L I B R A R.Y OF THL U N I VERS ITY OF 1LLI NOIS 6307 tio 445-4-57 A6R1CULTURE NOTICE: Return or renew all Library Materials! The Minimum Fee for each Lost Book is $50.00. The person charging this material is responsible for its return to the library from which it was withdrawn on or before the Latest Date stamped below. Theft, mutilation, and underlining of books are reasons for discipli- nary action and may result in dismissal from the University. To renew call Telephone Center, 333-8400 UNIVERSITY OF ILLINOIS LIBRARY AT URBANA-CHAMPAIGN Gelatiniz,ation and Retrogradation Changes in Corn o o and Wheat Starches Shown by Photomicrographs By SYBIL WOODRUFF and MAJEL M. MACMASTERS UNIVERSITY OF ILLINOIS AGRICULTURAL EXPERIMENT STATION Bulletin 445 CONTENTS I'AGI STARCHES USED IN" THK STUDY 3 Si nirees of Starches 3 M ethi nls of I 'reparing Starches 4 Si a roll Y it-It Is Obtained in Laboratory (> Comparison of Laboratory ami Commercial Methods 6 GELATINIZED STARCH (> Mi- tli ml ui ( it-latin i /.in ,u Starch 7 M ion >soi ipie Tt-ohnio 7 Curnstaroh Kt-siilts '' \\ lu-at Staroh Ri-snlts 1- l so nt Photomicrographs in Staroh Studios \2 l.itoraturo mi Staroh C hanuos lo STARCH RKTROC.UADKI) BY FRKKXINCi 15 Mothud nt Retrograding hy l-'reoxing lr KtToot of I )it"foront l-"roo/in,u Temperatures 17 Microscopic Appearance 17 Litoraturo mi Retrc>eradation hv Frooztnu 23 MICROSCOPIC' PARTICLES APPEARING IX GELATINIZED STARCH 24 Moth. M! of Freeing tho Particles 2 l ) \;i|ioaraiRX- ot Staroh I 'arttolos 32 Particles Separated by the Supercentrifuge 32 Relation ot Gelatinization and Free/.ing Temperatures to \"isibilit\ - of I 'articles i3 l.itoraturo on Particles in Starch 34 MEASUREMENTS OF PHYSICAL PROPERTIES i5 M t-t hods of Mrasu foment I 'sod 36 1 'it't'e renees Duo to Variety of Corn 3o Differences Due to Conditions During (innvth v Further Obser\ation on the I ndividualit v of Starches.. . 38 40 42 Gelatinization and Retrogradation Changes in Corn and Wheat Starches Shown By Photomicrographs I'v SYBIL \\"IKIUKITK and MA ICI\( >SO >PIC appearance has been relied upon as a means of identifying kinds of starch and of following physical changes in paste formation ever since anything has been known ot the nature of starch. There are surprisingly few photographic records in the literature, however, beyond those of original, unaltered starches ol different plant sources. Present-day research directed along lines designed to show the structure of the starch grain summons whatever assistance may be afforded bv photomicrographs. In this bulletin are reproduced photomicrographs showing such effects in corn and wheat starches as were brought about bv swelling the starches in water and by subsequently free/ing the gelatini/ed substance or by causing it to separate irom suspension bv means ot alcohol. Starches used in the work were of known origin and were pre- pared tor use bv methods thought to change them little from their native state. In this respect the materials were probably unique, since it lias been the more usual practice for starches of commerce to be employed in studies of this kind. It was learned early in the course of the work that cornstarch thus carefully prepared differed measurably trom commercial samples. Physical properties ot starch in original, gelatini/ed, and retro- graded states are of no small degree of interest to the person who uses starch tor "thickening" purposes in tood technology or in textile sixmg. as well as to one whose interest in starch is more or less academic. It has been one of the purposes of this studv to accompany with photomicrographs measurements ot differences in gel strength and viscosity as thev have appeared under different treatments of the starches used. STARCHES USED IN THE STUDY Sources of Starches 445 Champion White IVarl. Sutton White I )ent, and 1'ioneer Hi-l'red 305. \. In addition, two manufactured lirauds ot cornstarch have been used i"i>r comparison with samples made in the laboratory. In de- termine \\lu-lluT the conditions under which corn is grown have any effect on the cornstarch two samples ot Reid dent corn grown m ditlerent areas in Illinois were used. ( )ne sample was chosen trom corn grown in Champaign couutv under good conditions, except tor a period of verv hot weather. This corn vielded 05 bushels an acre and contained I 1 ' percent moisture when harvested. The second sample \\ as taken trom corn grown in Morgan countv under conditions so had. owing to heat, drouth, and grasshoppers, that it yielded only 10 bushels an acre. It had an unusuallv low moisture content, about 13 percent, when harvested. Since starch grains were earlv shown to be profoundly changed m appearance and behavior bv free/ing, it was thought worth while to tree/e ears ot corn while thcv still contained more than the amount of moisture usuallv present at time of harvest to see if free/ing at this stage affected their starch. The laboratory condition, however, exag- gerated such frosting of the ears as might occur in the field. After being iro/en for 24 hours at IS" to 20 F.. when the moisture content ot the kernels was still about 2/ to 32 percent, the ears were dried and starch was prepared in the usual wav. Sott and hard wheat (lours served for studies ot wheat i I'nticuui ruli/arc) starch in 1 ( >35 and l ( >^ \\lll \l . s I \NCI ! 1 :- 5 sulfur dioxid was therefore avoided in these experiments in spile of the tact that without it the separation ot starch troni corn was more diffi- cult and less complete. '1 o lie sure, the starch was later shaken with a solution dt sodium chlorid to dissoKe adhering globulins, hut this reagent acting for a short time at room temperature has not been IK ited to cause a change in the starch. The calcium salts of hard water were also avoided because of the marked effect on starch viscosity which thev have been reported bv Ripperton 18 * to have. The difficulty in washing the starch from wheat flour was greater than usual, because distilled water was used instead of tap water. The calcium salts of distilled water make the gluten more coherent and less likelv to contaminate the starch. Bleaching agents, containing as they do nitrogen trichlorid and ben/oyl peroxid, were not used with wheat flour starches except where the starches were not to be examined in their native state. In the preparation of cornstarch, one kilogram of washed, dried corn was ground in an electric mill to a fine, meal-like condition. Knough distilled water was added to make a moderately thick mash, the starch suspension from which was worked thru a 60-mesh sieve with repeated additions of water until the liquid came thru clear. The milky suspension was next put thru a Xo. 13 standard silk bolting cloth, the total volume of the strained starch suspension now being about n to 8 liters. After standing overnight in the refrigerator, as much as possible of the clear supernatant liquid was decanted, then water was removed from the starchv portion bv centrifuging. Layers of foreign matter on top of the starch in this and subsequent washings were removed with a spatula. The wet starch was next shaken up three times with a 2-percent solution ot sodium chlorid to remove traces ot protein and was washed with distilled water until free from chlorid ion : it was then washed successively with three or four portions each of 70-percent alcohol. 