,
without the anatomical elements being able to turn it aside or to appropriate it.
He gives as a proof of this, that when cane-sugar is injected into the veins or




292               ON FERMENTATION.
intestinal juice of a soluble alterative ferment, similar
to that of yeast; it forms one of the most important
and useful elements of this secretion. In order to prove
this, a solution of pure saccharose (not reducing Fehling's
liquid) must be injected into a portion of the intestine
included between two ligatures, or placed in contact
with an infusion of intestinal mucous membrane; we
then see, in a very short time, that the sugar reduces the
copper oxide. It may be ascertained by the saccharometer that the deviation of this physiologically altered
sugar is the same as that of chemically altered sugar.
M. C1. Bernard has ascertained the existence of alterative ferment in dogs, rabbits, birds, and frogs.  M. Balbiani has proved that it is to be found in the digestive
tube of silkworms. M. Bernard, by comparing these facts
relating to the digestion of saccharose in animals, with
the analogous transformations observed in the vegetable
economy, as he had before done for the saccharification
of starch, shows that the alterative ferment is generally
found in all parts of the living economy, and under all
the circumstances in which saccharose is necessary for
nutrition. Sugar-cane which is flowering, and beet which:runs to seed, transform by alteration the sugar laid up in
their tissues. The agent is always exactly the same, an
alterative ferment. M. Bernard obtained it from beetroot in course of evolution, as M. Berthelot had extracted
it from yeast. " Alcoholic fermentation," says our illustrious physiologist, "is a phenomenon correlative to the
nutrition of an organism, the  yeast of beer, torula
cellular tissue of an animal, it is returned, weight for weight, in the urinary
excretion, and consequently passes through the economy, without being modified or assimilated.




SOLUBLE FERMENTS,                 293
cerevisiw. But saccharose is unfit for the nutrition of this
microscopic form of life, as it is unsuited to that of
higher forms. It is therefore necessary that saccharose
should be modified, and transformed into glucose, before
it can conduce to the vital changes of the organic
ferment.  The cell of yeast, by producing this transformation, tends to its own development.  It digests the
saccharose for itself; alteration is here once more a
digestive action of the same nature as those which we
have already examined.
" In the same manner as starch, the saccharose, which
exists in a state of reserve in the tissues of a great
number of plants, is unfit to participate in the nutritive
circulation of the-plant.  It is for this reason that sugar
can accumulate and be laid up in store, as occurs in the
root of the beet, and in the stem of the sugar-cane.
The sugar forms a reserve there, while it waits for the
moment to begin to act.  This period arrives for beetroot, when it is about to put forth buds, flowers, and
fruit; then the sugar diminishes by degrees, and disappears soon after from the tissue and the stalk of the
beet, changing into glucose. The leaves at this time
contain glucose only; the root yields up its store, and
the reserves of sugar which it contained are distributed
through the stalk to serve for the purposes of the flower
and seed; but this is not possible, except by means of a
previous transformation, which changes the chemical
nature and the composition of the saccharose and causes
it to pass into the state of glucose. This is again a true
digestion.  Beet, therefore, like yeast, and like animals,
must digest its sugar."
I could not resist the pleasure of giving the very words




294             ON FERMENTATION.
of our great observer. Nothing can, in fact, give a
clearer idea of the importance of the subject of which
we are now treating, than the wide scope of the considerations by which these phenomena are connected
with the entirety of the great act of nutrition in living
organisms. We have not here to deal with isolated
chemical reactions, interesting on account of the obscurity which still reigns over the cause which produces
them, but with transformations which play an important
part in the acts of life -and nutrition, the remarkable
generality of which is placed in the most striking point
of view.
Enmulsizve and Sapozifying Fermcnt.-M. C1. Bernard
has shown that the pancreatic juice is the only one
among all the liquids secreted in the digestive tube,
which possesses the remarkable property of forming an
emulsion, and afterwards saponifying neutral fatty substances. It constitutes, therefore, the true agent of
digestion for fatty substances or natural glycerids.
The emulsion which precedes saponification is rather
a physical than a chemical transformation. It is the
mechanical division of the fatty liquid which is thus
separated into an infinite number of little globules,
maintaining themselves by means of a characteristic
molecular constitution. Under the microscope, we see a
great number of' granular bodies swimming in the liquid,
and animated by the Brownian movement. Emulsion
is the necessary condition for the absorption of fatty
bodies.
Among the organic secretions of the digestive canal,
the pancreatic juice is the only one which can furnish
with oils a complete and  persistent emulsion.  M.




SOLUBLE FERMENTS.                 295
Bernard attributes this emulsive power to a peculiar
soluble ferment. He considers that the persistent emulsion of the fat in milk is due to the presence of an
emulsive ferment, and that this nutritive liquid contains
the fatty matter ready prepared for absorption.
Emulsion by the pancreatic juice or the extract of the
gland is instantaneous. A slower action ensues; this is
saponification, that is to say, resolution of trimargarin,
triolein, tristearin, &c., by hydratation, into glycerin and
fatty acids. This resolution is not always effected in the
intestine. The emulsion penetrates into the chyliferous
vessels still retaining its lactescent character, and it is
not till afterwards, in the different tissues, that saponification takes place.  It has been thought.that the saponification excited by the pancreatic juice was due to the
alkalinity of this liquid. M. Bernard replies to this
objection by showing: (I) That other secretions quite
as alkaline are inactive; (2) That the tissue of the
pancreas, which has no alkaline reaction, rapidly produces phenomena of the  same  kind;  (3) We can
destroy by boiling, the specific power of the juice, without
weakening its alkalinity. The emulsive ferment may be
eliminated, mixed with albuminoid matter, and two other
indirect ferments, of which we have already spoken (pancreatic diastase, and the digestive ferment of the albuminoids). The emulsive ferment possesses the property
of casein, of being precipitated, in the cold, by magnesium sulphate. It is coagulated by heat.
We find bodies of the same order in plants. If we
steep oleaginous seeds in water, we obtain an emulsion
in which we soon find glycerin and the free fatty acids.
During the germination, the emulsive and saponifying




296              ON FERMENTATION.
ferment, placed with fat in the presence of water, causes
it to undergo true digestion, and renders it assimilable.
The emulsine of almonds has an analogous property;
we shall presently see that it acts on certain glucosides.
We cannot determine whether this complicated action
depends on a mixture of several ferments, or on a variable activity of the same body, according to the elements
subjected to it.  K6lliker and MUller have shown that
the pancreatic juice can effect the decomposition of
amygdalin. This is a very important indication.
Perhaps we shall find, when we have succeeded in
isolating the emulsive ferment of the pancreatic juice,
that it has the same properties as emulsin or synastase.
As a conclusion of that which relates to the fermentation of glycerides, we have only now to obtain definite
ideas as to the kind of reactions which take place, by
giving the equations of the decompositions, as the result
of the labours of M. Berthelot has shown them.
I. Emulsion; a physical change.
2. Splitting up.
C3 HS (C]8 H3' O. 0)3 + 3 H'+ O = C' -is 03 + 3 (C"' H33 0. HO)
Triolein.     Water.    Glycerin.    Oleic acid.
C3 H5 (C18 H35 0. 0)3 + 3 H' O  C3 H8 O- + 3 (C8 H5 O. HO)
Tristearine.            Glycerin.   Stearic acids.
Cs H5 (C4 H7 O. 0)' + 3 H   = -C3 H8 03 + 3 (C4 H7. HO)
Tributyrin.             Glycerin.   Butyric acid.
At/buziviosic Fermzclt.-Two digestive secretions, the
gastric and the pancreatic juices, and probably the
intestinal juice also, possess the power of transforming
the soluble or insoluble but indiffusible albuminoid
substances into soluble and diffusible principles.  It




SOLUBLE FERMENTS.               297
has been found that this property is due, as is the case
with the digestion of starchy bodies and saccharose, to
the intervention of special nitrogenous principles, to
which the name of pepsin has been given (the digestive
principle of the gastric juice), and to that of the albuminosic ferment of the pancreatic juice.
Pepsin, the method of extracting which we know
(processes of Schwann, Wasmann and Vogel, and de
Briicke), only acts on albuminoids in an acid medium,
and within very restricted limits of temperature. Its
action is principally exerted on fibrin. The short time
that food continues in the stomach only allows, in any
case, very slight transformations; and we must, indeed,
admit, that the stomachic, acid, pepsinic digestion of
albuminoid substances is, in every respect, a very incomplete operation, and rather preparatory than final.
Notwithstanding all that has been written and said
upon the chemical question, but very little is yet known
about the real transformations of the albuminoids in the
stomach, or under the influence of artificial gastric juice;
we will not, therefore, enter into long details on this subject; they would only give us superabundant proofs that
everything remains to be done. In fact, until the constitution of albuminoids is definitely ascertained, and we
know all the intermediate terms of their transformations
and progressive splittings up, we shall not be able to enter
on this question with advantage and success. I shall
therefore content myself with a few observations.
Action of the Gastric luice o0n the Fibrin of Blood.According to BrUcke, uncooked fibrin, placed in contact
at 40~ C. (IO4~ F.) with a sufficient quantity of gastric
juice of good quality, swells and dissolves rapidly,




