LV..-Phgtin and Phytic Acid.By GEORGE CLARKE.THE name phytin has been applied to a white, amorphous substancewhich has been observed by Palladin (Zeitsch. Biol., 1895, 31, 199)to occur in the seeds of many plants, and yields inositol and phos-phoric acid on hydrolysis under pressure with solutions of mineralacids or alkalis. An examination of the methods used in preparingthe material investigated by previous workers left no doubt thatdifferent substances have been described as phytin, and that thesesubstances were not always homogeneous. This conclusion was con-firmed by the divergent published results representing the composi-tion of phytin (compare Jegorov, Biochem. Zeitsch., 1912, 42, 433,and Plimmer and Page, Biochem. J., 1913, 7, 158).The acid obtained from phytin by the removal of the bases,calcium and magnesium, has been described as phytic acid, but nosalt or derivative of it of undoubted purity has been isolated andanalysed.Many of the substances described as phytic acid havebeen mixtures of phosphoric acid and an organic phosphoric acid.Schulze and Winterstein (Zeitsch. physiol. Chem., 1896, 22, 90 ;1903, 40, 121) prepared phytin by extracting the faf-free seeds ofSinap-s nigra with 10 per cent. sodium chloride solution, coagulatingthe proteins by boiling, and, after cooling and filtering, precipitatingphytin from the cold protein-free sodium chloride solution by heat-ing. The substance thus obtained was identical in properties withthat prepared by the methods recorded below.These authors men-tioned the fact that phytin was less soluble in hot than in coldacetic acid, but did not develop this method of preparation.Winterstein (Ber., 1897, 30, 2299) prepared phytin by extractingseeds with dilute acetic acid and precipitating it from the solutionby ammonia. The free acid was obtained by the removal of thebases by means of oxalic acid, and yielded inositol and phosphoricacid on hydrolysis under pressure with concentrated hydrochloricacid. The substances prepared by the latter methods were un-doubtedly mixed with calcium and magnaiufn phosphates andphosphoric acid.Posternak (Compt. rend., 1903, 137, 202, 338, 439) extractedphytin from various fat-free seeds by means of very dilute hydro-chloric acid, separating the substance by precipitation of the coppersalt, decomposing the latter by hydrogen sulphide, and treatingwith alcohol the Syrupy acid substance obtained by the evaporationof the acid solution.The final product was soluble in water, andthus differed from that obtained by Schulze and Winterstein (Zoc536 CLARKE: PHYTIN AND PHYTIC ACID.cit.) and the present author, which was insoluble. Posternak statedthat the substance prepared by him was free from nitrogen andinorganic phosphates.Treatment with alcohol of the syrupy acid residue obtained bythe latter worker would separate a large amount of inorganic phos-phoric acid from the final product, but an examination of the freeacid liberated from a product obtained by analogous methods, bycarefully fractionating the strychnine salt, showed that i t containedinorganic phosphoric acid. In this connexion it is of interest tonote that Page and Plimmer (Biochem.J., 1913,7, 168) have foundthat samples of commercial phytin always contained inorganic phos-phoric acid.Free phytic acid, obtained by Posternak, was described as a paleyellow, transparent syrup, yielding inosito-1 and phosphoric acid onhydrolysis, and giving no precipitate in the cold, but a characteristicyellow precipitate on warming with ammonium molybdate solution.He ascribed to the acid the formula C,H,O,P,, and the constitutionrepresented by anhydro-oxymethylenediphosphoric acid,[ CH,~O*PO(OH),],O.One of the arguments on which this formula was based was the factthat phytin and phytic acid were not hydrolysed by alkalis, a state-ment that Winterstein (Zeitsch.physiol. Chem., 1908, 58, 121) hassince shown to be incorrect. Neuberg (Biochem. Zeitsch., 1908, 9,557) has brought forward additional evidence to show