95-percent alcohol, and diethyl ether. It facilitated (Irving not to remove the last ether bv centrifuging but to pour the heavv starch suspension in ether directly onto a porous plate, on which after much of the ether had evaporated, it could be worked drv and free from lumps with a spatula. It was spread thinly mi clean paper and allowed to stand in air tor at least two days and was used in this air-dried form. Wheat starch was made bv working 500 grains of Hour into a dough with distilled water and, after letting the dough stand for one-halt hour, washing the starch from it bv working it i, r entlv under distilled (i BU.I.KTI.N Xo. 445 water. It was strained thru bolting cloth and treated from here on in the same manner as cornstarch. Starch Yields Obtained in Laboratory The yield of cornstarch was considered high if 30 percent of the weight of the corn was obtained; it was more usual to recover only about 22 percent. The factory yield for cornstarch is usually about 58 percent of the weight of the original corn. The yield of wheat starch varied from about 45 to 65 percent of the weight of flour used. \\ heat starch is not manufactured on a commercial scale, hence no comparison ol yields can be made. As stated before, the greatest concern was the avoidance of any treatment which might alter the starch ; si/e of yield was not a major consideration. Tho the Yields were fairly low, it is believed that the starch obtained was a representative composite of the starch from different parts of the kernels. Comparison of Laboratory and Commercial Methods The method of preparation employed differed notablv from the commercial process of manufacturing cornstarch 15 * in that the steep- ing of the kernels for 30 to 40 hours at a temperature of 115 : to 125 C F. in water containing .25 to .30 percent of sulfur dioxid. was omitted. This treatment greatly facilitates the freeing of starch from struc- tural tissues. The effect of sulfur dioxid mav be similar to that of chlorin dioxid. which was found bv Samec and Llm 21 " to have pro- duced chemical changes in potato and wheat starches to such degree that thev gradually lost their abilitv to form a thick, glutinous material even tho microscopic appearance remained unchanged. Samec 11 '" has attributed the gelatinizing effects of starch to the phosphoric acid- containing components in it. If this is true, tho some investigators doubt it. any reagent I possiblv sulfur dioxid ) which would remove the phosphoric acid might be expected to reduce swelling. The drying process emploved in the laboratorv was also different irom the usual commercial process for making powdered edible starch, which subjects the starch to 160' to 180 C F. for 18 to 20 hours. The alcohol and ether used in the laboratory tor drying mav have removed substances which would ordinarily be lett in the tactorv product. GELATINIZED STARCH 'I lie term (jelatimzc has long been used to refer to the changes which starch undergoes as it swells in water under the influence of CHANGE heat, salts, or other agents. Changes during the process are visible not only in gross but also in microscopic appearance. Photomicro- graphs in this bulletin illustrate some of the modified forms of starches ot different varieties ol corn and wheat alter thev have been gelatin- i/ed over a temperature range of 70 to 114' C. Provided there is present but little, if anv. excess of water over the amount which the starch is capable ot absorbing, as is the case when about ? grams ot starch is used per 100 grams ot suspension, a paste will result at a temperature of about 05 C. which will set to a gel alter cooling. The term r/iY is used to refer to gelatinized starch in the molded torm it assumes on cooling. Method of Gelatinizing Starch The method of gelatinizing the starch used in these experiments was as follows: 1-ive grams ot starch was moistened in a 250 ml. Krlenmever Mask with about 10 grams of distilled water, then 85 grams of hot distilled water, or enough to bring the total weight to 100 grams, was added with a pipette. The force of the stream kept the starch in motion and prevented lumping. '1 he onlv stirring given the paste was a swirling ot the tlask during further heating. The temperature ot the water was such that alter its addition the entire paste had a temperature of about (>() (.'. The flask was immersed in a boiling-water bath and brought <|uicklv to 70 . 80 , ( )(), or 05 C'.. according to the nature of the experiment. Complete swelling ol starch seemed to have occurred hv the time the temperature reached O.i C ., and the paste on cooling set to a gel. A suspension containing 5 percent concentration of starch was used in all experiments except those testing viscosilv, because it gave a consistency most like' that found in molded starch pastes used in preparing food products of iellied consistency. The swollen starch was either examined tinder the microscope at once or was allowed first to set to a molded gel. Microscopic Technic \\ith a tew exceptions, the swollen starch was alwavs mounted in a drop ot water and the cover slip sealed to the sink 1 to prevent evaporation and movement during photographing, A I.eitx. polari/ing biological microscope and camera were used tor the exposures. ( rossed mcols were used for detecting anisotropic properties of the materials, as indicated in the legends. An ocular of 30 times and an objective DIFFERENT \ ARIKTIES OF LORN GELATINIZED AT 80" C. IN" 5-PERCENT SUSPENSIONS 1. Reid Yellow Dent, not gelatinized -. Same ;is No. 1, between crossed nicols 3. Suite m \\'hite Dent 4. Champion \\"hite Pearl 5. Pioneer Hi-Bred 305A n. A commercial brand oi cornstarch of starches of Xo. 4 is more 1938] CHANCKS i\ COKN AND \\'HKAT STAIU IIKS 9 of 30 times magnification were employed in taking all photomicro- graphs, \vith the exception of Xo. 1, Fig. 14. The photomicrographs have been reproduced without change in si/.e. Cornstarch Results There was no characteristic microscopic variation in the cornstarch which could be assigned to varietal difference in the corn. Neither did free/ing the corn on the ear seem to change the appearance of the cornstarch. Starch of Reid Yellow Pent corn previous to gelatinixation, viewed without and with crossed nicols. and starches of four other corn varieties partially gelatinized in 5 percent suspensions in water at 80 C. are shown in Fig. 1. Xo differences occur in the four which might not be found in anv one starch sample with different adjust- ments of photographic field, light, and exposure, except in Champion White Pearl, which can easily be seen to be more swollen at 80 C. than are the others. It was characteristic of starch from this variety of corn to give a stronger gel ( see page 36) and to swell more com- pletely at a lower temperature than did the other varieties of corn- starch studied. The extent of swelling at three temperatures of gelatinixation ( 70, 80, and 95 C.) is shown in Fig. 2 for both a laboratory-prepared sample and a commercial cornstarch. Swelling was well started at 70 3 C., and material resembling masses of fish eggs could be seen to have exuded from the grains. Gelatinization progressed up to 95 C.. at which temperature the outline of swollen grains was indistinct and exceedingly difficult to bring into focus. This latter temperature is the one which permits of maximum swelling and gel strength. Later in the bulletin measurements are given which show that cornstarches from different sources varied considerably in the tirmness ot the gels they were able to produce. Three samples fluctuating widelv in gel strength are shown in Fig. 3 to appear on the whole verv much the same microscopically, altho Xo. 2. the strongest gel, derived from Champion White Pearl corn, is more blurred than Xos. 1 and 3 even with the best tocus obtainable for it. The high temperature of the autoclave (about 114 : C.) made the outline of the granule in Xo. 4 even a little more indistinct than it had been when gelatini/ed at 95 C. It is verv difficult to show the microscopic appearance of Hilly sv\ ollcn starch granules because of their transparency and lack of con- wSSRfr WT"**Arand of starch, ucl strength 101 mm. 4. Sutton White Dm! slarrh, antoclavnl for .^O minutes at 114 ('. alter urlatini/alion. (id strniuitli ISO mm. Champion White I 'earl starch, \\hieh LJUVI' the stnitljivsl uel of all kinds of corustarcli studied, was alwavs hlnrreil in mien >so ipir appeai'ance. ( )tlu'r\visc there \\~a> 110 indicutioii ot a dillerent decree ol liydra! ion. \utocla,vins4 \veak- ened the uel mea-nrahly from the reading of 2S 1 ' mm. \\hieh Suttou \\4iite Dent starch uavc oriuinalK' ; the mien >sci .pic appearance \\as only ^li.nluK altereil. ( .