298             ON FERMENTATION.
changing at first into a principle soluble in dilute acids,
but capable of being precipitated by neutralization; this
principle seems to have all the characters of syntonin,
the product of the transformation of muscular tissue
under the influence of acids. By prolonged digestion,
the syntonin disappears in its turn, and gives place to a
unique principle, peptone. It is doubtful whether peptone
has time to form in the stomach.
Meissner, on the contrary, asserts that the syntonin,
which originates at first, splits up by an ulterior action
into an insoluble indigestible substance, parapeptone, not
capable of being subsequently transformed by the gastric
juice, and into soluble principles (peptones a b c). The
acid stomachic digestion seems, according to these views,
to be a decomposition of proteinic substances.  This
manner of considering the question appears to me the
most probable, and the most conformable to the analogies that we can find, according to what is known of the
reactions of the albuminoids.
Other albuminoid substances behave, according to
Meissner, nearly in the same manner as fibrin, by becoming previously changed into syntonin,
The digestive action of the pancreatic juice, with
reference to albuminoids, is much more energetic and
efficacious than that of pepsin. It does not require
such limited conditions. Thus the albuminosic ferment
of the pancreas develops its power as well in an acid as
in an alkaline liquid. The products formed are distinguished from the initial compounds by' the property of
being no longer precipitated by the neutralization of the
liquid; they are incoagulable and easily diffusible, thus
resembling crystalloids. Without wishing to speak very




SOLUBLE FERMENTS.              299
decidedly on this question, I am inclined to consider
peptones as bodies closely allied to the diffusable, soluble,
uncrystallizable,and incoagulable substances which I have
obtained by the incomplete action of hydrated baryta on
albumin. As to the characters of the third order, by
which an attempt has been made to distinguish the different peptones, we pass them over in silence; they
present no definite characters to the mind. We can
form a clear idea of the nature of these bodies only by
bringing about by hydratation a complete resolution of
these bodies into definite chemical principles, such as
the elements of urea, tyrosine, leucine, and its homologues, and thus determining their constitution. An investigation of this kind, connected with that which I have
undertaken with respect to albuminoid substances, could
not fail to produce important results in a physiological
point of view.
The analogy of functions led M. Bernard to seek for
albuminosic digestion in plants. If, with him, we call by
this name every transformation of albuminoid matter
into soluble diffusible principles, it is certain that it must
exist there.
Thus the phenomena presented by yeast preserved in
a damp state without nourishment, and to which we have
referred when treating of alcoholic fermentation, ought
and must be considered as a true digestion of proteids.
It is the same with the continuous chemical transformations which take place in the protoplasm of vegetable
and animal cells. Nothing proves that the first products
of the change of albuminoids in the organism are entirely
excrementitious bodies, and that these products, as long
as they remain above a certain limit of splitting up,




300             ON FERMENTATION.
which they do not attain, as, for example, the crystalloids, such as leucine, tyrosine, &c., can no longer contribute to organic synthesis, and to the formation of new
cells or tissues. It is the water with which digested
yeast has been washed that contains the most suitable
nourishment for the development of this organic ferment;
but, excepting the albumin, which is inactive as nourishment for yeast, the nitrogenous principles of the water
of yeast are products of an order inferior to that of
proteinic substances; they are the results of their
splitting up. We do not here speak of leucine and
tyrosine, the presence of which has been recognized in
the water with which yeast has been washed, and which
have no nutritive power, but only of the nitrogenous
bodies contained in the uncrystallizable sirup.
BrUicke found pepsin in the blood, the muscles, and
the urine. This agent, therefore, is not confined to the
stomach only; it is diffused over the various parts of
the organism, wherever its presence may be necessary,
especially where there are albuminoids to be liquefied
and digested in order to render them fit for nutrition.
Bretonneau had already announced that meat introduced into a sub-cutaneous wound could be digested
there as in the stomach.
We have given these details of the indirect fermentations which are connected with the great biological
phenomena of digestion, the first act of nutrition in
living organisms, because of the great importance that
attaches to this subject.
As to the other phenomena of the same order, we
shall treat them more slightly, since all that is essential
may be described in general terms.




SOLUBLE FERMENTS.               30 1
Fermenlatlioz of Glzucosides.-The ferment which act 
most generally and energetically on these bodies is found
in sweet and bitter almonds; this is synastase or
emulsin. It acts best between 3o0 and 400 C. (86~
and Io4~ F.); but still it can bear an elevation of
temperature to 800 C. (1760 F.), without losing its
specific power.
Acids and alkalies, in small quantities, do not interfere with the action of emulsin. It is only after having
undergone a somewhat advanced state of putrefaction
that emulsin loses its qualities. Its intervention may
be supplied, up to a certain point, by other principles
of animal origin. Substances which are poisonous to
plants and to yeast have no influence on the reactions
produced by synastase.
All these facts, in addition to its mode of preparation
(see the process of Wittich, before described), leave no
doubt as to the nature of emulsin; it is a soluble ferment of the diastase family.
It will be sufficient to give a short summary of the
reactions over which it presides; for further details we
must refer to works on chemistry.
ist. Splitting up of amygdalin into glucose, hydrocianic acid, and benzoyl hydride. This reaction explains that which takes place when bitter almonds are
pounded with water.  The bitter almond contains at
the same time amygdalin and emulsin.  These two
bodies are in distinct cells, and can only react on each
other when a mechanical action and solution place
them in close contact.
This action may take place within the organism; as,
for instance, when we inject dissolved amygdalin and




302                 ON FERMENTATION.
emulsin into the veins of an animal, the subject
dies  with  the  symptoms  of   poisoning  by  prussic
acid.
2nd. Resolution of salicin into glucose and saligenin.
3rd.,,      of  chlorosalicin  into   glucose   and
chlorosaligenin.
4th.,,      of helicin  into  glucose and  salicylic
hydride.
5th.,,      of arbutin  into glucose and  hydroquinone.
6th.,,       of phlorizin  into  glucose and  phloretin.
7th.,,      of esculin into glucose and esculetin.
8th.,,      of daphnin  into  glucose  and  daphnetin.*
Vanillin, the odorous principle of vanilla, has quite
recently been artificially produced, by decomposing by
* These decompositions take place according to the following equations:C~0 H27 N 01   +  2 HIO -  2 C6 H'2- OG +  C7 -H6 O +  C N H
Amygdalin.       Water.      Glucose.       Benzoyl    Prussic
hydride.    acid.
C1' H;8 07 +  H2 0 =  C6 H]2 06 +  C7 H8 02
Salicin.               Glucose.     Saligenin.
C'3 H1 C1. 07 +  H2 0 -  C6 H12 06 +  C7 H7 C1. 02
Chlorosalicin.              Glucose.    Chlorosaligenin.
C13H~6O7 H6  0   H2   -- C6 H1206  +    C7 H6 02
Helicin.                 Glucose.    Salicylic hydride.
C12 H16 07 +  H2 0 -  C6 H12 06 +  C6 H6 02
Arbutin.                Glucose.   Hydroquinone,
C21 H24 0' +- H2 0 =  C6 H12 0O  +  C15 H14 0O
Phlorizin.               Glucose.       Phloretin.
C21 H24 013 +  3 H20 O      2 C6 H12 G6 +  C9 H6 04
Esculin.                    Glucose.        Esculetin.
C3' H34 019 +  2 HI2 0 -- 2 C6 H12 06 +  C19 H14 09
Daplhnin.                  Glucose.        Daphnetin.




SOLUBLE FERMENTS.               303
means of synoptase a glucoside contained in coniferous
plants (coniferin), and by oxidizing the principle formed
by this decomposition.
Emulsin, like most of the soluble ferments, may be
replaced, with respect to glucosides, by agents purely
chemical. Thus we arrive at the same results, in most
cases, by boiling with dilute acids.
A very interesting chemical reaction, of the same
order as that which we have observed in bitter almonds,
is found in the flour of mustard seed, mixed with water.
This mixture, the common mustard plaster, has, as we
know, a strong odour and a burning taste. The properties of this sinapism are due especially to the presence
of essence of mustard, or allyl sulphocyanide. Put this
essence does not exist fully formed in the seed any more
than the essence of bitter almonds in the kernel; it
originates at the expense of a special compound
contained in black mustard, under the influence of
a soluble ferment, myrosin, contained in white or
black mustard, and generally in the seeds of the
crtuciferae.
The initial product has been isolated in a crystalline
form; this is the potassic salt of a complex acid, myronic
acid. As to the ferment, we have nothing of importance
to say of it; it may be eliminated from the seeds of the
cruciferae, or of white mustard, which do not contain
potassium myronate, by the general methods already
described; its physical characters, as well as its composition, are those of all the soluble ferments.
Potassium myronate (C'0 H8 N. K. S2 0'~), whose
presence characterizes black mustard, is resolved, under
the influence of myrosin, into glucose, allyl sulphocyanide,




