LIBRARY STATE PLANT BOARD liarch 1945 E_636 United States Department of Agriculture Agricultural Research Administration Bureau of Entomology and Plant Quarantine A REVIEW OF INFORMATION ON ANABASINE AND NORNICOTINE, 1933-19U (Supplement to E-537 and E-561) By R. C. Roark, Division of Insecticide Investigations CONTENTS Page Introduction 2 Chemistry of anabasine ................ 2 Occurrence of anabaeine and nornicotine ....... 3 Anabasis 3 Duboisia 3 Nicotiana ............ 3 Occurrence of nornicotine in commercial nicotine sulfate ...... .... 6 Extraction of anabaeine from plants 6 Composition of commercial anabasine sulfate made from Anabasis 7 Analytical methods • 7 Separation of anabasine from nicotine and other alkaloids 8 Pharmacology of anabasine and nornicotine 11 Anabasine 11 Nornicotine 11 Insect icidal value of anabasine 12 Thysanoptera 12 Homoptera « . 13 Hemiptera •••• ik Dlptena Ik Coleoptera .......... 15 Anoplura . 15 Patents . 15 Literature cited . ........ 16 MAR 2 1 1945 - 2 - INTRODUCTION In April 1941 the Bureau of Entomology and Plant Quarantine issued E-557, A Review of Information on Anabasine, and in March 1942 E-561, A Review of Information on Nornicotine. These reviewed 163 and 50 publica- tions, respectively. The purpose of the present publication is to bring up to date all available information on both these promising insecticides* In preparing this review the following abstract journals, published from 1938 to 1943, inclusive, were searched: Biological Abstraots, Chemi- cal Abstracts, Chemisohes Zentralblatt, Experiment Station Record, Index Medious, and Review of Applied Entomology (Series A and B)« In addition the Quarterly Bibliography on Insectioide Materials of Vegetable Origin through No. 26 (January to March 1944), issued by the [Great Britain] Imperial Institute Consultative Committee on' Insectioide Materials of Vegetable Origin, -was consulted. CHEMISTRY OF ANABASINE The following constants for anabasine are given: Boiling point (p = 238 mm.) 280.9° C, freezing point 9°, specifio rotation -83. 1© . Its parachor ooincides closely with the theoretical parachor.— Sokolov (75) . In a report on the reactions of amino-N-methylanabasines the follow- ing melting points are given: ° C. alpha- Amino-N-methyl anabasine 95 - 95.3 Derivatives: Aoetyl ~ — — 72 - 73 Di acetyl, hydrate — — — 60.5- 62.5 alpha f -Amino-N-nethylanabasine 91.5- 92.3 Derivatives: Aeety} — — » _ ■ ■ ■■■■ 122 - 122*5 Propionyl — — — — — 97 - 98 Benzoyl ~— -— — — -— — 104 - 106 Dibenzoyl — — — 142 - 143 The picrate of the alpha compound decomposes at 238-239.5°, and that of the alpha 1 compound at 227.5-228°. The boiling point of the diaoetyl derivative is 160-162° (p=4 mm.), and that of propionyl derivative (p «5.5 mm.) 193-200°.— Kabachnik and JCitser (32). The dihydrochloride of dichloroanabasine is obtained by passing chlorine into anabasine in cold alcohol. The yield is 29-63 percent. It melts at 203.7°U. (dec.) Free dichloroanabasine dissolves in water with difficulty, is soluble in ethyl aloohol, slightly less soluble in ether, and melts at 51-3°. It reacts with picrio aoid, forming a picratt, ~3.- melting at 91-5°. Nicotinic acid is formed by oxidizing dlchloroanabasine with a permanganate solution. Bromination of anabasine under the same con- ditions forms bromo-substituted compounds, melting at 105° and 135°» which are very hygroscopic and very unstable. At 80-100° they are decomposed by water with the separation of anabasine. The amount of bromine decreases con- siderably on prolonged keeping of the bromo-substituted compounds. The pic- rate of one bromo-substituted compound melts at 199-201 6 r and according to its bromine content it corresponds to a monobromoanabasine.— Bashkevich (20). OCCURRENCE OF ANABASINE AND NORNICOTINE The most important sources of anabasine are the tree tobacco, Nicotiana glauca , a native of Argentina and Uruguay, and the Russian shrub, Anabasis aphylla , which is related to the American tumbleweed. Although N. glauca is a native of South America, it is very much at home in Mexico and in the southwestern United States.