\las4iiitication M) \ .ill) 12 Bn.LKTix Xo. 445 [August, trust. A certain amount of material \vas always found in each field, which might be residues of collapsed granules altho it is impossible to identify it as such. The quantity of this material did not seem to increase at the autoclave temperature; and yet other experimenters have attributed the inability of starch to form a gel to collapsing or rupturing of the outer envelop. The autoclaved gels were, however, measurably weaker, as shown on page 40. Wheat Starch Results In wheat starch as in cornstarch, the microscope failed to detect differences due to variety of wheat and bleaching treatment or differ- ences between hard and soft wheat types. Figs. 4 and 5 show char- acteristic markings of unswollen and swollen wheat starch, such as folds of the outer envelop which became apparent early in the swelling (Fig. 4, Xo. 5 and Fig. 5, Xo. 1) ; great divergence in size of granules (all samples) ; frequent cracking of granules as they swelled (Fig. 5, X T O. 5). Wheat starch completely swollen at 95 C. was even more difficult to show microscopically than cornstarch, and it will be noted that in Fig. 5. Xos. 3 and 6 are only indistinct blurs. In appearance wheat starch gels are much like cornstarch gels except weaker. Comment will be made later upon the concentrically arranged particles which were found in partially swollen wheat starch and which can be seen most clearly in Fig. 4, Xo. 5, and Fig. 5, Xos. 4 and 5. Use of Photomicrographs in Starch Studies The photomicrographs in Figs. 1 to 5 illustrate what happens dur- ing the swelling of starch in water, and at the same time they give evi- dence of the difficulties encountered in attempting to photograph material which is so transparent that but little definition is possible. Slightly different adjustments of light and focus and selection of different fields mav cause the photographs to appear more or even less different from each other than were the original materials. Each illustration has been chosen from the man}- exposures taken as being most typical of the conditions described. The lowest gelatini/ation temperature shown is 70 C. because only shortly before this tempera- ture dries anisotropy disappear. Onlv sixe and general appearance of the swollen granules are being considered here : the significance of microscopic particles is discussed on pages 29 to 35. Descriptions and drawings ot starch during the progress of its gela- tinixation are to be found in the literature for manv vears back, but Kit;. 4. M ICKOSCOPK: AITKAKAXCK OK WIIK.AT STAKCIIKS ( I K.I. \ i IXI/.KH AT 80 ('. IX 5-l'K.IU'KN T Sl'SI'KNSIONS 1. Soft wheat starch mounted in water, not gelatinized 2. Suit wheat starch, unbleached .^. I lanl wheat starch unbleached 4. Soft wheat starch, same field as N'o. 1, hetueen crossed nicols ; not gelatinized 5. Sot't wheat starch t'nmi heavily bleached flour 6. Hard wheat starch from heavily bleached Hour structural arrangement in unc ni tin- lar.^e u;ranti l uranulr in Xn. 5; ami tin- indistinct mass-like urlaiini/rd at ^5 ('. Alsn i;iite tin- irrcLnilarit v in . imts. X<- dit'u-rcncc can he- red vuni/rd as lifinji dtu ni .^i \ .id ) KS 15 photomicrographs have not appeared until recentlv. S jostronr ; ' lias lateK' published photographs of manv kinds ot starches under various degrees ot" pasting, tho he was more concerned that the photographs bring out evidences of granule structure than that thev show the progress ot swelling, lie therefore used acid-treated, thin-boiling corn and wheat starches. His records give a remarkable picture ot the nu'cellar arrangement within the granule. Woodruff and Webber' 1 -' a few years ago published photomicro- graphs of wheat starch at different stages ot gelatnn/.ation. Literature on Starch Changes The theories regarding changes which starches undergo as thev swell under the inlluence of heat or other agents are manv and com- plex. Xo attempt will be made to discuss them here. Excellent re- views in the literature cover such important aspects of the subject as temperatures of gelatnn/.ation, disappearance of birefringence, the part the constituent amvloses plav in the process, X-rav speclro- graphic changes and others. The reader is referred especiallv to a long series of articles bv Katz and others, 10 ' 11 * and to articles bv Linsbauer, 13 * bv Alsberg and Kask, 2 ' and bv I>adenhuizen. :; ' STARCH RETROGRADED BY FREEZING (ielatinized starch can be made to loosen its hold on imbibed water bv several means. Free/ing is one method which has been used with considerable interest in these studies, tor with tree/ing profound alteration in microscopic appearance ot starch can be' ettecled. I'hvsical changes in the starch substance during I reiving point to profitable applications of tins mode ot studying internal structure ot starch grains. The original degree ol swelling, i.e.. the temperature to which the paste was brought, would be expected to have some etiecl upon the wav in which it gives up its water again. 1 here- fnre starch gelatinized at temperatures ranging l nun /() to ''.^ I . uas Iro/en at both verv low and moderate! v low temperatures and changes in the resulting products recorded bv photomicrographs. I lie term retrograded is used to reler to the starch lorm ot lessened solubilitv thus produced bv tree/in^ ot a gelatinixed paste. Method of Retrograding by Freezing 16 Rn.i.ETiN Xo. 445 [August, Tastes previously gelatinized as described on page 7 and contain- ing 5 percent starch were poured while hot into shallow glass contain- ers, and after cooling to room temperature were allowed to stand in the free/.ing compartment of an electric refrigerator overnight or longer. The temperature here lluctuated considerably but averaged about 2 C to 3 C. After remaining at this temperature for about 24 hours, and then being allowed to thaw, the previously smooth gel had a fibrous, spongy structure from which the water was squeezed by hand, the gel being wrapped in bolting cloth during the pressing to prevent loss of material. The change in appearance due to freezing is apparent in Fig. 6; the gel on the right, frozen and then thawed, does not have the smooth consistency of the unfrozen gel on the left. FIG. 6. GROSS APPEARANCE OF FROZEN CORNSTARCH GELATINIZED AT 95 C. IN 5-PERCENT SUSPENSIONS Left, unfrozen. The gel is smooth and firm in outline. Right, frozen at 3 C. for 18 hours, then thawed. The gel has a fibrous, rough outline; water can be squeezed from its spongelike structure. The pressed material was dried completely in one of two ways. One way was to separate it loosely into shreds and allow it to dry in the air on clean paper, then grind it on a closely adjusted hand mill until it was a coarse powder. This powder was more coarsely flaky than that obtained by the second method. The second method con- sisted ot loosening the pressed substance and covering it with 95 per- cent alcohol, decanting after a few minutes, repeating this process three or lour times, and then washing three times with ether. The material was worked up on a porous plate until free from ether and reduced to a tine, slightly llaky powder. There was no evidence that drying the material with alcohol, after it had first been retrograded by treezmg. changed it: as pointed out on page 27, however, initial precipitation with alcohol gave a product very different from a frozen retrograded one. CHANT, KS ix CORN AND \Yni:vi Si \KVIIKS 17 Effect of Different Freezing Temperatures Early in the work with frozen retrograded starch it was observed that markedly different effects were produced by freezing in the electric refrigerator as described above and freezing at the tempera- ture of either liquid air or solid carbon dioxid. After being fro/en at the very low temperatures of the latter, where the temperature of the gel quickly dropped to probably less than 70 C.. the gel on thaw- ing still had most of the original slimy consistency of newlv gelat- inized starch. Probably in this procedure there was no opportunity for the reorientation changes to occur which always resulted from freezing the gels in the mechanical freezing unit and which gave the gels the appearance of a coarse fibrous structure from which water could be squeezed as from a sponge. The term which seems best to describe the microscopic appearance of the gel frozen at the lower temperature was "brittle." whereas heavy, flexible strands were to be found in the gel at a freezing temperature but slightlv under O 3 C. An attempt was made to determine whether there was a "transi- tion temperature." above or below which this difference could be dis- tinguished in the fibrous appearance of the gross and microscopic specimens. For this experiment one dee]) pan was insulated from the warmth of the room by being set in another larger one separated from the first by asbestos wool. The inner pan contained a mixture ot alco- hol and water previously frozen to a mush on solid carbon dioxid. This mush was stirred during the operation and its temperature, not that of the freezing starch, was noted. The starch gels in thin layers in aluminum cups were set in this mush of alcohol and water and allowed to remain there until frozen. P>v varving the proportions of alcohol and water, temperatures ranging from K) ' to 10' C. were obtained with a fluctuation of :4 during an interval of several minutes. A transition temperature of 25 C. (' 2"') was found in this way for cornstarch first gelatinized at either 00 or o : C'. and for wheat starch gelatinized at ( '0 C. I'elow this transition temperature, the gel after thawing had the brittle microscopic appearance mentioned; above this temperature the heavy strands appeared. However, the coarsest tibers alwavs formed in gels frozen at temperatures but a little under treez- ing: 2 to .V C. Microscopic Appearance Reid -t;ircli \\as uclatinized in 5-percent snsin.