304              ON FERMENTATION.
and potassium, bisulphate, as shown in the following
equation:C~0 H18 N K S2 010   C6 H51 06 + C N C3 H5 I S + S 04 K H
Potassium myronate.   Glucose.    Allyl sul-  Potassium
phocyanide.    bisulphate.
The potassium myronate is extracted in the following
manner by the process  of M. Bussy, modified  by
Messrs. Witt and K6rner.  A  kilogramme of black
mustard seed pulverized is boiled with IY  litres of
alcohol at 82 per cent., until a quarter of a litre of
alcohol is distilled over. It is pressed while hot, and
the same operation is repeated with the remainder.
This is again pressed, dried at Ioo~ C. (2I20 F.), pulverized, and digested for twelve hours with three parts
of cold water; it is pressed again, and the remainder
diffused in two parts of cold water. The watery solutions are evaporated, after the addition of a little barium
carbonate, till they are of a sirupy consistence. The
remainder is treated with boiling alcohol at 85 per cent.
(I or I 2 litres); it is then filtered, distilled, and left in
flat open plates to crystallize.
We will now mention the other analogous fermentations, without dwelling on them.
I. The fresh root of the madder contains alizarin and
other colouring matters, insoluble of themselves, under
the form of soluble glucosides (Rubian, ruberythric acid).
Together with these principles, the same root contains
a soluble ferment, erythrozyme, which is not slow in
splitting up these glucosides, when the powdered madder-root is mixed with water.   Under its influence,
the colouring matters, at first dissolved, separate whilst




SOLUBLE FERMENTS,                305
sugar is formed. M. E. Kopp has succeeded in stopping
the action of the ferment, by the addition of a certain
quantity of sulphurous acid; he founded on this fact a
very interesting process for the extraction of the colouring matters of madder.
2. The infusions of the seeds of buckthorn, when they
are kept for some time, also undergo a fermentation
which decomposes the soluble colouring glucosides contained in this infusion or decoction; glucose and an
insoluble pigment are formed. Boiling with acids produces the same transformation.
3. The biliary acids, the taurocholic and glycocholic,
are susceptible, as we know by the works of Strecker, of
being resolved by hydratation into taurine and cholalic
acid, or into glycocoll and cholalic acid. All that need
be done, in order to attain this result, is to boil it for a
sufficient time with baryta. The same effect of decomposition is observed when bile is left to itself, to undergo
spontaneous change. Taurocholic acid is transformed
more easily than its accompanying acid.  This easy
transformation, due to the intervention of ferments, has
for a long time obscured the chemical history of bile.
4. The decomposition of hippuric acid, in the urine of
the herbivora, into benzoic acid and glycocoll is a reaction of the same order.
5. This is the case also with two vegetable glucosides,
phillyriin, contained in the bark of the PAzillyrea laizfolia,
and popzilin, from  the bark of the aspen. These substances are attacked neither by beer-yeast nor by
emulsin; but when they are placed under the conditions of lactic fermentation, they are resolved, the one
into glucose and phillygenin —
14




306               ON FERMENTATION.
C'7 Hf:4 01 +- H2 0 -  C6 H1'2 06 + C21 H24 06
Phillyrin.            Glucose.    Phillygenin.
the other into glucose, saligenin, and benzoic acidC13 H17 (C? Hh O) 07 + H2 0  =  C'3 Hl8 07 + C7 H6 02
Populin or benzoyl salicin.        Salicin.    Benzoic acid.
The salicin formed at first is resolved in its turn into
glucose and saligenin.
6. Infusions of gall-nuts, left to themselves, ferment;
the tannin disappears, and is replaced by gallic and
ellagic acid, and glucose.
Strecker, considering the tannin as a glucoside, represented its decomposition by the equationC27 H22 0O7 + 4 120 =  C6 H12 O6 + 3 C7 t16 05
Tannin.                Glucose.     Gallic acid.
7. According to M. Fremy, pectose is accompanied, in
the vegetable tissues in which it is found, by a ferment,
pectose, sometimes soluble and at others insoluble, which
possesses the  property  of transforming  pectose and
)ectill into pectic and metapectic acids successively.
P'cctic fermentation plays an important part in the
conversion of ripe fruits into an over-ripe, half-rotten,
or "sleepy" state.  It also assists in the formation
of vegetable jellies.  In  fact, the  transformation  of
the natural juices of fruits into jellies is a  result of
the metamorphosis of pectin, contained in these juices,
into the pectosic and pectic acids.  If, then, a natural
juice-that of the currant, for example-does not contain pectose, we must add to it another juice, or pulp,




SOLUBLE FERMENTS.                307
which contains it, in a soluble or unsoluble form,
remembering that boiling, by coagulating the ferment,
renders it inactive. Pectic fermentation is effected at
about 350 C. (95~ F.). (Fremy, Ann. Chim. Phys. (3), vol.
24, p. I.)




308            ON FERMENTATION.
CHAPTER III.
ON THE ORIGIN OF FERMENTS.
TILE question of the origin of ferments is intimately
connected with that of spontaneous generation. In fact,
from the time of Van Helmont and others, who, even
in the seventeenth century, gave directions for the production of mice, frogs, eels, &c., the partisans of this mode
of generation have, by the progress of the tendency to
examine into the causes of things, been driven from the
larger animals or plants visible to the naked eye, to the
smallest living productions, which we can observe only
by the aid of the microscope. But ferments are found
among these inferior microscopic organisms. Redi, a
member of the Academy of Cimento, showed that the
worms in putrefied flesh, which were at first thought to
be of spontaneous origin, are only the larvae from the
cggs of flies, and that all that was necessary to prevent
entirely the birth of these larvae, was to surround the
decomposing meat with fine gauze; he was the first
to ascertain that parasitic animals are sexual and able
to lay eggs.
The invention of the microscope, and the numerous
observations by which it was followed, towards the end
of the seventeenth, and the commencement of the eightcenth, century, gave fresh impulse to the doctrine of




ON THE ORIGIN OF FERMENTS.          309
spontaneous generation, which had lost all credit in questions concerning the origin of living beings of a higher
order.
The question now was how to explain the origin of
the various living' productions, revealed by the microscope in infusions of vegetable and animal substances,
among which no apparent symptom of sexual generation could then be found.
The subject was studied for the first time in a scientific manner by Needham, who published in i745, in
London, a work on this subject. This observer did for
infusoria what had already been done for the higher
organisms. He protected, or rather endeavoured to protect, vegetable or animal infusions from the action of
germs, seeds, or any other agents of mutiplication which
could come from without. At the same time he destroyed
by a physical agent, heat, the. germs which might be
supposed to exist beforehand in the liquid. Under these
conditions, either living beings will be produced in the
midst of the infusion, or none will be found there; in
the former case, it must be admitted that these organisms are developed in the medium which is suitable
to them, without the intervention of any germ; in the
second, that the doctrine of spontaneous generation is
false. In reality, the question can only be resolved in
this manner, and all experimenters who have entered
upon it from Needham's time to the present day ought
to have made use of it.
The serious and grave difficulty, on which, during this
period, all discussions raised between heterogenists and
panspermists have turned, is so to arrange the experiments as to remove every suspicion of the intervention




3 I0            ON FERMENTATION.
of germs brought from without, or pre-existing in the
liquid..
If the result is negative, if when all precautions that
seem to be necessary have been taken, and all causes
of error have been removed, there is no formation of
infusoria, it will be difficult to raise any serious objection
to the inevitable conclusion, provided that the methods
employed for the purpose of eliminating the pre-existing
germs are not of such a nature as to modify the medium,
and to render it unfit for the development and the nutrition of living organisms. If, on the contrary, we still
meet with the birth of living beings, the suspicion will
always revive that the experiment has been badly performed, and that a contrary result would have been
obtained by conducting it more carefully. The heterogenists, therefore, find themselves in a more disadvantageous situation than their opponents, and notwithstanding the success which they may obtain, they will
never convince them.
We think therefore that it is useless to give here a
detailed account of their minute researches; they must
be consulted in the original memoirs. A single experimenzt which proves, by a negative result, thzat organic
infusions, protected from germs fromZ without, do not give
birthz to infusoria, is worth more, scientzfically speaking,
than ten experiments tending to establish the contrary
opinion.
If, therefore, we pass over the details of the fundamental experiments of the heterogenists, and speak of
those the results of which are conformable to the ideas
of the panspermists, it will not be in a spirit of partiality. We are convinced that the latter are the only




ON THE ORIGIN OF FERMENTS.          311
ones free from all objections, the relative skill of the
operators being disregarded, and considered as nothing
in the estimate formed. We may, however, say, that
M. Pasteur's researches may serve as a model for all
those who may wish to conduct investigations of this
kind, whatever may be the preconceived opinion by
which they are guided. By their precision, and the care
taken to remove every source of error, they leave nothing
to be desired.
As the results obtained by M. Pasteur lead him to
deny spontaneous generation, his opponents must above
all prove that he is mistaken, by adopting the same
rigorous experimental conditions.
Needham's experiments, of which we have before
spoken, wlich led this observer to admit and sustain the
doctrine of spontaneous generation, consisted essentially
in placing organic substances which were capable of
decomposition, in vessels hermetically sealed, which
were subsequently submitted to a high temperature, in
order to destroy the pre-existing germs.
The work of the English writer attracted great notice
on account of the support of Buffon, whose ideas he
upheld.
Soon after began the great controversy between Needham and Spallanzani, a celebrated Italian physiologist.
The latter, in his treatises on animal and vegetable
physics, translated by Sennebier, in I777, refuted by
experiment the conclusions arrived at by Needham.
The controversy turned principally on this point;
Spallanzani was not satisfied with heating the hermetically sealed vessels containing the infusions, for several
minutes, merely the lime zvhiich is required io cook a leu's