— Higbee (27)« A review of information on anabasine discusses sources, chemistry, alkaloids of Anabas i e aphylla , physiological action, insecticidal proper- ties, and commercial forms used for pest control.— Holman (28, pp. 21-25). A comprehensive review on the alkaloids of tobacco includes nornicotine and anabasine. — Jackson (^1) . Anabasis The anabasine content of Anabasis aphylla was found to be inversely proportional to the degree of moisture. However, this relationship cannot be accepted as an absolute rule without further verifications. Kudryavtsev ( ^U ). Anabas i s aphylla contains anabasine, lupinine, aphyllidine, and aphylline. The preparation of salts and other derivatives is described. Anabasine, upon reacting with boiling 5-percent hydrochloric acid, under- goes ring opening.— Spath, Galinovsky, and Mayer (82). In a study of saline plants of Turkmania Anabas is aphylla was found to contain 2 percent (dry basis) of potassium nitrate. — Sabinin, Baslavskaya, Beloussova, and Schocklender (60). Dnboieia Analyses of samples of Duboisia hopwoodii -received from Australia showed nornicotine to be the only alkaloid present, as follows: Leaves 3.31, leaves on twigs 1.92, twigs without leaves 0.95, and larger stems 0.52 percent.— Bowen (10). Nicotiana From the steam-volatile fraction of extracts of cured tobacco leaves, volatile bases other than nicotine amounting to at least 6.5 percent of the total alkaloid content were obtained. The dipicrate of a base isolated from this nornicotine fraction decomposed at 179.5-180.5° C. (uncor.). This base reacted with nitrous acid and with benzoyl chloride in the man- ner described by Pictet and Rotschy (ref . 30 in E-56l) for nicotimine. Yickery and Pucher (85). The volatile and nonvolatile alkaloid contents of the following species of Nicotiana are given: acuminata , alata, chinensis, glauca, glutinosa, langs- - U - dorffi, longiflora , paniculata , p lumba gi ni fo li e, t guadrivalvis , repand.a , rusbyi , rustics,, Sf.rier^e, suaveolens , sylvestris , taba.cum. The alkaloid content of a number of hybrids is given. — ■ Shmuk and Xhmura (68). Ni c o t i ana t aba cum normally contains nicotine but no anabasine, whereas N. glauca contains only anabasine. When N. glauca w as grafted on N.tabacum, formation of the characteristic alkaloid proceeded in stock and scion with a consideraoxe increase in the anaba.sine content of th« scion over that of the control plant. Anabasine appeared to be formed in the stock also. When N. tabacu m was grafted on N, glauca , nicotine was completely replaced by anabasine in the scion while a trace of nicotine appeared in the stock. — Shmuk, Kostov, and Borozdina (69) . When Ni cotiana rustica was grafted to N. glauca , it lost nicotine and accumulated anaofsine. — Shmuk (63) . The grafting of nicotine-containing tobacco on the nicotine-free species, Nicotiana glauca , resulted in plants in which the nicotine was completely replaced by anabasine. — Evtushenko ( 22) . A study was made of the inheritance of nicotine and anabasine content by Nicotiana tabacum x Nicotiana glauca hybrids, and interaction of stock and scion when these species are grafted. In hyurids a change was noted in the type of alkaloids with reference to position of the leaves. — Kuz'menko and Tikhvinskaya (35) » Nornicotine was identified in the following species of Nicotiana : bentha m ian a, ingulba , longiflora , palmeri , sanguin ea, rolanifolia, suaveolens , sylvestris , and trigonophylla . — ShinukTo^) . A large number of representative Nicotiana species and hybrids were examined, for alkaloids. Ail 29 of the wild species contained 1 or more alkaloids. N. alata , bigelovii , go ssei , and wigandioides apparently con- tained only nicotine, N. glutino°.q t iraritima , o_top_hora, tomentosa , and trigonophylla apparently contained only nornicotine. N, debney i and g lauca contained largely d,l_- anabasine. In debneyi about 15 percent of the total alkaloid was nicotine, whereas in glauca a much lower percentage of this alkaloid was found. The remaining 18 species contained mixtures of nicotine