-nsn>ns. I he inix.rn retro- irr;i in tin- same field as Xo. 3. After the fro/en retrograded starch had been dried it gave, on being gelatinized a second time, a verv different appearance I Xo. 4) trom freshly gelatinized starch I Xo. 1). The gross appear- ance of the gel in Xo. 4 was shapelv but granular and lacking the uni- tormitv ot consistency of the unfrozen gel in Xo. 1. Hven tho regelat- imzed and refrozen several times, the appearance changed but imperceptibly after the starch was once retrograded in this wav. Kttects produced bv freezing temperatures both higher and lower than about 25"' C\. the transition temperature of reorientation, art- shown in I'ig. 8. Xos. 1 and 2 were trozen at a temperature abo\ e tin- transition temperature and differ mainlv in that Xo. 2 was more com- pletelv gelatinized in the beginning than was Xo. 1. Xos. 4 and 5 are the same fields as Xos. 1 and 2 but were photographed between crossed nicols and show amsotropic strands. Xos. 3 and u were trozen at the temperature of solid carbon dioxid. which is much lower than the transition temperature. Xo. 3 had originallv been incomplete! v gelatinized at 80 ? C. and. \vlien frozen quickly at the low teni])erature, the starch grains seemed to assume a chamhke arrangement as the water was frozen out. This arrangement mav be due to tin- greater freedom ot movement in the more walerv suspension and also niav be the arrangement which the forces are striving tor in the closelv packed. completely swollen starch grains of a still gel at the time the aniso- tropic strands appear. At least the same appearance as that observed in Xo. 3 was noted repeatedly when the same set of conditions was provided. 'I he appearance of Xo. (). a completely gelatinized starch irozeii at the temperature of solid carbon dioxid, is best described as brittle. The gel did not acquire a veined structure nor the fibrous spon^v nature of the gels trozen at higher temperatures. Photomicrographs were made frequently between crossed nicols m the series frozen at temperature's near zero, because thev show the wav in which gelatinized starch regained amsotropv after such re-ori- entation had occurred. Thus far the eft eel has been shown in moist gels onlv: but it can be even more vividlv shown by mounting tin- specimen drv. as was done in I'ig. ''. Xos. 1 and 2 show the drv fibrous material after freezing retrogradation, and Xos. 4 and r> the same fields between crossed nicols. Xos. 3 and <> show sonic- of the same 5 V " / '' 1 L3* *1 &U'*\m DIFFERENT FREEZING TEMPERATURES ox APPEARANCE I\i'i'l starch was gelatinized in 5-iH'rccnt suspensions. 1. At 80' C. ; frozen at 3 C. 2. At ')()' C'.; frozen at 3 3 C. 3. At 80 C. ; frozen on solid carbon dioxid 4. Same as Xo. 1, with crossed nicols 3. Same as Xo. 2. with crossed nicols 0. At ')() C". ; trozen on solid carbon dioxid The lower temperature altered the appearance of the cornstarch much less than the higher U j mprraturr. Xo. 3 shows that when not Completely ~W'illrti the starch granules tend to torm chains dnrin.cf treezing more visibly than wlien more completely gelatinized. Xo. ft shows the brittle, le^s fibrous appearance characteristic of the loucr temperature. (Magnification 30x30') jc,. 10. KI-TKCT 01 UKK/IM; ox \\IIKAT STAKCII CiKi. ATI. \IZKD IN ielatinixed at 70' ('. 2. Fro/en at - 3 C. ,v l-'ni/i'ii on solid carbon dioxid material which has been mounted in water. Anisotropic strands arc visible even in the water-swollen mass of Xo. b. 1 he great difference between the appearance of these gels and of gelatinized starch before free/ing can be seen bv comparing Fig. ^ with Fig. 2. Xo. 1. Wheat starch is shown in Fig. 10 in forms both incompletely and fullv gelatinized, before and after tree/ing. It has undergone free/ing changes of the same character as cornstarch. Free/ing at the low temperature of solid carbon dioxid left wheat starch even less changed than cornstarch ; the higher free/ing temperature produced aniso- tropic strands in the gel very similar to those of cornstarch. In their gross appearance the \\dieat starch gels underwent about the same changes as cornstarch but the changes were not so marked. Literature on Retrogradation by Freezing Freezing has been used occasionally by workers in the held of starch chemistry as a means of effecting a separation of constituents of the starch granule. Little descriptive data have been given by authors, however, to indicate what temperature and time have been employed or even how the precipitated material appeared. Probably the earliest account of the effect of freezing was that reported in 1 S44 hv Scharling."" who found his starch paste reduced to "soup" by an unexpected freeze. The solid material was porous and paperlike and ii clipped in water, swelled immediately like a sponge. I'nder the microscope the mass looked like pure, small powder shells. Mallilano and M oschkott ' ' in 1'MO reported Ireezmg a potato starch solution, with a filamentous coagulum as a result. HersteiiT' described in 1 ( M2 a "modified" starch, obtained trom a lairlv thick starch paste' sub- jected to a low temperature, lie said that the resulting spongy, hbrous mass had been seriously proposed as a material tor papermaking at one time. Keillv. ( >' I )ono\ an. and Murphv 17 ' centrifuged the clear liquid trom libers lormed when a live-])ercent suspension ni gelatinized potato starch was irozen. and made molecular weight determinations (>n the amylose so recovered. Ling and Xan]i'- employed tree/ing as a means ol separating amvlopectin trom soluble amylose ol potato starch, a paste' ol which had been kept lor several hours at a temperature near ( . I'aldwm' 1 ' also se])arated the /:?- irom the a-amvlose ol potato starch In ireezmg a paste' coutaimng !..> percent starch winch had been lirst gelatinized at a temperature ol bo ( . lie did not identity his I reiving tempera lure but described the line particles produced by rapid Ireezing as perfect spheres and those obtained with slow ! reiving as coarser 24 BULLETIN Xo. 445 [Auyitst, particles \vhich were large spheres or conglomerates of spheres. He also warned that if the material remained frozen for several hours, a precipitate so coarse was formed that it did not dissolve even in several days. That there are different degrees of retrogradation was pointed out by Taylor and Morris. 274 who have called the insoluble substance sep- arating from a once clear dispersion of /3-amylose when it is frozen, retrograded amylose. They also admitted the possibility of retro- gradation within the starch granule itself. The process is described by them as an association of glucose units into shearlike bundles which in turn associate to a varying degree, depending upon the extent to which retrogradation has gone. These sheaflike bundles correspond no doubt to the heavily veined, anisotropic areas of the frozen gels depicted in the photomicrographs in this bulletin and in those of an already published paper by "Woodruff and Hayden. 