312             ON FERMENTATION.
egg, and io destroy the germs, as Needham expresses it;
but he kept them for the space of an hour in boiling
water. He then observed no production of infusoria.
But, objects the English observer, from the manner in
which he treated and put to the torture his nineteen
vegetable infusions, it is evident that he not only much
weakened, or perhaps totally destroyed, the vegetative
force of the substances infused, but also entirely corrupted, by the exhalations and the odour of the fire, the
small portion of air which remained in the empty part
of his vessels. It is not therefore surprising that his
infusions, thus treated, gave no signs of life. Such must
necessarily have been the case.
This idea, that the action of the temperature of boiling water destroys the vegetative force of infusions, is
maintained even at the present day, and has served as
an argument to the heterogenists; as they were unable
to attack the material correctness of Pasteur's experiments, they did not accept the conclusions which he
sought to derive from them.
We find also in the passage just cited, the necessity
for the experiments made by Schwann and Helmholtz
on calcined air, and for those of Schrceder and F. Dusch,
on strained air.
The objection of a possible change in the air contained
in the phial, under the influence of prolonged boiling,
in presence of organic substances, was a serious one at
the time that it was brought forward; it becomes more
so, when we know that the air confined over preserved
meats, prepared by Appert's process, contains no oxygen.
It was, therefore, absolutely necessary to place the infusions in contact with air in a normal condition, after




ON TIlE ORIGIN OF FERMENTS.          313
that boiling had deprived them  of their pre-existing
germs, avoiding at the same time any new germs brought
by the air.
For this purpose, Dr. Schwann heated flasks containing the infusions, until the destruction of the germs
was insured; but his flask was not closed; it communicated freely with the surrounding air by means of a
glass tube bent in the form of an U, and heated, in one
part of its length, by means of a bath of fusible alloy.
Under these conditions, the air may be renewed in the
flasks, but the fresh atmospheric air admitted has undergone, like the infusion, the action of heat, which destroys
the germs.
Schwann's experiment was very decisive, as to broth
made from meat; and the negative result (no development of infusoria) was quite satisfactory.  But it was
not the same with analogous trials on alcoholic fermentation, which gave contradictory results.
Ure and Helmholtz repeated and multiplied these
experiments with the same success.
To obviate the objection of a possible change by heat,
in a mysterious and undefined principle, different from
germs, but whose presence in the air was necessary to
the production of infusoria, Schultz (Ann. des Sciences
Naturelles, vol. 8, (2), 1837) caused the renewed air to
pass through energetic chemical re-agents, such as concentrated sulphuric acid. He half filled a glass vessel
with distilled water containing various animal and vegetable substances; then stopped the vessel with a cork
through which passed two bent tubes, and exposed the
apparatus thus arranged to the temperature of boiling
water. Then, while the vapour was still escaping through




3 I4           ON FERMENTATION.
the tubes of which we have just spoken, he adapted
to each of them a Liebig's bulb apparatus, one containing concentrated sulphuric acid, and the other concentrated caustic potash.  The high temperature must
necessarily have destroyed every living thing, all the
germs that might happen to be in the inside of the
vessel, or of its appendages, and the communication
from without was intercepted by the sulphuric acid on one
side and the potassa on the other. Nevertheless, it was
easy to renew by aspiration at the end of the apparatus
which contained the potassa, the air thus enclosed, and
the fresh quantities of this fluid which were introduced
could not carry with them any living germ, for they were
forced to pass through a bath of concentrated sulphuric
acid. M. Schultze placed the apparatus thus arranged
at a well lighted window, side by side with an open
vessel, which contained an infusion of the same organic
substances; then he was careful to renew the air in his
apparatus several times a day for more than two months,
and to examine with the microscope what took place in
the infusion. The open vessel was soon found filled
with vibrios and monads, to which were soon added
polygastric infusoria of a larger size, and even rotifers;
but by the most attentive observation he could not discover the least trace of infusoria, confervae, or mildews,
in the infusion contained in the apparatus.
The latest researches of Schrceder and V. Dusch
(I854-I859) tended to raise another objection, the possible change in a special principle in the air, by a re-agent
as energetic as sulphuric acid. Guided by the experiments of Loewel, who ascertained that common air,
when it had been previously filtered through cotton, was




ON TI-IE ORIGIN OF FERMENTS.        315
unfit to cause the crystallization of supersaturated solutions of sodium sulphate, they placed one of the tubes
of Schutze's apparatus in communication with a tube
3 centimetres (I'IS in.) in diameter, and from 50 to 60o
centimetres (19i68 to 23'62 in.) in length, filled with
cotton-wool. The other tube was connected with an
aspirator.
When the liquid, the interior of the flask, and the
tubes, had been been deprived of air by boiling, the
apparatus was removed to its place, and the aspiration
continued night and day.
The two observers thus proved that meat to which
water had been added, the wort of beer, urine, starch,
paste, and the various materials of milk taken separately, remained intact in the filtered air.
On the contrary, milk, meat without water, and the
yolk of egg, grew putrid as rapidly as in common air.
The result of these experiments is, that there are
spontaneous decompositions of organic substances which
require nothing but the presence of oxygen gas to cause
them to commence; while others need, besides oxygen,
the presence in the atmospheric air of those uzlzkwzovwz;/ibz.gs, which are destroyed by heat or sulphuric acid,
or are retained by the cotton.
The two observers do not then decide on the nature of
these things, and do not assert categorically that they
are germs, and, in reality, nothing allows us to draw
these conclusions.
M. Pasteur's experiments (Memoire on the Organic
Corpuscles which exist in the Atmosphere, Ann.
Chim. Phys. (3), vol. 64, p. 27) have advanced the question another step, by proving that they are really germs




3 I6            ON FERMENTATION.
of ferments and infusoria, which are destroyed by heat, or
arrested by the sulphuric acid or cotton in the experiments alluded to above.
M. Pasteur made a hole in a window-shutter, several
metres above the ground, and through this he passed a
glass tube half a centemetre ('I96 in.) in diameter, and
containing a plug of soluble cotton I centimetre
('39 in.) in length, kept in its place by a spiral platinum
wire. One of the ends of this tube passed into the
street; the other communicated with a continuous
aspirator.  When the air had passed for a sufficient
time, the plug of cotton, more or less soiled by the dust
which it had intercepted, was placed in a small tube with
the mixture of alcohol and ether, which dissolves guncotton. It was left for the space of a day. All the
dust was deposited at the bottom of the tube, where it
is easy to wash it by decantation, without any loss, if
care is taken to separate each washing by an interval of
repose of from twelve to twenty hours. The deposit,
and the liquid which covers it, are put in a watch-glass
together; after the evaporation of the alcohol, the
remainder is placed in water, and examined with the
microscope. M. Pasteur also made use of ordinary
sulphuric acid in order to moisten the dust. This agent
had the effect of separating the grains of starch and
calcium carbonate, which are always found in greater
or less quantities in deposits collected on the plug of
cotton.
Figs. 24 and 25 represent organic corpuscules, associated wih amorphous particles, as seen through the
microscope, under a power of 350 diametres; the liquid
containing them was common sulphuric acid.




ON TIHE ORIGIN OF FERMENTS.              3I7
Fig. 24 applies to dust collected from the 25th to the
26th of June, IS6o; fig. 25 to dust from the very intense
fog of January, I86I.
It was not enough to discover with the microscope
organic particles mixed with amorphous substances, but
FIG. 24.                   FIG. 25.
FIt's. 24 and 25.-Organic corpuscules of dust, mixed with amorphous particles.
it was necessary to prove that these particles really consisted of fertile germs, capable of producing the infusoria
which are developed in such abundance in organic liquids
exposed to the air.
For this purpose, M. Pasteur arranged the experiment
in the following manner:Into a flask capable of containing from 250 to 300
cubic centimetres (I5 to I8 cub. in.), he introduced Ioo
or I5o cubic centimentres (6 to 9 cub. in.) of albuminous
saccharine water, prepared in the following proportions:
Water Ioo
Sugar Io
Albuminoid and mineral matter from beer-yeast'2 to'7.
The neck of the drawn-out neck flask communicated
with a platinum  tube, as shown in Fig. 26.  In this
first stage of the experiment the T-shaped tube with