that theinmito1 ring formation exists in phytin and phytic acid.The methods used in the preparation of the material, on theanalysis of which Posternak based the, formula C2Hs09Pz, did notpreclude the possibility of admixture with phosphoric acid.The phytin previously examined had been prepared by one ofthe methods described, or some slight modification of them.Starkenstein (Biochem. Zeitsch., 1911, 30, 59) has stated thatair-dried commercial phytin contains a considerable quantity ofinorganic phosphoric acid, and that the amount is increased bydrying a t looo.He attributed this to the decomposition of phytininto inositol and phosphoric acid a t that temperature, but did notrecord the isolation of inositol. Anderson (J. Biol. Chem., 1912,11, 473) failed to confirm his conclusions that phytin was so easilydecomposed.The results of experiments recorded in this communication haveshown that the free acid liberated from air-dried phytin of homo-geneous composition-which was separated from admixed mineralPhosphates by precipitation from cold dilute acetic acid by boiling-consisted of a mixture of approximately equal quantities of anorganic phosphoric acid (phytic acid) and phosphoric acidCLARKE: PHYTIN AND PHYTIC ACID. 537A solution of the ammonium salt of the organic phosphoric acid,prepared from a pure strychnine salt, gave no precipitate on warm-ing to 60° with a nitric acid solution of ammonium molybdate, andonly a very slight one on remaining a t that temperature for severalhours.An explanation of the behaviour of phytin is that it is not simplya salt of an inositolphosphoric acid, but a complex substance,possibly a complex calcium-magnesium salt of an inositolphosphoricacid and phosphoric acid, and, on removing the bases, yields thetwo acids.The fact that the composition of pure phytin, preparedas described below, corresponded with no calcium-magnesium saltof a simple acid ester of inositol and phosphoric acid, gave supportto this view. The strychnine salt of the organic phosphoric acid,isolated from the mixture of acids obtained from phytin, on theother hand, gave results on analysis in agreement with salts ofsimple inositolphosphoric acids.Anderson (J. Biol.Chern., 1912, 11, 471) prepared from com-mercial phytin obtained from two sources a series of salts, which onanalysis gave figures corresponding with salts of inositolhexaphos-phoric acid, CGH,O,[PO(OH),],. He described an acid tribariumphytate prepared by precipitation from 0.5 per cent. hydrochloricacid solution by the addition of an equal volume of alcohol. Thissalt was probably one of the purest derivatives of phytic acidhitherto isolated, but i t seemed not impossible from the methodsof preparation that the salt as well as the acid prepared from itmight contain some phosphoric acid.EXPERIMENTAL.Preparation of Phytin.The seeds of Indian field mustards, a mixture of Srussica junceu(H.fil. and T.) and Brassica campestris (Linn.), were extractedwith petroleum f o r several days. The petroleum extract wasseparated by means of a centrifugal machine, and the seeds driedin the sun for a few hours. Twenty-seven kilograms of air-dried,fat-free seeds were extracted for seven days with 100 litres of 4.5per cent. acetic acid. The extract was separated from the seeds ascompletely as possible in the centrifuge, boiled for fifteen minutes,and allowed to cool. This procedure coagulated a large portion ofthe proteins. After remaining overnight, the supernatant extractwas easily syphoned off.The dark brown extract thus obtained wasagain heated to boiling. A small quantity of phytin separated out,but the solution was too dilute and impure for any quantity ofmaterial to be obtained in this manner. Aqueous ammonia wa538 CLARKE: PHYTIN AND PHYTIC ACID.then added to the boiling extract until it was just alkaline, and theboiling continued for a few minutes. A large quantity of dark-coloured precipitate (A) was thus obtained, which was separatedby filtration while still hot, and well