29 * STARCH RETROGRADED BY ALCOHOL The luminous appearance of alcohol-retrograded wheat starch between crossed nicols had been observed by the authors even before the study of freezing retrogradation had been undertaken. Because the microscopic appearance of a starch precipitated from its paste by alco- hol was found to differ so markedly from that retrograded by freezing, photographs were taken to show the contrast. Method of Retrograding by Alcohol The starch paste was first gelatinized at a desired temperature, allowed to cool to about 75 C.. then poured into five times its volume of alcohol solution, containing 2 parts of alcohol by volume to one part of water. After the suspension had cooled to room temperature. 2 volumes more of 95-percent alcohol were added to insure complete precipitation of the starch, and the suspension was allowed to settle overnight in the refrigerator. The liquid was removed by centifuging; the retrograded starch was shaken three times with 95-percent alcohol and centrifuged. Xext it was shaken with absolute alcohol from five to nine times, and lastly three times with ether. The last ether was worked out of the starch on a porous plate with rapid handling to pre- vent its absorbing moisture and becoming gummy. Numerous washings in absolute alcohol were necessarv to remove all moisture Vie fore final drving with ether. KKTKdi.K. \I)ATI()\ P,V ALCOHOL AMI I'A I M.IKI-: EFKKCTS IN COKXSTARCII v S * < v % " fS^v**^ v s k^o ^v%-^^\^; , * - s . wn@n7llMr *, * ' * I* \ ' 4. RctroL, Results With Corn and Wheat Starches I he starch so retrograded was a white powder almost as tine as the original starch before gelatinization. It would swell readilv in water even in the cold or would gelatinize when heated to ( h (/. to give a somewhat molded gel. similar to the original tho not so uni- torinly smooth. Retrogradation and regelatini/ation could be repeated several times without much change in results. Alcohol precipitation quite obviously merely withdrew the water from the swollen granules and permitted of no such phenomenon as occurred with free/ing. Cornstarch retrograded by alcohol and mounted dry. moist, and subsequently regelatini/ed at 95'" C. is shown in Fig. 11. Xos. 2, 3. and r> respectively. Xo. 5 appears verv much like the starch gel which had undergone swelling but once at 95 c C. ( Fig. 2, Xo. 3). For comparison a tlake of starch retrograded bv free/.ing and mounted dry is shown in Xo. 1 ; Xo. 4 is a starch retrograded bv free/.ing. first dried then re- gelatini/ed : Xo. (i. a starch retrograded bv alcohol, regelatmi/ed and subsequently fro/en. The difference between starches retrograded bv tree/ing and bv alcohol is evident. Starch retrograded by alcohol is capable ot later being regelatmi/ed and tro/en to give the same appear- ance it might have had if it had been fro/en in the first place. The same set of conditions for wheat starch is shown in Fig. 12 and with results similar to those for cornstarch. It max be noted in Xo. 3 on this plate that alter alcohol retrogradation the wheat starch resumed its original peculiar swollen shape. Interesting photomicrographs demonstrating the abilitv of the drv material to transmit light when examined between crossed nicols were obtained with Starches retrograded bv alcohol. '1 he same held without and with crossed nicols is shown in Fig. 13. \Vilh crossed nicols there was not onlv a brilliant transmission ot light but occasional amsotropic particles could be seen scattered thru the mass. I'olh corn and wheat starches behaved in this wav. The effect of the alcohol is apparently not one which permits the same kind ot reonentation within the granule as that produced bv slow tree/ing. Xothmg in either gross or microscopic appearance of the gel suggests the fibrous structure- of fro/en gels. Moreover, when starch retrograded bv alcohol is ottered water again, il resumes ap- proximatelv the same shape and si/.e it had before it was dehvdrated. It reacts toward subsequent free/ing much as a newlv gelatini/ed starch does. ( hi the other hand, something more than dehydration must have occurred to cause the starch to appear so luminous between crossed nicols. 'I here is an interestm;/ similarity here to the formation CHANGES IN CORN AND WHEAT STARCHES 29 of spherocrystals reported by Van cle Sande-Bakhuyzen 28 * to have occurred in alcohol to \vbicb an amylose solution \vas added and allowed to stand for three weeks. The crystals were said to be built up of radial needles and the larger ones exhibited the cross of a native starch granule. MICROSCOPIC PARTICLES APPEARING IN GELATINIZED STARCH Occasional reference has been made earlier in this paper to oval or round particles which stand out in the photomicrographs because of their ability to transmit light in a manner different from the remainder of the gelatinized or gelatinized-and- frozen material. The nature of the particles, or micelles as they have been called by some, is important because such units are believed to hold a kev to internal structure of the starch granule. The particles are thought by the authors to have been either set free or actually formed bv the freezing of gelatinized starch, since they were found in greatest abundance after the gel had been frozen, especially when it was frozen at the temperature of solid carbon dioxid. The photomicrographs in Figs. 14 and 15 demonstrate whatever degree of success was attained in separating these particles from the rest of the starch and in showing their nature. Some of the reproduc- tions show them as they appear in the gelatinized starch and others picture them after they have been freed more or less from the material enveloping them. Method of Freeing the Particles The particles could be seen in greatest number and to best advan- tage in starch which had been gelatinized at a temperature no higher than 70 C. Suspensions containing 5 percent of starch bv weight were used. The gelatinized starch was frozen at the temperature of solid carbon dioxid, thawed, and treated with a 1:1 solution of ammonium hydroxid, which seemed to loosen the encrusting material. I "articles could then be almost completely separated from this sur- rounding substance bv a Sharpies supercentrifnge at 3.000 to 5,000 r.p.m. Partial separation was obtained even in an nrdinarv centrifuge at about 1,600 r.p.m. The quantitv of particles obtained from 10 grams of starch was, however, verv small. The duration of the free/ing period on solid carbon dioxid did not Fit;. 14. MICROSCOPIC ['ARTICLES IN GELATINIZED STAKCH All starches except Xo. 4 were first partially gelatinized at 7d C. in ^-percent suspeiisii ms. Corn: 1. Treated \\itli 1:1 HC1 2. Frozen mi >olid carbon dioxiil ,\ [ ; rozen at ^3 C. ; \\itlimit anil \\'ith cnissnl mo>ls \\'lu-at: 4. Xot gelatinized; treated with 7.5 percent IU'1 ?. I'rozen mi siilid carhdii dmxid f). Frozen on solid carbon dioxid; treated with 1:1 XH.OH 1 . ' a >* , t 1. Treated with 2\\ C'a ( X( ) :: )... ; starch matrix not removed; stained \\ith lodin, nionnted in water. I 'article- round and isotropic. 2. Treated with 1:1 XII, Oil; starch matrix not removed; stained with iodm. mounted in water, ['articles oval and anisotropic, ,v Treated with 1:1 XII, OH; starch matrix partially removed hy snper- centri I tiiie ; mounted dry, unstained. Oval particles did not appear amsotropic. 4. Same as Xo. .^. Show- clustering durum drying on -lide. 5. Treated with 1:1 XII, OH; particles partial!) removed hy superccni n- Itme; rcsuspended in water and I ro/en auain at ,> ( . Mounted dry. d. Same as Xo. 5 except tin- tield contains some -larch matrix which reoriented characteristically with final Iree/inu at - 3 (.'