and nornicotine. Nicotine was predominant only in N. benavidesii , langsdorffi , and stocktoni ; in longiflora and plumb agini folia it was not possible to de- termine which of the two alkaloids '> r as present in the higher concentration; and in the rest nornicotine predominate J.. No wild species studied had as much as 2 percent total alkaloid content, and. most species Had less than 0.5 percent. In 23 species 8U to 100 percent of the total alkaloid content was taken up by the main alkaloid. In crosses between N. tabacum, which contains mostly nicotine, and species whose alkaloid complex was made up largely or entirely of nornicotine, the hyorids contained mainly nornicotine, together with small amounts of nicotine. The F]_ and amphidiploid N« tabacum (nicotine) x N. glauca (anabasine) contained mostly anabasine with some nicotine. In one sample of this 7\ a trace of nornicotine was detected. — H. H, Smith and C R. Smith ( ]k) . A method is presented for classifying tobacco^ broadly into three types according to their picrate melting points — (l) the - 5 - nicotine type, 21 5-224 °C.; (2) the mixed type, 190-215°, indica- tive of both elk^loids 8S mp.lor components; and (3) the nornicotine type, l 7 5-200*. Of 90 samples of tobacco examined, 2 were of the nornicotine type. One, Robinson's Maryland Medium Broadlesf, was investigated by ?/'arkwood (refs. 2_5 and £6 in E*56l) ; the other, Flue-cured Cash, may prove equally valuable as a source- material of nornicotine. Tiro tobaocos were of the mixed type; the others were of the nicotine type.— Markwood and Barthel (46) • The 42 species of Nicotiana , whan analyzed by an improved pro- cedure which is described, can be divided into 4 groups as follows: (l) The main alkaloid is niootine, seoondary pyridine alkaloids being absent or present only in insi^iificant quantities j (2) the main alkaloid is nornicotine, tertiary pyridine alkaloids being absent or present only in insignificant quantities; (3) a mixture of nicotine and nornicotine, and (4) the main alkaloid is anabasine. ~ Shrouk and Borozdina ( 67 ) • A simplified method is presented for the identification of alkaloids of tobacco. The essence of the method is the formation of the picrates of the secondary base and submission of it to methylation without isolation of the pure auine; for effective- ness it is sufficient to have 2 to 10 mg. of the alkaloid. The picrate of the base is methylated quantitatively by heating with formaldehyde and formic acid. The presence of nornicotine by this procedure was shown in various members of the tobacco family. Determination of the picrate melting point both before and after methylation enabled the nornicotine to be identified in the presence of nicotine. Anabasine and piperidine nr^ also methylated by this procedure.— Shmuk (64). In order to find a more logical basis for a classification of tobaccos, the melting points' of picrates prepared from known mixtures of nicotine and nornicotine were determined. The steam- volatile alkaloids were separated from the plant material by dis- tillation and the melting point of the mixed picrates of the alka- loids was oompared with that of the known-mixture picrates. In the curve prepared from the known -mixture-pi crate melting points and composition the slope of the right-hand side was sufficient to warrant establishment of a division at composition two-thirds nicotine and one-third norniootine, and temperature 211° C. Hence, tobaccos, the alkaloids of which give a picrate with the upper limit of the melting range above this point, are considered as the "nicotine type. 1 * The division point at composition one-third nicotine and two-thirds nornicotine, and temperature 196° C, falls in a region of merked discontinuity of melting-point be- havior and is therefore even more suitable to set off the "nornicotine type" of tobacco. This leaves the interval 196° to 211° C. as characteristic of "mixed type" of tobacco. Tobaocos were analytod for nicotine and nornicotine, and the analysis and the melting point of the picrate of the steam-volatile alkaloids from the sample were