washed with boiling water. Thecrude precipitate (A) contained phytin, calcium and magnesium-ammonium phosphates, and a considerable quantity of protein andother organic impurity. It was intimately mixed with 16 litres of8 per cent. acetic acid, and extract,ed for two or three hours withconstant shaking.The extract was separated from the undissolvedorganic impurities by filtration through cloth, boiled, allowed tocool overnight, and the cold solution thoroughly stirred. The un-dissolved and precipitated matter was then easily separated byfiltration through paper, and a perfectly clear filtrate obtained.Aqueous ammonia was added t o the boiling filtrate until it wasjust alkaline, the white precipitate (B) being collected on a filterand washed with boiling water until almost free from ammonia.The white precipitate (B) consisted of phytin and calcium and mag-nesium-ammonium phosphates. It was dissolved in 6 litres of8 per cent. acetic acid, and a very small amount of insoluble matterseparated by filtration.The clear solution thus obtained wasboiled for some minutes. Much phytin was precipitated, and i t wasseparated by filtration through a Buchner funnel while the liquidwas still hot, then well washed with boiling water, and finally withethyl alcohol, 82 grams being obtained. The hot filtrate fromwhich the phytin had been separated was again made alkaline withammonia, and the precipitate (C) separated and washed. It wasdissolved in 2 litres of 8 per cent. acetic acid, and phytin separatedby boiling, filtering while still hot, and washing with water andalcohol as described above. Fifteen grams were obtained. Thefiltrate from this was again subjected to similar treatment, a smallervolume of 8 per cent. acetic acid being used (1 litre), and yielded8 grams of phytin.The residual filtrate on treatment with excessof ammonia yielded a further precipitate, which consisted mainlyof calcium and magnesium-ammonium phosphates, and containedonly a very small amount of carbon.The total yield of phytin was 105 grams, or 0.38 per cent. of theair-dried, fat-free seeds.The following yields of phytin were obtained in other prepara-tions :27.0 kilograms of air-dried fat-free seeds gave 125 grams of phytin = 0.46 per cent.*28*8 ) y Y ) > ) s> 165 > > y , = 0.57 ) )31.5 ,) Y Y 9 7 9 ) 152 Y > ,, = 0.47 ,)Seeds extracted with 0-2 per cent. hydrochloric acid.Very dilute hydrochloric acid can be used instead of dilub acetiCLARKE: PHYTIN AND PHYTIC ACID. 539acid for the extraction of the seeds, and some of the material usedin this investigation was prepared by extraction with 0.2 per cent.hydrochloric acid, the phytin being subsequently purified byseparation from 8 per cent.acetic acid. It was found, however,that the extracts obtained by the use of dilute hydrochloric acidwere more difficult to handle than those obtained by dilute aceticacid.Phytin prepared in the manner described above was a snow-white,amorphous powder, resembling in properties the substance describedby Schulze and Winterstein (Zeitsch. physiol. Chem., 1896, 22, 90).It contained carbon, hydrogen, phosphorus, calcium and mag-nesium, but no trace of nitrogen could be detected. It was in-soluble in hot and cold water, readily soluble in very dilute mineralacids, and soluble in cold, but sparingly so in hot dilute acetic acid.It was precipitated from a cold 8 per cent.acetic acid solution onboiling, completely redissolving when allowed to cool.A solution of phytin in very dilute nitric acid gave an abundantyellow precipitate with acid ammonium molybdate solution onwarming to 60°.The following fractions from different preparations were preparedfor analysis :(A) Fifty grams of phytin dissolved in 2000 C.C. of distilledwater and 100 C.C. of glacial acetic acid. The solution obtained wasquite clear and free from undissolved material. It was heated, notquite to boiling. The precipitated phytin was separated by filtra-tion, well washed with boiling water until free from acid, finallywith alcohol, and dried on a porous plate.Phytinwas again precipitated, separated, washed with boiling water, butnot with alcohol, and dried on a porous plate.