. The separated particles in Xos. ? and showed no tendency to reorient in this uay. ( Mamuticatii MI ,i() x 30 ) 32 BUM.ETIX Xo. 445 [August, seem to affect the number of particles obtained. Altho not so successful as solid carbon dioxid, the temperature of a mechanical freezing unit i about 3 C.) could be used also. Other reagents used with somewhat less satisfaction than ammonium hydroxid were hydro- chloric acid, sodium hydroxid. and calcium nitrate, each in varying concentrations. Appearance of Starch Particles Partially swollen starches showed the microscopic particles more readily than did completely gelatinized starch ; cornstarch showed them in greater abundance than wheat. Several photomicrographs of the particles as they appeared in gelatinized starch of both kinds are shown in Fig. 14. Xo. 1 is of cornstarch mounted in concentrated hydrochloric-acid solution (1:1). showing concentric arrangement of these units within the granule. Occasionally a few particles were found which were anisotropic. either because they were unusually free from encrusting material or for some other reason unknown. The same field taken without and with crossed nicols in Xo. 3 shows a cluster of anisotropic particles in a cornstarch granule. Xo. 2 shows an abundance of particles in starch after it had been frozen. A some- what different effect can be noted in Xo. 4 where ungelatinized wheat starch was allowed to stand in 7.5 percent hydrochloric acid for 24 days. This was the usual concentration of acid used by other workers for lintnerizing starch. The granules appeared to have split into seg- ments which were striated in orderly arrangement of particles. Xos. 5 and 6 also show wheat starch with particles somewhat loosened from the matrix. In addition to the microscopic fields of Fig. 14, which were especially chosen to depict these particles, the same formations have occasionally appeared in other photomicrographs. See Fig. 2, Xos. 1 and 5: Fig. 10. Xo. 3: Fig. 12. Xo. 1 ; and Fig. 13. right mount. Particles Separated by the Supercentrifuge The supercentrifuge was found to be capable of separating these microscopic particles from the enveloping matrix of gelatinized starch with such a degree of completeness that particles could be found in dense masses. Photographs of particles freed in this way are shown in Fig. 15. Xos. 3 to 6. The particles appear somewhat larger than they were in reality, because of light diffraction in the dry mounts. The reason for allowing the- particles to dry on the slides before they were photographed, rather than mounting the material wet as usual, was that 1938] CHAXC.KS IN C'OKN AND \\'IIKAT STAKCHI-:S 33 the particles \vere in violent Hrownian movement in the absence of starch matrix. Clustering on the slide, as is especially noticeable in Xo. 4, was due to currents during drying, not to the nature of the particles themselves. Water suspensions of the separated particles did not exhibit reorientation into anisotropic strands as did fro/en starch matrix. Xo. 5 shows the particles unaffected by being frozen after centrifuging; X T o. 6, containing a fragment of matrix, shows the matrix itself in strand formation after free/ing, but the particles not re- arranged so far as can be seen. Anisotropy was not found in any particles separated from frozen starch paste by the use of the supercentrifuge. The particles in Fig. 15, Xos. 3 to 6, were in no case anisotropic, tho they were otherwise given the same treatment as those shown in Xo. 2 which were anisotropic. Even in violent Brownian movement, the anisotropy of the freed particles should have been still visible if present. Xo explanation of this observed difference is at present known. Aside from this varia- tion in optical behavior, all the particles obtained by treating frozen starch with ammonium hydroxid ( Xos. 2 to 6) were small oval ones appearing alike. Ammonium hydroxid, of all reagents used, proved most effective in unveiling anisotropic particles. When the particles were loosened from the starch matrix bv digestion in 2M calcium nitrate solution, as in Xo. 1, large round particles resulted. These were not found to be anisotropic. Xo attempt was made to separate them from the matrix. Relation of Gelatinization and Freezing Temperatures to Visibility of Particles Xoteworthy is the observation that free/ing at the low temperature of solid carbon dioxid made the greatest number of particles visible; it was also at this temperature that the gelatinized paste reoriented into fibrous strands to the least extent. There mav be some connection between the two points. There seems as great likelihood that the freezing produces retrogradation of other material, thus causing the particles to become visible, as that the particles are initially formed at the time of freezing. Possibly at a temperature of about --3 C., where a tibrous gel is obtained, the particles disappear as such when reorientation into anisotropic strands occurs. It may be that the strands are composed ot these particles in orderly arrangement. ( hi the other hand, fro/en water-suspensions of particles, freed from the starch matrix, gave no evidence ot such strand lormation. The particles could not be found easily in starch which had first 34 been gelalini/.ed at temperatures much higher than 70' C., even alter llie paste had been fro/en. This \vas noted l>v the authors, and others have also reported that the niicellar structure could best be seen belore gelatini/.ation had gone verv tar. ( )n the other hand, amsotropic strands were formed verv promptlv in frozen gels which had been gelatini/ed at a temperature as high even as ( )r> ' C. T\vo kinds ot particles have been observed bv the authors, one a large round bodv, isotropic so far as can be determined ; the other a considerablv smaller oval one. freqnentlv anisotropic. It is thought possible that the larger round particle is reallv the small ])article still enveloped in other material which prevents it trom exhibiting aniso- tropv. The round particles were obtained bv treatment with either calcium nitrate solution or with ammonium hvdroxid. and the small oval ones onlv with ammonium hvdroxid. lioth particles stained a tvpical starch blue with iodin-potassium iodid solution. The X-rav s]>ectrum a of supercentrifuged particles was judged to be that of neither ungelatinized, gelatinized, nor retrograded starch. This statement is based u])on preliminary observations, and measure- ments of this kind have not vet been carried far enough that a further statement can be made. Literature on Particles in Starch Particles in starch have been described in the literature. Sjostrom's recent photomicrographs 23 * show them verv clearlv in slightlv swollen starches of many sources and he identities them with Xiigeli's 1 "" micelles. Samec wrote 20 *: "\\hen a separation (of a starch solution) into a sol and a gel fraction has occurred, there are observed i bv ultranncroscopic methods) a great number of round or egg-shaped particles in a highly viscous Hind which contain luminous particles imbedded in a transparent mass. The viscous fraction becomes less prominent on dilution or continued heating, so that under these condi- tions, the ultramicroscope shows particles in verv active movement in an armcroseopic held." I lanson and Kat/"- : ' used chemical means of splitting starch, granules radiallv and tangenliallv into anisotropic "blocks." showing the effect with drawings. A little earlier Kat/ and Derksen"' in claim- ing a crystalline niicellar structure for starch as a result of X-rav data, noted that at 80 C". or higher the crystal structure is lost and the material is either molecillarly dispersed or the micelles become re- versiblv amorphous, \\oodrurl and flavdeir" have published photo- micrographs also showing anisotropic particles in gelatim/ed starch. Kvcr since Xageh'" tirst proposed his theory concerning the micellar structure of the starch granule, it has been much discussed in the literature with the particular purpose ot explaining optical prop- erties of starch. All this has been reviewed l>v manv authors. 1 - In recent vears X-rav spectra have been commonly employed as a means of studying crystalline arrangement in the granule ot gelatmi/ed and retrograded forms of starch as well as in native starch. Reference has already been made to the extensive writings of Kal/. and co-\yorkers on this subject. 9 ' '" MEASUREMENTS OF PHYSICAL PROPERTIES Kven tho microscopic examination ot gelatinized starch tailed to show detectable difference between cornstarches of different sources, some ot these same starches produced different pasting results, which tact is ot practical significance in starch usage and has therefore been studied. For example, one of the important uses of starch in food preparation is that of "thickening," the desired effect being an in- creased viscosity of the liquid so "thickened." Furthermore, it is sometimes an advantage for the thickened material to set to a gel as it cools, as in molded puddings, fillings, and even the crumb ot bread itself, which is a very stiff starch gel. File consistencies ot pasted starches may be compared in several ways. Viscosity measurement is one which the factory uses tor deter- mining whether a starch is a thin-boiling or thick-boiling one. I Ins property of gelatinized starch has been reported on most frequently. An instrument called a consistometer has been devised and used 1>\ one group ot workers"* lor measuring differences in very stilt pastes. Photographs of starch gels will at least tell whether the cooled cooked paste will hold the shape of the mold or not : sometimes this is a matter of considerable importance in practical food preparation, \\orkmg with concentrations ot starch on the borderline between molded gels and llmd consistencies, \\oodrult and others ha\e found photographs to be one useful means ot recording physical differences between starches. 