oompared with the oomposition-melting point curve. The close agreement of the relative percentage compositions and melting points obtained from the tobacco samples with t..ose of known solutions has been used with considerable success in extimat- ing the relative composition of these alkaloids in samples. — Rowen and Barthel (15). - 6 - Nornicotine may "be identified in tobacco, insecticidal tobacco preparations, and nicotine preparations by comparing the melting point of the mixed picrates of the steam-volatile alkaloids with the picrate melting point of a methylated sample thereof. Methylation of the nor- nicotine gives nicotine; consequently, the picrate of the methylated alkaloids will melt at the same point as nicotine picrate and no de- pression of melting will occur in a mixed-melting-point determination with nicotine picrate in those cases where steam-volatile alkaloids other than nicotine and nornicotine are substantially absent. — Bowen and Barthel (1_6) . When nightshade ( Solanum nigrum) , Datura stramonium , and tomato plants, usually free of nicotine, were grafted separately on tobacco ( Nicotiana tabacum and N.rustica) , they were enabled to form this alkaloid and store it in the leaves and fruit. Large quantities of nicotine will form in the scions, if the leaves are removed from the tobacco stock at the time of intensive growth of the scions and the root of tobacco is present. V/hen tobacco is used as the scion in graft upon the same plants, nicotine disappears entirely from the graft system and cannot be detected in the stock or scion. When N. glauca , which contains anabasine, was grafted on a tomato plant, as much anabasine was formed by the scion as occurred in the control plant of N. glauca raised under normal conditions. — Shmuk, Smirnov, and Il'in (JOT OCCURENCE OF NORNICOTINE IN COMMERCIAL NICOTINE SULFATE Two commercial samples of nicotine sulfate were found to contain nornicotine, in one amounting to nearly 12 percent of the alkaloid content. This finding is important because nornicotine has been re- ported to be more toxic than nicotine to aohids. Commercial nicotine preparations containing nornicotine may be expected to be better con- tact insecticides than those of pure nicotine. Since nornicotine is known to be less volatile than nicotine, when a mixture of the two is used as a fumigant, the predominant effect is believed to be that of nicotine. The presence of nornicotine in materials considered to be pure nicotine products could easily account for erratic and nonre- producible entomological results. — Bowen and Barthel (12) . EXTRACTION OF ANABASINE FROM PLANTS Steam distillation under pressure can be used to remove anab- asine from plant material. — Sokol&v (J8) . Of the water- immiscible solvents for the extraction of anab- asine, ethylene dichloride is the best for yield and for velocity of extraction. Anabasine combines rapidly with the impurities in tech- nical ethylene dichloride only during the initial stage of standing, after which anabasine is not decomposed and is only partially trans- ferred to the residue with the slowly separating products of the reaction. The optimum ratio of solvent to raw material is 2:1. A semi-production-scale apparatus and experiments are described. — Sokolov and Trupp (81) . - 7 - The diffusion process of producing anabasine by the water- kerosene method is described* Proteins, which are especially harmful in the further treatment of the alkploids, ere decreased shnrply by the presence of sodium chloride. The maximum yield of the alkaloids was 98 percent* In the countercurrent-diffusion process the average period of extraction depends on the temper ature of the water; it could be reduced to 20 to 30 minutes.— Sokolov and Demonterik (79). A report on the production of anabasine sulfate from Anabasis aphyll a at Tohiakent, U.S.S.R., has been published.— Iljin tSOTT" COMPOSITION OF COMMERCIAL ANABASINE SULFATE MADE FROM ANABASIS Commercial anabasine sulfate solution is reported to contain 21*52 peroent of anabasine, 7.62 percent of lupine, 10*45 percent of aphyllidine and aphylline, 18*19 peroent of total sulfates, 1.68 peroent of free sulfuric acid, and 1*24 peroent of inorganic im- purities.