3 I8            ON FERMENTATION.
three stop-cocks is removed, and its place supplied by a
simple india-rubber connecting piece. The platinum
tube is raised to a red heat by means of a small gas
furnace. The liquid is boiled for two or three minutes,
and is then allowed to grow completely cold.  It is
filled with common air, at the ordinary pressure of the
atmosphere, but which has been wholly exposed to a
red heat; then the neck of the flask is hermetically sealed.
This, being thus prepared, and detached, is placed
in a stove at a constant temperature of about 30~ C.
(86~ F.); it may be kept there for any length of time
without the least change in the liquid which it
contains.  It preserves its limpidity, its smell, and
and its weak acid reaction; even a very slight absorption of oxygen is mainly to be observed. M. Pasteur
affirms that he never had a single experiment which was
arranged as described above, which yielded a doubtful
result; while water of yeast mixed with sugar, and
boiled for two or three minutes, and then exposed to
the air, was already in evident process of decomposition
in a day or two, and was found to be filled with bacteria
and vibrios, or covered with mucors. These experiments
are directly opposed to those of Messrs. Pousset, Mantegazzo, Joly, and Musset.
It is therefore clearly proved that sweetened yeast
water, a liquid very liable to be decomposed by the
contact of common air, may be preserved for years
unaltered when it has been exposed to the action of
calcined air, after having been allowed to boil for a few
minutes (two or three).*
M. Pasteur has pointed out a cause of want of success, which has led
many experimenters into error; by showing that the mercury of a mercurial




d~~~~~~~~~~~~~~~~~
X~~~`  -   B'  -v D avrl;.
MG2.IPsu'aaasoteirutnfE=,ariofscoaigracnuo
F
~c\\l\\\~\\~.\. 
Y ~ ~ ~        ~                     ~         ~         ~         URGUIY.~~~~~~~~
z
H
FIn. a6.-M~. Pasteur's apparatus for the introduction of calcined air into flasks containing orgnanic infusions   \




320              ON FERMENTATION.
This being determined, M. Pasteur adapted, by means
of an india-rubber tube, the closed point of his flask
filled with sweetened yeast water, which had been kept
for two or three months in a heated stove, without any
development of organisms, to an apparatus arranged like
that in figure 26.
The pointed end of the flask passed into a strong glass
tube  o or I2 millimetres ('39 to'46 in.) in its inner diameter, within which he had placed a piece of tube of small
diameter, open at both ends, free to slip in the larger
tube, and enclosing a portion of one of the small plugs
of cotton loaded with dust. The larger glass tube is
bound to a brass tube in form of a T, furnished with
stop-cocks, one of which communicates with the airpump, another with the heated platinum tube, and the
third with the flask, by means of the large tube which
contains the smaller one with the cotton. These various
parts are joined together by means of india-rubber.
The experiment is commenced by exhausting the air,
after having closed the stop-cock connected with the
red-hot metallic tube. This being afterwards opened,
allows calcined air to enter the tubes slowly; this operation (exhaustion and readmission of calcined air) is
repeated several times. The point of the flask is then
broken off within the india-rubber, and the small tube
containing the dust is allowed to slip into the flask, the
neck of which is again sealed with the lamp.  As an
additional proof, and to obviate all objections, the same
arrangements were made with similar flasks, prepared
trough is a complete receptacle for living organisms, and consequently that all
experiments made with such a trough must necessarily induce a development
of infusoria.




ON THE ORIGIN OF FERMENTS.           32I
like the preceding, but with this difference, that instead
of cotton charged with atmospheric dust, there was
substituted a small piece of tube containing calcined
asbestos; (as an additional precaution, it had been
ascertained that calcined asbestos, loaded with atmospheric dust, by the same means as the cotton, gave
identical results).
The following are the observations obtained constantly
by M. Pasteur;In all the flasks, into which dust collected from tile
air were introduced, I. Organic productions began to
make their appearance in the liquid after 24, 36, or
48 hours at the most. This was precisely the time
necessary for the same phenomena to appear in
sweetened yeast-water exposed to contact with the
atmosphere.
2. The products observed are of the same kind as
those which are seen to make their appearance in the
liquid when left freely exposed to the air, such as
mucors, common mucidines, torulacei, bacteria, and
vibrios of the smallest species, the largest of which, the
mo0zas lvcis, is only'004 millimetre (oo000157 ill.) in
diameter.
It was a remarkable circumstance that MS. Pasteur
never saw any alcoholic fermentation appear, although
the composition of the liquid employed was very appropriate to this kind of change.
When the water of yeast is replaced by urine, the
experiment being conducted exactly in the same manner,
we always notice the absence of any change as long as
atmospheric dust has not been introduced, whilst with
the addition of this, numerous organisms are developed,




322             ON FERMENTATION.
in every respect similar to those which appear and are
developed in urine kept in the open air.
If, on the contrary, the experiment be repeated with
common milk, we may be sure that it will in every case
curdle, and become putrid. We shall observe the birth
of numerous vibrios of the same species, and bacteria,
and the oxygen of the flask will disappear.
M. Pasteur thinks that this result, so different from
those observed in other liquids, arises only from the fact
that milk contains germs of vibrios which resist the
boiling heat of water. To prove this, he boiled milk,
not at Ioo0 C. (2I2~ F.), or at the usual pressure of the
atmosphere, but at I 10 C. (230~ F.), under a greater
pressure, and he found that the flasks thus prepared,
and hermetically sealed, could be kept for an indefinite
time in the stove, without giving rise to the smallest
production of mould or infusoria. The milk preserves
its taste, its smell, and all its properties; and the atmosphere of the flask is only slightly modified in its composition.  (After forty days there were still found I8'37
volumes of oxygen per Ioo parts of air.)
This difference between milk and urine, or sweetened
yeast-water, must be attributed to the alkaline condition
of the former medium, whereas the two others are
acid.
In fact, if we previously neutralize the acid of the
sweetened yeast-water, by means of calcium carbonate,
we obtain organisms under the same conditions of the
experiment as those under which they were not before
developed.
These facts led M. Pasteur to make researches on the
comparative action of temperature on the fecundity of




n       ~             li i~ il                             1at]Bt
0
me~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~t
FIG. 27.-M. Pasteur's apparatus for studying the resistance of germs and spores to temperatures more or less elevated
YI   lil  13    1~ ~ ~ ~'`   lii;, iiiiillllllll~~~~~~~8~~'111           ~( 3
to
tJ




324            ON FERMENTATION.
the spores of the mucidines, and of the germs which
exist suspended in the atmosphere.
The following is, in few words, the method followed
by him. He passed a small portion of asbestos over
the small heads of the moulds which he wished to study;
he then placed- this asbestos, covered with spores, in a
small glass tube, which he introduced into an U tube
(Fig. 27) of larger diameter, in which the smaller tube
could move freely; one of the extremities of the U tube
is joined by india-rubber to a metal tube in form of a
T, with stop-cocks. One of these cocks communicated
with the air-pump, another with a red-hot platinum tube.
The other extremity has an india-rubber tube which is
connected with the flask into which the spores are to be
introduced; this flask is hermetically sealed, and has
been filled with calcined air, and suitable nutritious
liquid previously raised to the boiling point. Finally,
the U tube dips into a bath of oil, of common water, or
salt water, according to the temperature which we wish
to attain. Between the U tube and that of platinum,
there is a drying tube with sulphuric pumice-stone.
When all the apparatus which precedes the platinum
tube has been filled with calcined air, and the spores
have been maintained at the desired temperature for a
sufficient time, which may be varied at pleasure, the point
of the flask is broken with a blow of a hammer, without
unfastening the india-rubber connecting pieces which
attach the flask to the U tube; then inclining to a proper
angle, this latter tube, when removed from its bath, the
asbestos with its spores is slipped into the flask. The
flask is then hermetically sealed, and is carried to the
stove at 20o or 30~ C. (68~ to 86~ F.). The experiment




ON THE ORIGIN OF FERMENTS.           325
with the dust from the air is also made in the same
manner with asbestos.
Without any humidity, the fecundity of the spores of
Peizecillium glaucum is preserved up to I20~ C. (2480 F.)
and even a little above-I25~ C. (257~ F.). It is the same
with the spores of the other common mucidines. At
I30~ C. (266)~ F.), the power of developing or multiplying is destroyed in all of them. The limits are the
same for the dust from the air.
In all these careful experiments, the most scrupulous
precautions were taken to prevent the access of the
slightest portion of common air. But, say the partisans
of heterogenesis, if the smallest portion of common
air develops organisms in any infusion whatever, it must
necessarily be the case that, if these organisms are not
spontaneously generated, there must be germs of a
multitude of various productions in this portion of
common air, however small it may be; and if things
were so, the ordinary air would be loaded with organic
matter which would form a thick mist in it.
M. Pasteur has shown that there is a great deal of
exaggeration in the generally received opinion that even
the smallest quantity of air is sufficient to develop
multitudes of organisms; that, on the contrary, there
is not in the atmosphere a continuous cause of these
so-called spontaneous generations; that it is always
possible to procure, in any determined place, a sufficient
but still limited quantity of common air, having undergone no kind  of modification, whether physical or
chemical, and nevertheless quite unsuited to set up any
decomposing action in a liquid eminently putrescible.
The method of experimenting is very simple. Into a