(C) Seventy grams of phytin from a separate preparation weredissolved in 2500 C.C.of 5 per cent. acetic acid. The clear solutionwas heated for some time by passing steam into it. The precipi-tated phytin was separated, washed with water and alcohol, anddried.(D) Seventy grams of phytin from a preparation made by ex-tracting mustard seeds with 0.2 per cent. hydrochloric acid, andsubsequent purification by precipitation from 8 per cent. aceticacid, were' dissolved in 2500 C.C. of 5 per cent. acetic acid. Phytinwas separated from the perfectly clear solution by heating, washedand dried.It is somewhat difficult to obtain phytin in an anhydrous con-dition.After heating f o r several hours at l l O o in a vacuum overphosphoric oxide, it still continued to lose weight. When heated(B) The filtrate from A was boiled for fifteen minutes540 CLARKE: PHYTIN AND PHYTIC ACID.under similar conditions a t 180° for five hours it became constantin weight, and remained so after prolonged heating for many hours.The anhydrous substance, dried a t 180° and dissolved in diluteacetic acid, was precipitated again unchanged by boiling.Plimmer and Page (Biochem. J., 1913, 7, 162) have referred tothe difficulty of analysing phytin, and have critically examined themethods of determining calcium and magnesium in the presence ofphosphoric acid.They mention also the difficulty of completelyburning carbon in the presence of phosphoric acid.The following methods were used in the analyses of phytinrecorded below :Curb on and hydro yen.-The anhydrous substance was intimatelymixed with the finest powdered copper oxide.Calcium and 2Clagnesium.-The organic matter was oxidised byconcentrated sulphuric acid. After diluting with water, calciumwas separated from the solution as calcium sulphate by the additionof an equal volume of alcohol, and magnesium estimated in thefiltrate as Mg,P,O,, after the removal of alcohol by evaporationand oxidation with nitric acid as described by Plimmer and PagePhosphorus was determined (after the oxidation of the organicmatter by concentrated sulphuric acid) by means of ammoniummolybdate, and weighed as Mg,P,O,.The following results were obtained on analysis of the fractionsdescribed above, after drying in a vacuum over phosphoric oxidea t MOO:(loc.cit.).Fraction A .0.4176 gave 0.1482 CO, and 0.0620 H,O.0.4866 ,, 0.3044 CaSO, and 0.041 Mg,P,O,. Ca=18*39;0-4421 gave 0.3210 Mg,P,O,.C=9*67; H=1*64.Mg = 1-84.P = 20.21.*0*4866 ,, 0.359 Mg,P,O,. P=20*53.Fraction B.0.4650 gave 0.1694 CO, and 0.0698 H,O.0.4592 ,, 0.2812 CaSO, and 0.0458 Mg,P,O,. Ca=18*01;0.2820 gave 0.2072 Mg,P,07.C=9*93; H=1.66.Mg = 2-18.P = 20.45.* Estimated as Mg,P,O,, after separation of CaSO,CLARKE: PHYTIN AND PHYTIC ACID. 541Fractzon C.0.4330 gave 0.1530 CO, and 0.0756 H,O.0.6210 ,, 0.3880 CaSO, and 0'0520 Mg,P,O,.Ca=18*37;0.4504 gave 0.3370 Mg2P,0,.C =9.63 ; H = 1.94.0.4070 ,, 0.1440 CO, ,, 0.0684 H,O. C=9.65; H=1*86.Mg = 1.83.P = 20.82.Fraction D.0.3758 gave 0.1334 CO, and 0.0530 H,O.0.4950 ,, 0.3100 CaSO, and 0.0404 Mg,P,O,. Ca=18*40;0.3680 gave 0.2736 Mg,P,O,.C =9.68; H= 1.56.*0.4774 ,, 0.1626 CO, ,, 0.0680 H,O. C=9.29; H=1.58.Mg = 1.78.P = 20.69.C12H,,04,P,,Ca,Mg requires C= 9.70 ; H = 1.48 ; Ca = 18.87 ;Mg = 1-61 ; P = 20.88.Cl2H2,O4,Pl,Ca7Mg requires C = 9-82 ; H = 1-34 ; Ca = 19.09 ;Mg = 1-63 ; P = 21.14.C12H,,0,,P,,Ca,Mg requires C = 10.06 ; H = 1.12 ; Ca = 19.58 ;Mg=1*6'7; P=21*67 per cent.The composition of separate preparations of phytin, purified byseparation from cold dilute acetic acid, was constant.Phytin is decomposed by heating under pressure with 30 per cent.sulphuric acid into inositol and phosphoric acid (Posternak, Compt.rend., 1903, 137, 439; Winterstein, Ber., 1897, 30, 2299).Fifteen grams of phytin were heated with 67 C.C.of 30 per cent.sulphuric acid in a sealed tube a t 130° for ten