— Dashkevioh (19). An improved method for the isolation of lupinine from commercial anabasine sulfate solution was devised* The crude alkaloid mixture was separated by the Orekhov method (ref* 98 in E-537) and distilled in vacuo . The low-boiling fraction 136-9 °"Tp = 12 mm.) consists of lupinine and anabasine. The mixture is dissolved in dry toluene and treated with metallic solium with stirring and heating. On oooling the sodium lupinate is filtered off, washed with dry toluene, and treated with water, the mixture is extracted with petroleum ether, and the «-rtraot dried and conoentrated to yield crystalline lupinine (97 percent recovery). The mother liquor after distillation yields anabasine* The use of petroleum ether for washing sodium lupinate also appears to give a better product than toluene. The best resuT- 1 -- were obtained when petroleum ether was used as the medium for th^ reaction with met* 1 Mo sodium*— Sadykov and Lazur'evakii (61). In 1935 it was reported that Russia produced 2,500 tons of insecticides (powders, soaps, and solutions) containing anabasine or anabasine sulfate*— Bocharova (8) • NALYTICAL METHODS A method for the micro chemical detection of anabasine in legal ohemioal investigations has been desoribed. Whan Dragen- dorf *s reagent is added to solutions of anabasine, oharaoteristio erystals are obtained which may be used for the micro ohemioal de- tection of the alkaloid in material from the cadaver. In this way as little as 1 microgram of anabasine in a drop of solution oan be detected. The anabasine is extracted from the organs by repeated digestion of the ground material with aloohol to which tartaric aoid has been added in amounts sufficient to give an acid reaction to litmus* The extract is oonoentrated in vacuum, the protein re- moved, the aloohol evaporated, and the material subjected to pre- liminary purification with ohloroform. The tartrate is then con- verted into the base by treatment with ammonia, and the base is ex- tracted with ohloroform. After removal of the chloroform the an- abasine oan be tested for directly, or after further purification of the solution.— Shvaikova (71). - 8 - A rapid method is proposed for determining total alkaloids in semimanufactured produots and waste materials in the production of anabasine* The alkaline extract is extracted with ethylene di chloride and the ethylene di chloride extract titrated with hydrochloric acid (litmus indicator)* For determination in Anabasis the raw material is extracted with water, a part of the water extract is made alkaline, and the analysis conducted as above*— SokoloT (7fi)* Separation of anabasine from nicotine and other alkaloids A titration method for determining the total alkaloid content of a mixture is proposed which is based on the fact that the acetyl derivative of anabasine does not form a pi orate and niootine does*— Khmura (53)* Anabasine can be separated from the alkaloids associated with it in Anabasis aphyll a by precipitation as f luosilicate from a solu- tion in ethyl or methyl alcohol* The fluosilicate, CjoHi4N2H2SiF6 • H2O, m*p* 239°(dec*), can be converted to the free base by reaction with sodium hydroxide* Anabasine is precipitated with alcoholic fluosilicio acid from an alcoholic medium* The precipitate is dissolved in hot water and titrated with a 0*1 N sodium hydroxide solution (thymolphthalein indicator)* Nicotine can be determined similarly* Nicotine can be distinguished from anabasine by the solubility of its fluosilicate in methyl aleoholj fluosilicates of both are insoluble in ethyl alcohol*— Sokolov ( 75 , 77)* A comparative study was made of the reactions of anabasine, nioo» tine, and oaniine* The Meltser reagent (carbon disulfide, ethyl alcohol, and dilute copper sulfate solution) causes a white turbidity with nicotine and a black-brown turbidity with anabasine and ooniine* All three alkaloids form characteristic crystals In saturated aqueous solution of piorio acid* An ether solution of anabasine or