326             ON FERMENTATION.
flask of 250 or 300 cubic centimetres (15 to I8 cub. in.),
I5o cubic centimetres (9 cub. in.) of a liquid that has a
tendency to decomposition are introduced; the neck of
the flask is drawn out with the lamp, leaving the point
open; then the liquid is boiled till the vapour escaping
from the extremity has expelled all the air; at this
moment the point of the flask is closed by the lamp, by
means of a blow-pipe, and it is allowed to grow cool. The
flask then contains no air; if we break off the point in
any particular place, the air re-enters suddenly, carrying
into it the germs held in suspension; it is again closed
with the lamp, and kept in a stove at a temperature of
200 or 300 C. (68~ to 86~ F.). In the generality of cases,
organisms are developed; these organisms are even
more varied than if the liquid were freely exposed to
the air, which M. Pasteur explains by saying that, in this
case, the germs in small number, in a limited volume of
air, are not hindered in their development by germs in
greater number or more precocious in their fecundity,
which are able to occupy the space, and leave no room for
them. But it is especially important to notice in the results
obtained by this method, what frequently happens many
times in each series of trials, that the liquid continues
absolutely intact, however long it may have remained
in the stove, as if it had been filled with calcined air.
This phenomenon is the more striking, and shows itself in more marked proportions, when the air received
into the flasks is taken from a greater height. Thus,
among twenty flasks opened in the country, eight contained organic productions; out of twenty opened on
the Jura, only five contained any; and out of twenty
flasks opened at Montanvert, in a rather high wind,




ON THE ORIGIN OF FERMENTS.            327
blowing from the deepest gorges of the "Glacier des
Bois," one only was affected by any change.
We may also draw other conclusions from this series
of observations. Since the putrescible liquid, which
had been previously boiled, and which was contained
FIG. 28.-M. Pasteur's flask to deprive the air of its germs.
in the flasks, was filled with organic productions in a
great number of instances, after the introduction of a
limited quantity of air, the genetic power of the infusions had not been destroyed by the material conditions of the experiments. Besides, this objection, which
has been raised ever since the earliest controversies
between the heterogenists and the panspermists, has
been definitely answered by an experiment made by
M. Pasteur; he received in a flask, exhausted and
deprived of living germs by the momentary application of a sufficiently high temperature, some blood at
the instant that it left the organism, and without allowing this liquid, which is so peculiarly putrescible, to come
in contact with air.   By permitting air- deprived of
germs, either by calcination or simple filtration, to enter
the flask, and then hermetically sealing it, he found that




328            ON FERIMENTATION.
the blood was preserved for an indefinite period intact,
although it had not been exposed to heat.
M. Pasteur has also shown that air may be deprived
of germs by its passage through a capillary tube bent
upon itself. It is, therefore, sufficient, in most cases, to
draw out the neck of the flask so as to form a very long
narrow tube, which is bent in several directions, as, for
example, in Fig. 28. When the air originally contained
in it had been expelled, and the pre-existing germs
killed by prolonged boiling, the flask is allowed to cool
slowly.
In closing our account of M. Pasteur's interesting
memoir, in which heterogenesis was driven to its last
intrenchments, we must add that this learned chemist
endeavoured to deprive his adversaries of one of their
principal arguments. Experiments on spontaneous generation have always been conducted with vegetable or
animal infusions; it was supposed by Needham, Buffon,
and Pouchet that organisms were only thus produced at
the moment of expiring nature, when the elements of
the beings on which they are developed are entering
into new chemical combinations, and are passing fully
through the phenomena of fermentation or putrefaction.
In other words, albuminoid matters preserve in some
degree a certain reserve of vitality, which would allow
them to become organic by contact with oxygen, when
the conditions of temperature and humidity are favourable. Starting with the idea that albuminoid substances
are only aliments for the germs of infusoria, mucidines,
or ferments, M. Pasteur has proved directly that organic
substances may be replaced by those which are purely
mineral or artificial, or, at least, by substances on which




ON THE ORIGIN OF FERMENTS.           329
this imaginary vegetative force cannot be supposed to
have any influence.
We have spoken elsewhere, at some length, of the
experiments made by M. Pasteur and M. Raulin on the
nutrition and development of ferments and of mucedines in artificial media composed of pure sugar candy,
ammonium tartrate, and phosphates.
The reader will doubtless remember an observation
of M. Pasteur's, which we mentioned in passing, but
without dwelling upon it. When he introduced into
sweetened yeast-water that had been previously boiled
and preserved in calcined air atmospheric dust collected
at different times and in various places, he never met
with alcoholic fermentation. Yet the liquid employed is
one of those which are most suitable for the development
of alcoholic ferments. The fact may be explained by
admitting that the air does not contain spores or germs
of sacc/haromyces or apiculated ferment; but then how
can we explain the prompt and constant appearance
of alcoholic fermentation in the juice of grapes or of
fruits in general? The cause of this apparent contradiction is very simple. It is not the air which brings
the germs of the alcoholic ferments which propagate and
multiply so rapidly in the must of grapes, or, if it brings
any, they are in so minute a quantity that they would
not be sufficient to set up fermentation in so short a
time. These germs are found on the very surface of the
fruit, on the grapes which contain the saccharine liquid,
the decomposition of which they excite as soon as they
are placed in contact with it when the fruit is pressed.
In order to prove this, M. Pasteur prepared a series of
forty flasks, with sinuous necks like those which we have
15




330             ON FERMENTATION.
described above (Fig. 28), with this difference only,
that the neck of the flask, which is drawn out to a fine
"swan's neck" is not the only one. Each flask has another
straight neck closed by an india-rubber tube furnished
with a glass stopper. Into these forty flasks he introduced limpid and filtered grape juice, which, after being
boiled, like all liquids which are slightly acid, remains
intact, although the extremity of the neck is open. On
the other hand, he washed in a few cubic centimetres
of water part of a bunch of grapes. Under the microscope, we may perceive in this water the existence of a
multitude of organic corpuscules, resembling so closely
as to be indistinguishable from them, either spores of
minute fungi, alcoholic ferment, or the Mycoderma vilzi.
This having been done, M. Pasteur sowed nothing in
ten of the forty flasks; in ten others, he placed, by
means of the second tube, some drops of the liquid in
which the grapes had been washed. In a third series
of ten other flasks, he placed some drops of the same
liquid, previously raised to the boiling point, and then
cooled. Into the ten remaining flasks, he introduced
one drop of the juice of a grape taken from those that
were not crushed.
The first series gave no product, the juice having
remained unaltered. In the second series, there appeared
some flakes of mycelium, somc alcoholic ferment, and,
afterwards, Mycodaerma vizi; at the end of forty-eight
hours the ten flasks were in full fermentation, if they
had been experimented upon at the temperature of the
air. The third series had not a single flask affected;
the liquid remained limpid. In the fourth, a single flask
was changed. The conclusion which may be drawn




ON THE ORIGIN OF FERMENTS.            331
from these facts is simple.*  M. Bechamp had already
proved, by former experiments, that grapes bear on their
surface all that is necessary to cause saccharine water to
ferment, even when protected from the air.
With the question of the origin of ferments and of
spontaneous generation, is connected another, which may
be discussed independently, whatever may be the origin
of ferments, whether spontaneous or no; namely, whether
a fermenting organism can transform itself into a different ferment, endowed with distinct active properties,
when the conditions of its development are modified.
It is evident that this question may be seriously discussed; it is connected with the general development
theory which has been applied to higher organisms; it
is, therefore, still more applicable to the simplest organisms of the living creation. The facts observed are not
entirely opposed to the idea of a transformation of ferments into each other; we have even had occasion to
mention some which seem favourable to it.  In the
meanwhile, these facts are still too few in number to give
rise to important inferences; some of them are even
contested. We shall therefore merely call attention to
this branch of the study of inferior organisms; it has
already been the object of valuable researches, which
for several reasons deserve to be continued.
* Pasteur, Comp.-Rend., vol. 75, P. 78i.








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but also original and suggestive in the highest degree. Nowhere since the publication
of Sir Henry Maine's'Ancient Law,' have we seen so many fruitful thoughts suggested in the course of a couple of hundred pages....   To do justice to Mr. Bagehot's fertile book, would require a long article. With the best of intentions, we are
conscious of having given but a sorry account of it in these brief paragraphs. But we
hope we have said enough to commend it to the attention of the thoughtful reader."Prof. JOHN FISKE, in the A tlantic Monthly.
"Mr. Bagehot's style is clear and vigorous. We refrain from giving a fuller account of these suggestive essays, only because we are sure that our readers will find it
worth their while to peruse the book for themselves; and we sincerely hope that the
forthcoming parts of the'International Scientific Series' will be as interesting." —
A t/hezazn.
"Mr. Bagehot discusses an immense variety of topics connected with the progress
of societies and nations, and the development of their distinctive peculiarities; and his
book shows an abundance of ingenious and original thought."-ALFRED RUSSELz
WALLACE, in Nataere.
D. APPLETON  & CO., Publishers, 549 & 55I Broadway, N. Y.