1 "'" An instrument originally designed for measuring the strength of pectin jellies has been emploved with considerable success tor measuring differences in lirmness ot starch i>els. Variations due to M) BULLETIN Xo. 445 [.-Ingitst, variety of corn and to conditions of treating the corn before extract- ing the starch have been measured not only in gel strength but in viscosity. A few such data will serve to point out the differences in physical behavior which may exist even in the absence of photomicro- graphic distinctions between the various starches. Methods of Measurement Used Gel strength was measured with a Tarr and Baker jelly tester 20 * which measures millimeters of hydrostatic pressure required to force a plunger thru the surface of the gel. Starch was gelatini/ed as usual in suspensions containing 5 percent of starch and poured into glass molds holding 30 ml. of hot paste when filled to within 3 mm. of the top. Each mold was covered with a watch crystal during cooling to prevent a ''skin'' from forming and to collect condensed moisture. Without removing the gel from the glass, readings of pressure were made on the top surface at the time it was first observed to break under the pressure of the plunger. Because of the only relative exact- ness of the test, readings were made on at least twenty replicate molds of the same paste and each starch was gelatinized on as many as ten occasions. Viscosity readings were made with a Stormer viscometer. employing starch in 2-percent concentrations. The paste was first gelatinized in an Erlenmeyer flask at the desired temperature and was then trans- ferred to the viscometer cup where the gelatini/ation temperature was maintained thruout the readings. The same number of readings was made on viscosity as on gel strength. Relative viscosities are recorded on the graph for comparative purposes. Differences Due to Variety of Corn The strongest gels in the series of cornstarches measured were those made from white corn. Champion White Pearl produced the strongest and Sutton White Dent the next strongest. Starches from Pioneer 1 Ti-Bred 305 A and Reid Yellow Dent, two yellow varieties, produced gels less strong than those made of starch from the white corns but far stronger than those of the two commercial brands of table cornstarch. These rather widely fluctuating results on gel strength are shown graphically in Fig. 16. Altho not included in the graph, a Reid starch, prepared in the laboratory by steeping corn in water containing sulfur dioxid. gave as weak a gel as the commercial starch samples. \ iscosity readings on the same starches showed less marked devia- 193S] CHANGES IN CORN' AND \\'HF,AT STARCHES 37 tion among the samples than did gel-strength measurements ( Fig. 16). Apparently the two sets of measurements emphasi/e different points of behavior and both may be important. 300 a. 100 2 > COMMER. STARCH A COMMER. STARCH B REID YELLOW DENT PIONEER HI-BRED 305A SUTTON WHITE DENT CHAMPION WHITE PEARL FIG. 16. GEL STRENGTH AND VISCOSITY OF STARCH DIFFERED WITH VARIETY OF CORN Starches from the above corns were all gelatinized at o C. The weakest gels were those of commercial cornstarch ; the strongest those of white corn. \ iscosity fluctuated but little compared with the fluctuations in gel strength. Differences Due to Conditions During Growth Corn grown in Illinois under unfavorable conditions, yielding 10 bushels an acre, gave a weaker starch gel than the same Reid corn grown under good conditions where the vields were o5 bushels an acre I Fig. 17). The gross appearance of the two sets of gels is shown in Fig. IS. These photographs also give an idea of the general outline and the appearance of the starch gels used lor the gel-strength measurements. Free/ing the corn on the cob before it had ttilly matured also 38 350 I'. i I.I.K-J IN XD. 445 300 0250 150 UNFAVOR. FAVORABLE GROWING GROWING CONDITIONS CONDITIONS REID YELLOW DENT NOT FROZEN REID YELLOW DENT NOT FROZEN SUTTON WHITE DENT Al-TECTKI) i:v (iKn\Yl.\r, Starches irmii the above corn? were all gelatinized at ( h* ( . Relatively weak uel- ucre obtained from the starch of corn LIT own under ]ioor conditions and from corn that had been frozen on the ear. Viscosity was affected little in comparison with Liel strength. atlected tin- ,L(cl ad\ erselv. Tlif ,yx-l strcn^tli of starch irom both \'cllo\v and \\liitc- corns \\TIS lo\\'cr \vhcn the starch was obtained from corn that had been fro/en on the cob. I'.ut a^'ain neither growing conditions nor tree/in^" ot corn on llie cob seemed to alter the viscositv ol the starcli ]>astes to a marked decree i Vig. 17). Free/in^ tlie corn before its starcli had been ^elatini/ed must not be confused with the effect free/in.^ lias on starch pastes, as discussed in a previous section of the bulletin, for the effect is not of the same nature. Further Observation on the Individuality of Starches 'I he individuality of the starches of different varieties of corn is shown in the phvsical behavior of their pastes even tho not in their microscopic appearance. for some investigations it might lie necessarv to identify all conditions pertaining to the source and treatment of the >tarch material, it misinterpretations are to lie avoided. Further evidence of the individualit v of starches from different kinds of corn was made in an experiment with Reid and Sutton White I >ent starches, winch should he mentioned at least hriellv here. The starches were first pasted at 95 C . Then boiling was continued for l-'n;. 18.--(il)SS Al'l'K. \KA.\CK OK C'OKNSTARCH (il-'.I.S ( /"/vr: Reid starch, urown under favorable conditions, 1 ( ,M(>; gelatinized in 5-percent suspensions at 05, 00", 80', and 70 ('. respectively, iett to nuht. /.o;ivr: Reid starcli thrown under very unfavorable conditions, 1036; gelatinized in 5-percent suspensions at 05", 00", 80", and 70 (.". respectively, lefl to ri.uht. '1'be tzel at the extreme left was the one used in all gel-Strength measnre- HH'iits. Xcite that the uels of the lower row were \\eaker thnnnit than those oi the ii])per row. various lengths of time in an autoclave where a temperature ot 1 C'. \\'as maintained. Changes in ^el strength and in viscositv I measured at ( '5 C .) u'ere noted to he of quite different order in the two starches, as shown by the figures on the following pa^e. Autoclaved Reid starch tormed an extremelv \\i-ak <^el. much like wheat starch j^els in strength. \"iscosit\- in this starcli \\as so lowered by autoclaving that the ]>aste \\hile hot \\ as \\ater\\ The i;el of Sutton \\hite Dent starch lost some in strength during autoclaving but not to the same extent as the Reid starcli. \\ith all the diminution in gel strength it exhibited even atter ( '0 minutes, the Sutton cornstarch was still stiff compared with the regular gels made with commercial corn- 40 Hru.KTix X<>. 