niootine, when mixed with an ether solution of iodine, first becomes turbid and then crystallises* Coniine does not undergo this reaction* Hecke reagent (selenic acid and concentrated sulfurio acid) colors the anabasine residue pink, but does not change the oolor of nico- tine or ooniine with hydrochloric aoid (sp* gr* 1*19)* The coniine residue forms beautiful violet crystals after evaporation* No such orystals are formed from anabasine or nicotine* With nltrio aoid (sp* gr* 1*4) the niootine residue assumes an Slow heating a pink oolor, which changes to green* Under the same conditions no coloration of anabasine and ooniine is observed* A maximum yield up to 95 percent of anabasine alkaloids is obtained by extraction of cadaver material with acidified alcohol*— Ratvadovskii (65). The following directions for determining niootine in the presence of anabasine are givent Treat the aqueous solution of the alkaloids with 3 ml* of 10-percent sulfurio aoid and 10 ml* of freshly pre* pared 5-peroent sodium nitrite, heat for 30 minutes at 40-60* # neutralise approximately by introducing 10-percent sodium hydroxide and then aoourately by titration with 0*1 N sulfurio aoid (with methyl orange), and add 3 to 4 drops of exoess aoid* Precipitate the picrates by addition of piorio aoid (12 gm* per liter) and cool for 4 hours* Filter, wash twice with 0.1-peroent piorio aoid and twice with water, transfer to a glass-stoppered flask, add 16 to 20 - 9 - ml. of water, and titrate with a 0*1 N alkali (phenolphthalein in- dicator), adding 5 ml* of toluene toward the end of titration* The amount of 0*1 N sodium hydroxide used aultiplied by 0,0081 gives the oontent of nicotine. The method is based on the fact that nitrous acid, without affecting the nicotine, transforms the anabasine into the nitroso compound, which is not precipitated by picrio acid under the above conditions.-- Shmuk and Borozdina (65). Nicotine and anabasine, treated with an aqueous solution of aniline and a 5-peroent solution of ammonium thiocyanate or potassium thiooyanate in bromine water, give a yellow color, whioh remains constant after the addition of 0.5 to 3.0 cc. of 5 to 10 percent sodium carbonate for nicotine, and ohanges to a bright rose for anabasiae. The colored solutions can be compared in the colorimeter with corresponding standard solutions. However, the determination of anabasine and nicotine by the coloriraetric method in plants was complicated by the other compounds containing the pyridine ring.— Shmuk and Borozdina (66). To identify the alkaloids of to v acco the picrates of nornico- tine and anabasine can be methylated quantitatively by her ting with formaldehyde-formia aoid. Picrio acid does not methyl ate under the conditions used. Determination of the melting point of the piorate before and after methylation enabled nornicotine to be identified in the presence of nicotine.— Shmuk (64 ). Nicotine forms an azeotropio mixture with water in a concentra- tion of 2.5 gnu per 100 ml. This property is used in separating it from nomiootine or anabasine, or both. The separation of nicotine from a mixture of the three alkaloids was 97 to 103 peroent of theory when the procedure consisted in distilling the mixed alkaloids from 126 ml* of water solution through the Widmer oolumn to a low volume (about 15 ml.), adding 60 ml. of water oontaining 2 gnu of sodium ohloride, and continuing again to a volume of 15 ml. The combined distillate was titrated and calculated as nicotine, but represents all the nicotine with some of the accompanying alkaloid or alkaloids. The titrated distillate was made alkaline with a slight excess of standard alkali to neutralize the standard acid, and the dis- tillations were repeated as before but without the addition of sodium chloride. The oombined distillate was titrated and the niootine calculated.