Oinioans of thze Press on the "Inlerlnalional  Scienltiqc Series."
III.
Foods.
By Dr. EDWARD SMITH.
I vol., 12mo.  Cloth.  Illustrated........         Price, $1.75.
In making up THE INTERNATIONAL SCIENTIFIC SERIES, Dr. Edward Smith was selected as the ablest man in England to treat the important subject of Foods. His services
were secured for the undertaking, and the little treatise he has produced shows that the
choice of a writer on this subject was most fortunate, as the book is unquestionably the
clearest and best-digested compend of the Science of Foods that has appeared in our
language.
" The book contains a series of diagrams, displaying the effects of sleep and meals
on pulsation and respiration, and of various kinds of food on respiration, which, as the
results of Dr. Smith's own experiments, possess a very high value. We have not far
to go in this work for occasions of favorable criticism; they occur throughout, but are
perhaps most apparent in those parts of the subject with which Dr. Smith's name is especially linked."-London Examiner.
" The union of scientific and popular treatment in the composition of this work will
afford an attraction to many readers who would have been indifferent to purely theoretical details.... Still his work abounds in information, much of which is of great value,
and a part of which could not easily be obtained from other sources. Its interest is de.
cidedly enhanced for students who demand both clearness and exactness of statement,
by the profusion of well-executed woodcuts, diagrams, and tables, which accompany the
volume.... The suggestions of the author on the use of tea and coffee, and of the va.
rious forms of alcohol, although perhaps not strictly of a novel character, are highly instructive, and form an interesting portion of the volume."-N. Y. Tribune.
IV.
Body and Mind.
THE THEORIES OF THEIR RELATION.
By ALEXANDER BAIN, LL. D.
I vol., I2mo.   Cloth......                        Price, $I.50.
PROFESSOR BAIN is the author of two well-known standard works upon the Science
of Mind-" The Senses and the Intellect," and "The Emotions and the Will."  He is
one of the highest living authorities in the school which holds that there can be no sound
or valid psychology unless the mind and the body are studied, as they exist, together.
" It contains a forcible statement of the connection between mind and body, studying their subtile interworkings by the light of the most recent physiological investigations. The summary in Chapter V., of the investigations of Dr. Lionel Beale of the
embodiment of the intellectual functions in the cerebral system, will be found the
freshest and most interesting part of his book. Prof. Bain's own theory of the ccnnection between the mental and the bodily part in man is stated by himself to be as follows:
There is'one substance, with two sets of properties, two sides, the physical and the
mental —a double-faced -unity.' While, in the strongest manner, asserting the union
of mind with brain, he yet denies'the association of union in   lace,' but asserts the
union of close succession in time,' holding that'the same being is, by alternate fits, under extended and under unextended consciousness."'-Christinz  Register.
D. APPLETON  & CO., Publishers, 549 & 551 Broadway, N. Y.




Opinions of Mtie Press on Mhe' Inlernalional Scienltifc Series."
v.
The Study of Sociology.
By HERBERT SPENCER.
I vol., I2mo.  Cloth.............                         Price, $X.5o.
"The philosopher whose distinguished name gives weight and influence to this volume, has given in its pages some of the finest specimens of reasoning in all its forms
and departments. There is a fascination in his array of facts, incidents, and opinions,
which draws on the reader to ascertain his conclusions. The coolness and calmness of
his treatment of acknowledged difficulties and grave objections to his theories win for
him a close attention and sustained effort, on the part of the reader, to comprehend, follow, grasp, and appropriate his principles. This book, independently of its bearing
upon sociology, is valuable as lucidly showing what those essential characteristics are
which entitle any arrangement and connection of facts and deductions to be called a
science. "-Eziscopalizaz.
" This work compels admiration by the evidence which it gives of immense research, study, and observation, and is, withal, written in a popular and very pleasing
style. It is a fascinating work, as well as one of deep practical thought."-Bost. Post.
" Herbert Spencer is unquestionably the foremost living thinker in the psychological
and sociological fields, and this volume is an important contribution to the science of
which it treats.... It will prove more popular than any of its author's other creations,
for it is more plainly addressed to the people and has a more practical and less speculative cast. It will require thought, but it is well worth thinking about"-Albany'vening Yourazal.
VI.
The New Chemistry.
By JOSIAH P. COOKE, JR.,
Erving Professor of Chemistry and Mineralogy in Harvard University.
I vol., t2mo.  Cloth......       Price, $2.00.
"' The book of Prof. Cooke is a model of the modern popular science work. It has
just the due proportion of fact, philosophy, and true romance, to make it a fascinating
companion, either for the voyage or the study."-Dazily Grahiuc.
"This admirable monograph, by the distinguished Erving Professor of Chemistry
in Harvard University, is the first American contribution to'The International Scientific Series,' and a more attractive piece of work in the way of popular exposition upon
a difficult subject has not appeared in a long time. It not only well sustains the character of the volumes with which it is associated, but its reproduction in European countries will be an honor to American science."-l\rez York Tribune.
" All the chemists in the country will enjoy its perusal. and many will seize upon it
as a thing longed for. For, to those advanced students who have kept well abreast of
the chemical tide, it offers a calm philosophy. To those others, youngest of the class,
who have emerged from the schools since new methods have prevailed, it presents a
generalization, drawing to its use all the data, the relations of which the newly-fledged
fact-seeker may but dimly perceive without its aid.  To the old chemists, Prof.
Cooke's treatise is like a message from beyond the mountain. They have heard of
changes in the science; the clash of the battle of old and new theories has stirred them
from afar. The tidings, too, had come that the old had given way; and little more than
this they knew.... Prof. Cooke's' New Chemistry' must do wide service in bringing
to close sight the little known and the longed for.... As a philosophy it is elemen.
tary, but, as a book of science, ordinary readers will find it sufficiently advanced."Utica Morning Herald.
D. APPLETON  & CO., Publishers, 549 & 551 Broadway, N. Y.




Op2inions of the Press on the  "International Scienltfic Series."
VII.
The Conservation of Energy.
By BALFOUR STEWART, LL. D., F. R. S.
W/ith ant A.bendix treating of thie Vieal and Mfental A&Zican tios of the Doc/rine.
I vol., I2mo. Cloth. Price, $x.50.
"The author has succeeded in presenting the facts in a clear and satisfactory manner,
using simple language and copious illustration in the presentation of facts and principles, confining himself, however, to the physical aspect of the subject. In the Appendix the operation of the principles in the spheres of life and mind is supplied by
the essays of Professors Le Conte and Bain."-Ohizo Farmter.
" Prof. Stewart is one of the best known teachers in Owens College in Manchester.
" The volume of THE INTERNATIONAL SCIENTIFIC SERIEs now before us is an excellent illustration of the true method of teaching, and will well compare with Prof.
Tyndall's charming little book in the same series on' Forms of Water," with illustrations enough to make clear, but not to conceal his thoughts, in a style simple and
brief." — Christian Register, Boston.
"The writer has wonderful ability to compress much information into a few words.
It is a rich treat to read such a book as this, when there is so much beauty and force
combined with such simplicity.-Eastern Press.
VIII.
Animal Locomotion;
Or, WALKING, SWIMMING, AND FLYING.
With a Dissertatlio on A,,ronauzics.
By J. BELL PETTIGREW, M. D., F. R. S., F. R. S. E.,
F. R.C. P.E.
I vol., I2mo.                   Price, $i.75.
"This work is more than a contribution to the stock of entertaining knowledge,
though, if it only pleased, that would be sufficient excuse for its publication. But Dr.
Pettigrew has given his time to these investigations with the ultimate purpose of solv.
ing the difficult problem of Aeronautics. To this he devotes the last fifty pages of his
book. Dr. Pettigrew is confident that man will yet conquer the domain of the air." —
N. Y. Y. ourna of Commnzerce.
" Most persons claim to know how to walk, but few could explain the mechanical
principles involved in this most ordinary transaction, and will be surprised that the
movements of bipeds and quadrupeds, the darting and rushing motion of fish, and the
erratic flight of the denizens of the air, are not only anologous, but can be reduced to
similar formula. The work is profusely illustrated, and, without reference to the theory
it is designed to expound, will be regarded as a valuable addition to natural history."
-Omaha Republic.
D. APPLETON  & CO., PUBLISHERS, 549 & 55I Broadway, N. Y.