445 ("HANI.KS IN GF.I. STKKNUTH AMI VISCOSITY Gel strength I'iscosity Reid Ycllozi 1 I } cnt starch mm. at95 e C. Gelatinized at 95 3 C 208 1.24 Autoclaved for 5 minutes 15 1.14 Autoclaved for 30 minutes 6 1.15 Autoclaved for 90 minutes 4 1.13 Snt ton II 'hit e Dent starch Gelatinized at 95 C 289 1.34 Autoclaved for 5 minutes 221 1.42 Autoclaved for 30 minutes 180 1.37 Autoclaved for 90 minutes. . .155 1.37 starch, the strength of which was only 100. The weakening of gel strength was progressive as the autoclaving time was prolonged. Viscosity measurements on Sutton cornstarch were even higher after autoclaving than before, which is further evidence that viscosity and gel strength seem to measure two different sets of properties in the starches. Too few data on wheat starch were accumulated to make similar comparisons for wheat. Wheat starch gels are enough weaker than cornstarch gels of the same percentage concentration to make it impos- sible to measure them with the instrument. In outline and general appearance, however, the wheat starch gels were well formed in spite of their lesser strength compared with cornstarch gels. Gels made with starch of heavily bleached wheat tlour did not indicate that the bleach- ing chemicals had an effect at all comparable with that of sulfur dioxid on cornstarch. Subjective observations of gel strength were thought to be of measurable value in view of the fact that sensory impressions of firm- ness of cornstarch gels agreed remarkably well with the ranking made according to gel-strength measurements. SUMMARY This bulletin shows photomicrographs of corn and wheat starches in native, gelatinized, and retrograded states. Starches used for the experiments included those from four varieties of corn and from two commercially manufactured samples; they included also both soft and hard wheat starches. There was no microscopic appearance which could be assigned to varietal difference, to chemical treatment during preparation of the starch or wheat flour, or to propitiousness of growing season. 41 Subjecting corn and wheat starch gels to temperatures hut little below tree/ing produced retrogradation ettects conspicuouslv ditterent from those observed when the gels \vere fro/en at the temperature of solid carbon dioxid. The temperature 25 r C. was found to be a critical one tor corn and wheat starch, below and above which this difference became apparent. Anisotropic strands of reoriented material were produced at temperatures higher than 25 C. In gross appear- ance gels fro/en at ,V C. were fibrous and spongv. and the starch from them could not be regelatinized to the original gel state. dels retrograded bv alcohol \vere unmistakably different from those retrograded by free/ing. They were luminous thruout between crossed nicols. dranules in the gels seemed to have been dehvdrated but not reoriented into strands, as bv free/.mg. ( )n regelatini/ation, the gel resumed much of its former appearance. The free/ing of partiallv gelatinized starch exposed to view micro- scopic particles, occasionally amsotropic. which were thought to have some possible connection with the anisotropic strands alwavs found in gels troxeu at .V C. These particles were centrifuged from the thawed starch matrix with a tair degree ot success alter it had been treated with ammonium hvdroxid. The particles were found in greater abundance in cornstarch than in wheat starch. They were also more plentiful alter the starch was Iro/.en at the temperature ot solid carbon dioxid than after it was fro/en at 3 C. Thev were not alwavs aniso- tropic. Thev showed violent Brownian movement when treed trom the starch matrix. Thev sometimes appeared more nearly round and larger than at other times, but it seemed doubtful that the two torms were separate entities. Several photomicrographs illustrate the particles as thev were found in this studv. Kven tho it was not manifested bv microscopic appearance, the individuality of starches was apparent in the phvsical aspects ot their gels. ( iel strength fluctuated widelv with the varietv of corn from which the starch was obtained and also with the growing conditions under which the corn was produced. \\ bite corns gave starches with stronger gels than vellow corns, and all samples prepared in the 1 laboratory gave firmer gels than commercial starches, \\heat starch gels were weaker than cornstarch gels. Viscosity differences were very small compared with differences in gel strength. BULLETIN Xo. 445 LITERATURE CITED 2. - and RASK, O. S. On the gelatinization by heat of wheat and maize starch. Cereal Chem. 1, 107-116. 1924. 3. BADEXHUIZEX, X. P., IR. Die Struktur des Starkekorns. Protoplasma 28, 293-326. 1937. 4. BALDWIN, M. E. Separation and properties of the two main components of potato starch. Jour. Amer. Chem. Soc. 52. 2907-2919. 1930. 5. CAESAK, G. Y., and MOORE, E. E. Consistency changes in starch pastes: tapioca, corn, wheat, potato and sweet potato. Indus, and Engin. Chem. 27, 1447-1451. 1935. 6. HANSON, E. A., und KATZ. J. R. Abhandlungen zur physikalischen Chemie der Starke und der Brotbereitung. XYII. Uber Yersuche, die gewachsene Struktur des Starkekorns mikroskopisch sichtbar zn machen, besonders an lintnerisierter Starke. Ztschr. Phys. Chem.. Abt. A, 168, 339-352. 1934. 7. - Abhandlungen zur physikalischen Chemie der Starke und der Brotbereitung. XY1II. Weitere Yersuche, die gewachsene Struktur des Starkekorns mikroskopisch sichtbar zu machen. Ztschr. Phys. Chem., Abt. A, 169. 135-142. 1934. 8. HERSTKIX, B. Modified starch. Orie. commun., 8th Internatl. Cone. Appl. Chem. 13, 177-181. 1912. 9. KATZ, J. R. The X-ray spectrography of starch. Gelatinization and retro- gradation of starch in the bread staling process. In "A comprehensive survey of starch chemistry," edited by R. P. Walton, pp. 68-76, 100-117. The Chemical Catalog Co"., Xew York. 1927. 10. - - ct al. (The reader is also referred to numerous papers on the various aspects of starch chemistry by Katz and others in the following publications: Biochem. Ztschr. 1933-34: Ztschr. Phys. Chem. 1930-35; 11. - der Starke und der Brotbereitung. X1Y. 1st die Starke im Starke- kleister kristallinisch oder amorph? Ztschr. Phys. Chem., Abt. A. 167. 129-136. 1933. 12. LINO. A. R.. and XAXTI. D. R. Starch. I. Xature of polymerized amylose and of amylopectin. Jour. Chem. Soc. 123. 2606-2688. 1923. 13. LIXSBAUER, K. Mikroskopische Studien iiber den Yerkleisterungsprozess. P.ot. Centbl.. Beihefte. Abt. A. 53. 172-199. 1935. 14. MALFITAXO. CT., and MOSCHKOFF, A. Sur la coagulation de la matiere amylacee par congelation. Compt. Rend. Acad. Sci. [Paris] 150. 71(1-711. 1910. lr. MOFFETT, G. M. Manufacture of corn starch. In "A comprehensive survey of starch chemistry," edited by T. P. Walton, pp. 130-138. The Chemical Catalog Co.. Xew 'York. 192"". 16. X.\i, I.LI. C. Die Micellartheorie (1858). Ostwald's Klassiker der exakten Wissen-chaften Xr. 227. Alb. Ercy, ed. Akad. Yerlausuesellschaft M. P.. H.. Leipzig. 1928. 17. RF.II.I.Y. J.. O'DoxovAX, P. P., and MTRTHV, H. A note on the molecular complexity of amylose in potato starch. Roy. Dublin Soc. Sci. Proc. 21. 37-42. 1034. CHAXCES i.v CORX AMI WHEAT STARCHES 43 18. RIITERTOX, J. C. Measurement of consistency of starch solutions. Indus ami Engin. Chem., Analyt. Kd. 3, 152-154/1931. 19. SAMEC, M. Kolloidchemie dor Starke. Yerlag von Thoodor Steinkopt'f. Dresden und Leipzig. 1927. 20. - A summary of the colloid chemistry of starches. In ''Colloid chemistry, theoretical ami applied," edited by Jerome Alexander, pp. 167-180. The Chemical Catalog Co., Xe\v York. 1932. 21. - - und L'r.M, F. Studien iiber Pflanzenkolloide. XI. 11 1. Kiniluss von Chlonlioxyd aut den Chemismus der Kartotiel- und \\eizeiistarke. Kolloidchem. Beihefte 43, 287-294. 1930. 22. ScuARi.ixii, E. A. Einige \"ersuche iiber Amylum. Ann. Choin. 49, 315-316, 1844. 23. STOSTROM, C). A. Microscopy of starches and their modifications. Indus, and Kngin. Chem. 28, 63-74.' 1936. 24. SPOXSLKR. O. L. The structure of the starch grain. Amer. Jour. Bot. 9, 471-492. 1922. 25. - - Structural units of starch determined by X-ray crystal structure method. Jour. Gen. Physiol. 5, 757-776. 1923. 2(t. TARR, 1.. \Y. Fruit jellies. 111. Telly strength measurements. In Del. Agr. Kxp. Sta. Bui. 142. 1926. 27. TAYLOR, T. C., and MORRIS, S. G. The properties of the amyloscs. Corn n-arnvlose and retrograded /3-amylose. lour. Amer. Chem. Soc. 57, 1070-1072. 1935. 28. VAX HE SAXDE-BAKHUYZEX, H. L. The crystallization of starch. Proc. Soc. Kxp. Riol. Med. 23, 506-507. 1926. 29. WoonRn-T, SYBIL, and HAYIIEX, HKXRIETTA. The effect of freezing on the physical and microscopic character of gels ot corn and wheat starches. Jour. Agr, Res. 52, 233-237. 1936. 30. - and M AC.\! ASTERS, M. M. The effects on corn and wheat starch gels produced by pretreating the starches with freezing and with chemical reagents". Trans. 111. State Acad. Sci. 29, 107-109. 1936. 31. - - and Xicor.r, LAURA. Starch gels. Cereal Chem. 8, 243-251. 1931. 32. - and \\ EBHER, L. 1\. A photomicrographic study ot gelatinized wheat starch, lour. AST. Re<. 46, 1099-1108. 1933. "iVERSITYOFILLINOIS-URBANA