-- C. K. Smith (73). The determination of niootine and nornicotine in the presenoe of each other in aqueous solution is described. One portion of the solution is treated with sodium nitrite and acetic acid. The nitroso- nornicotine formed is not volatile in steam from a solution buffered at pH 10. The unchanged niootine is steam-distilled off, and determined by acidimetrio titration or "By precipitation with siliootungstic aoid. To another portion formaldehyde and formic aoid are added to methylate the nomiootine to niootine. The total alkaloid, now niootine, is distilled off after addition of excess sodium hydroxide and determined as before. The difference between the two values represents the nomiootine. Details were developed -for attaining an accuracy of 97 to 98 percent.— Markwood (46). - 10 - An improved apparatus for the steam distillation of niootine, nornicotine, and anabasine has been designed* This apparatus per* nits a longer contact between the steam vapor and the solution con- taining the alkaloids, -which results in more rapid distillation than is possible by the conventional (A*0*A*C«) method*— Bowen and Barthel (11). A method is proposed for analysis of nicotine and nornicotine* Sodium chloride and strong sodium hydroxide greatly aid the distilla- tion of nornicotine* The steam distillate containing both alkaloids is concentrated, and an aliquant treated with nitrous add to form nitrosonornicotine, which is not volatile with steam under the con- ditions at which the nicotine may be distilled (alkaline to phenol- phthalein)* Precipitation of the silicotungstates of the distillate containing the nicotine and of another aliquant containing both alka- loids furnishes the means of calculating the percentages of these alka- loids* The results of analyses of several samples of tobacco* N* rustics , and Tranter cial nicotine sulfate solutions lndioate an unexpeotedTy wide- spread occurrence of nornicotine* From a comparison of analyses made aeoording to the A. 0* A* C* method* it is evident that when nornicotine is present the accepted method of analysis for nicotine cannot be de- pended upon to give reliable results.— Bowen and Barthel (IS)* A test for the completeness of extraction of alkaloids from plant material is described which employs a slight modification of the method employed by Markwood and Barthel (see 46) for the liberation of the alkaloid from plant material* Approximately 5 gnu of the metre is treated with 10 ml* of strong sulfuric aoid (9 ♦ 1), whereby the cellular structure is broken down with the formation of the sulfates of any remaining organic bases* After mixing for several minutes, water is added and the undissolved portion removed by filtration* The filtrate is neutralised with sodium hydroxide just aoidifedwith hydroohlorie aoid and tested witk siliootungstic acid solution* The test solution and reagent art warmed and then cooled with strong agitation. No precipitate in- dicates less than 0*01 peroent of nicotine, nornicotine, or anabasine — Bowen and Barthel (14)* A method of determining nicotine and nornicotine in mixtures of *■' the alkaloids without preliminary separation is described* The method depends on the reactions of these oompounds with cyanogen bromide to give, with niootine, a pale yellowish* green color, and with nornicotine a red color* A 0.64 to 2-gm. sample of tobacco is steam- distilled from a solution containing salt and sodium hydroxide* This treatment insures a quantitative recovery of norniootine, which is much less volatile with steam than is niootine* A volume of the dis- tillate containing not more than 80 micrograms of nicotine or 160 micrograms of nornicotine is treated with potassium dihydrogen phosphate, and then with freshly prepared cyanogen bromide reagent. After the color has been allowed to develop, the solution is trans- ferred to a spectrophotometer and the intensity of the color de- veloped by norniootine is read at 540 millimicrons and that by nicotine at 575 millimicrons. Concentrations of the alkaloids are determined by inspeotion of previously prepared calibration curves. Since the color developed by nornicotine has an additive effect on the intensity of the color given by niootine, the true concentration of - 11 - nicotine is equal to its apparent concentration minus the concentra- tion of nornicotine. Factors influencing the development of the colors are discussed. — Larson and Haag (37) « PHARMACOLOGY OF ANABASINE AND NORNICOTINE Anabasine Farmers in the Union of South Africa "believed Nicotiana glauca to be deadly to ostriches, and it. had been reported to be poisonous to cattle and sheep. In tests on rabbits the flowers were found to be slightly less toxic than the leaves. Rabbits died within 1.5 hours after being fed 30 gm. °f the fresh flowers and within half an hour of eating the same dosage of fresh leaves. A sheep receiving 300 gm. of the fresh leaves through a stomach tube died within 2.5 hours.