O5inions of the Press on the "I nlerznational Scient.fic Series."
IX.
Responsibility in Mental Disease.
By HENRY MAUDSLEY, M. D.,
Fellow of the Royal College of Physicians; Professor of Medical Jurisprudence
in University College, London.
I vol., I2mo.  Cloth   o.  Price, $I.5o.
"Having lectured in a medical college on Mental Disease, this book has been a
feast to us. It handles a great subject in:a masterly manner, and, in our judgment, the
positions taken by the author are correct and well sustained."-Pastor and People.
"' The author is at home in his subject, and presents his views in an almost singularly clear and satisfactory manner.... The volume is a valuable contribution to one
of the most difficult, and at the same time one of the most important. subjects of investigation at the present day."-N. Y. Observer.
"It is a work profound and searching, and abounds in wisdom."-Pizsbaurg Cornmsercial.
" Handles the important topic with masterly power, and its suggestions are practical and of great value."-Providence Press.
X.
The Science of Law.
By SHELDON AMOS, M. A.,
Professor of Jurisprudence in University College, London; author of "A Systematic
View of the Science of Jurisprudence," " An English Code, its Difficulties
and the Modes of overcoming them," etc., etc.
I vol., 12mo.  Cloth.....  Price, $x.75.
c The valuable series of' International Scientific' works, prepared by eminent specialists, with the intention of popularizing information in their several branches of
knowledge, has received a good accession in this compact and thoughtful volume. It
is a difficult task to give the outlines of a complete theory of law in a portable volume,
which he who runs may read, and probably Professor Amos himself would be the last
to claim that he has perfectly succeeded in doing this. But he has certainly done much
to clear the science of law from the technical obscurities which darken it to minds which
have had no legal training, and to make clear to his'lay' readers in how true and high a
sense it can assert its right to be considered a science, and not a mere practice."-The
Christian Register.
"The works of Bentham and Austin are abstruse and philosophical, and Maine's
require hard study and a certain amount of special training. The writers also pursue
different lines of investigation, and can only be regarded as comprehensive in the departments they confined themselves to. It was left to Amos to gather up the result
and present the science in its fullness. The unquestionable merits of this, his last book,
are, that it contains a complete treatment of a subject which has hitherto been handled
by specialists, and it opens up that subject to every inquiring mind.... To do justice
to' The Science of Law' would require a longer review than we have space for. We
have read no more interesting and instructive book for some time. Its themes concern
every one who renders obedience to laws, and who would have those laws the best
possible. The tide of legal reform which set in fifty years ago has to sweep yet higher
if the flaws in our jurisprudence are to be removed. The process of change cannot be
better guided than by a well-informed public mind, and Prof. Amos has done great
service in materially helping to promote this end."-B-zffalo Cocrier.
D. APPLETON & CO., PUBLISHERS, 549 & 551 Broadway, N. Y.




Opinions of the Press on the " IZnternational Scienitzic Series."'
XI,
Animal Mechanism,
A  Treatise on Terrestrial and Abrial Locomotion.
By E. J. MAREY,
Professor at the College of France, and Member of the Academy of Medicine.
With 117 Illustrations, drawn and engraved under the direction of the author.
x vol., x2mo. Cloth...  Price, $1.75
"We hope that, in the short glance which we have taken of some of the most important points discussed in the work before us, we have succeeded in interesting our
readers sufficiently in its contents to make them curious to learn more of its subjectmatter. We cordially recommend it to their attention.
" The author of the present work, it is well known, stands at the head of those
physiologists who have investigated the mechanism of animal dynamics-indeed, we
may almost say that he has made the subject his own. By the originality of his conceptions, the ingenuity of his constructions, the skill of his analysis, and the perseverance of his investigations, he has surpassed all others in the power of unveiling the
complex and intricate movements of animated beings."-Populasr Science Molot/dly.
XII.
History of the Conflict between
Religion and Science.
By JOHN WILLIAM DRAPER, M. D., LL. D.,
Author of "The Intellectual Development of Europe."
I vol., i2mo....                   Price, $1.75.
"This little'History' would have been a valuable contributidn to literature at any
lime, and is, in fact, an admirable text-book upon a subject that is at present engrossing the attention of a large number of the most serious-minded people, and it is no
small compliment to the sagacity of its distinguished author that he has so well gauged
the requirements of the times, and so adequately met them by the preparation of this
volume. It remains to be added that, while the writer has flinched from no responsibility in his statements, and has written with entire fidelity to the demands of truth
and justice, there is not a word in his book that can give offense to candid and fairminded readers."-N. Y. Evening Post.
" The key-note to this volume is found in the antagonism between the progressive
tendencies of the human mind and the pretensions of ecclesiastical authority, as developed in the history of modern science. No previous writer has treated the subject
from this point of view, and the present monograph will be found to possess no less
originality of conception than vigor of reasoning and wealth of erudition.... The
method of Dr. Draper, in his treatment of the various questions that come up for dis,
cussion, is marked by singular impartiality as well as consummate ability. Throughout his work he maintains the position of an historian, not of an advocate. His tone is
tranquil and serene, as becomes the search after truth, with no trace of the impassioned
ardor of controversy. He endeavors so far to identify himself with the contending
parties as to gain a clear comprehension of their motives, but, at the same time, he
submits their actions to the tests of a cool and impartial examination."-N. Y.- Tribune.
D. APPLETON  & CO., PUBLISHERS, 549 & 55I Broadway, N. Y.




QOjiniozs of the Press on the "Inernational Scientific Series."
XIII.
THE DOCTRINE OF
Descent, and Darwinism.
By OSCAR SCHMID T,
Professor in the University of Strasburg.
WITH 26 WOODCUTS.
I vol., 12mo.  Cloth.......    Price, $x. 50.
"The entire subject is discussed with a freshness, as well as an elaboration of detail, that renders his work interesting in a more than usual degree. The facts upon
which the Darwinian theory is based are presented in an effective manner, conclusions
are ably defended, and the question is treated in more compact and available style
than in any other work on the same topic that has yet appeared. It is a valuable addition to the'International Scientific Series.' "-Boston Post.
" The present volume is the thirteenth of the'International Scientific Series,' and
is one of the most interesting of all of them. The subject-matter is handled with a
great deal of skill and earnestness, and the courage of the author in avowing his opinions is much to his credit.   This volume certainly merits a careful perusal."Hartford Eveniozg Post.
" The volume which Prof. Schmidt has devoted to this theme is a valuable contribution to the Darwinian literature. Philosophical in method, and eminently candid,
it.shows not only the ground which Darwin had in his researches made, and conclusions reached before him to plant his theory upon, but shows, also, what that theory
really is, a point upon which many good people who talk very earnestly about the
matter are very imperfectly informed." —Detroit Free Press.
XIV.
The Chemistry of Light and
Photography;
In its Application to Art, Science, and Industry.
By Dr. HERMANN VOGEL,
Professor in the Royal Industrial Academy of Betlin.
WITH I0o ILLUSTRATIONS.
I2mo..........     Price, $2.00.
" Out of Photography has sprung a new science-the Chemistry of Light-and, in
giving a popular view to the one, Dr. Vogel has presented an analysis of the principles
and processes of the other. His treatise is as entertaining as it is instructive, pleasantly combining a history of the progress and practice of photography-from the first
rough experiments of Wedgwood and Davy with sensitized paper, in 8o02, down to
the latest improvements of the art-with technical illustrations of the scientific theories
on which the art is based. It is the first attempt in any manual of photography to set
forth adequately the just claims of the invention, both from an artistic and a scientific
point of view, and it must be conceded that the effort has been ably conducted."Chicago Tribune.
D. APPLETON & CO., PUBLISHERS, 549 & 55I Broadway, N. Y.




Opinions of the Press on the "' International Scienliffc Series."
xv.
Fungi;
THEIR NATURE, INFLUENCE, AND USES.
By M. C. COOKE, n. A., LL. D.
Edited by Rev. M. J. BERKELEY, M. A., F. L. S.
With  o09 Illustrations.  Price, $I.5o.
"Even if the name of the author of this work were not deservedly eminent, that of
the editor, who has long stood at the head of the British fungologists, would be a sufficient voucher for the accuracy of one of the best botanical monographs ever issued
from the press.    The structure, germination, and growth of all these widely-diffused organisms, their habitats and influences for good and evil, are systematically
described."-New York World.
"Dr. Cooke's book contains an admirable rdsusumd of what is known on the structure, growth, and reproduction of fungi, together with ample bibliographical references
to original sources of information.'"-London A thenwuzm.
"The production of a work like the one now under review represents a large
amount of laborious, difficult, and critical work, and one in which a serious slip or fatal
error would be one of the easiest matters possible, but, as far as we are able to judge,
the new hand-book seems in every way well suited to the requirements of all beginners
in the difficult and involved study of fungology."-The Gardener's Chronicle (London).
XVI.
The Life and Growth of Language:
AN OUTLINE OF LINGUISTIC SCIENCE.
By WIL IAM DWIGH' WHITNrEY,
Professor of Sanskrit and Comparative Philology in Yale College.
I vol., I2mo.  Cloth.  Price, $1.50.
" Prof. Whitney is to be commended for giving to the public the results of his ripe
scholarship and unusually profound researches in simple language. He draws illustrations and examples of the principles which he wishes to impa rt, from common life
and the words in frequent use.
"The topics discussed in this volume are, for the most part, those which have
been already treated by other writers on philology, and even by the author himself, in
his volume on'Language, and the Study of Language,' published a few years ago,
and, though many of the truths here set forth are those with which students in the
same line of investigation are generally familiar, all will rejoice to see them restated in
such a fresh and simple way.
" This work, while valuable to scholars, will be interesting to every one."-The
Churchman.
" This work is an important contribution to a science which has advanced steadily
under conditions that appear constantly to throw an increasing light on difficult questions, and at each step clear the way for further discoveries." -Chcago Inter-Ocean.
" Prof. Whitney is undoubtedly one of the foremost of English-speaking philologists,
and occupies an enviable position in the wider circle of European students of language.
" His style, clear, simple, picturesque, abounding in striking illustrations, and apt
in comparisons, is admirably fitted to lie the vehicle of a popular treatise like the work
under consideration."-Portlzed Daily Press.
D. APPLETON  &  CO., PUBLISHERS, 549 & 551 Broadway, N. Y.