— Steyn (SU) . Anabasine solutions are fatal to animals- when injected under the skin. Solutions of the equivalent compound anabasine-proto acid (obtained from soybeans and pea seeds) under the stme conditions caused only slight poisoning. The rule of Traube stating the relation between the surface tension of a solution and its poisonous properties cannot oe applied to solutions of anabasine-proto acid, since in spite of low surface tension such solutions do not kill experimental animals. Leont'ev and Smirnova (39) • Alpha-aminoanabasine possesses the fundamental pharmacological properties of anabasine but is less active. Its depressing action on the sympathetic ganglion of white mice is l/75, its stimulating action on the cerebral layer and the kidney 1 /75» its stimulating action on respiration l/20 to 1/100, and its toxic action l/l6. The primary phase of the depression of respiration is absent; action on the blood pressure of decerebrate cats is less, and production of convulsions in white mice is greater, than with anabasine. — Poluektov The introduction of acetyl, propionyl, and benzoyl groups in alpha- and alpha' -amino-N-methylanabasine decreases their respiratory stimulation but does not lower their toxicity. — Kabachnik and Zltser (}2) . The intravenous injection into rabbits of 2.5 to 6 mg. of ana- basine sulfate per kilogram of body weight followed by repeated in- jections of 0.5 to 1.5 mg. per kilogram caused a decrease in the velocity of blood circulation in the lesser circulation system. The decrease was due to the action of anabasine sulfate in contraction of the blood vessels and the increased emission of adrenaline. — Pleshchitser (^2) . Nornicotine On mice and dogs the toxic and vasopressor effects of smoke from a tobacco containing 0.13 percent of nicotine and 0.27 percent of nornicotine were due to the nicotine present, and relatively in- dependent of the nornicotine content of the tobacco. On man the smoke from these low-nicotine cigarettes produced much less pro- nounced 091, Nov. 30. (19) -— 1939** Tne chemical properties of commercial anabasine sulfate. Trudy Leningrad. Inst. Sovet. Torgovli 1939 (2): 8-13. [Abstracts in Ehim. Ref. Zhur. 4 (2): 71. 1941j Chea. Abe. 37* 3228, 1943.] (20) — — 1939^. The chloro- and bromo- substituted anabasine alkaloids and their structure. Trudy Leningrad. Inst. Sovet. Torgovli 1939 (2): 14-21. [Abstracts in Khta. Ref. Zhur. 4 (2)t 47. 1941; Chea. Abs. 37: 3095. 1943.1 (21) Brmoloev, M. F. 1940. The biology of Thrlps [linl Lad.] llnarlus Uz. and control meaeures against it. [Leningrad] Inst. Zashch Raet. (Lenin Acad. Agr. Scl., U. S. S. R. , Inst. Plant Pro- tect.) Trudy po Zashch. Raet. (Bui. Plant Protect.) 3: 23-24 t illus. [In Russian. Abstract in Rev. Appl. Unt. (A) 30: 234- 235. 19*2.] (22) Ivtuehenko, G. A. 19^0. The most important results of the inveetlgations of the phyeleloglcal section of the Tobacco and Machorka Institute (WITtM). Tabakwelt 10 (6): 32-41. [in Russian. Abstracts in Chem. Zentbl. 112 (1): 591. 1941; Chea. Abe. 36: 7066. 1942.] (23) Freak, G, A. 1942. Insecticides. Chem. and Indue. 6li 429-431. (24) Oalakhov, P. N. 194l. Ooabating the summer cabbage fly. Sady - IS - (25) Goodhue, L. D., and Sullivan, W. N. 19*4-5. Method of allying parasiticides. U. S. Patent 2,321,023, June 8. To Secre- tary of Agriculture. (26) Haag, H. B., and Larson, P. S. 191*3, Some chemical and phar- macological observations on 'low nicotine" tobacco. Science 97: 187-188. 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