hours. The dark-coloured solution was diluted with water, and calcium sulphate,which had separated out, removed by filtration. Excess of sulphuricacid was removed by treatment with finely powdered barium car-bonate. The solution, which contained inositol, was evaporated toa small bulk, removing from time to time the slight deposits ofmineral matter. The concentrated solution was acidified with twodrops of nitric acid, poured into five times its volume of ethylalcohol, and ether added.Inositol separated out as a viscid mass,which quickly solidified. Yield = 3.5 grams.Six grams of inositol from the above and other similar prepara-tions were boiled for two hours with 50 grams of recently distilledacetic anhydride and 0.5 grams of zinc chloride, and the reactionmixture poured into cold water. Hexa-acetylinositol separated, andwas purified by several recrystallisations from ethyl alcohol. Itmelted sharply a t 211O (uncorr.), and was dried a t l l O o for analysis.Anhydrous substance mixed with powdered lead chromate542 CLARKE: PHYTIN AND PHYTIC ACID.(Found, C = 50.0 ; €I = 5.6.per cent.)takes place in accordance with the following equation :C18H24O12 requires C = 50.0 ; H = 5.5The decomposition of phytin into inmitol and phosphoric acidCl,H,,044PloCa7Mg + 8H20 + 8H2S04 =Fifteen grams of phytin gave 3.5 grams of inositol.2C6H,,06 + lOH,PO, + 7CaS04 + MgSO,.C12H22044P10Ca,Mg requires 3.6 granis of inositol.Examination of the Acid from Phytin, Cl,H,,O,,PloCa,Mg.Seventy grams of air-dried phytin were dissolved in 2.5 litres of5 per cent.acetic acid, and basic lead acetate solution was addeduntil no further precipitate was produced. The lead salt wasseparated, washed with boiling water until free from acetic acid,and decomposed by hydrogen sulphide. Excess of the latter wasremoved from the acid solution by boiling under diminishedpressure, and sufficient cold saturated copper acetate solution addedto precipitate the acid.The copper salt was separated, washedwith boiling water, and deprived of copper by means of hydrogensulphide. The strongly acid solution thus obtained was evaporatedto a syrup under diminished pressure, and treated with 500 C.C. of95 per cent. ethyl alcohol. A large quantity of a white, flocculentsubstance (I) separated. This was coagulated by boiling for a fewminutes, filtered from the alcoholic extract, and washed with alittle alcohol.The white substance (I) contained phosphorus, calcium andcarbon, and readily dissolved in cold water. Its aqueous solutionwas precipitated with cold saturated copper acetate solution. Thecopper salt was decomposed by hydrogen sulphide, and the acidsolution evaporated t o a syrup, This syrup was treated with 95 percent.ethyl alcohol (300 c.c.). A smaller amount of a white sub-stance (11) separated, which was filtered from the alcoholic extractafter boiling, and washed with alcohol. Its aqueous solution wasagain precipitated with copper acetate solution, the copper saltseparated, deprived of copper, and the acid solution treated asbefore. This procedure was repeated until the syrupy acid residuewas completely soluble in 95 per cent ethyl alcohol. About fiveoperations were necessary.The alcoholic solutions of the acid obtained by the above opera-tions were mixed together, evaporated under diminished pressure,and finally dried in a vacuum over sulphuric acid.The acid thus obtained wits a viscid, dark-coloured syrup, freefrom calcium and magnesium.It was very readily soluble iCLARKE: PHYTIN AND PHYTIC ACID. 543water or alcohol, and gave a yellow precipitate with acid ammoniummolybdate solution on slightly warming.Twenty-two grams of acid produced from phytin in the mannerdescribed above were dissolved in 3 litres of water, and 50 gramsof recently precipitated strychnine, in as fine a state of division aspossible, added to ths solution. On heating, nearly all the strych-nine dissolved. A small amount of resinous impurity separated,and was removed. The solution of strychnine salts was evaporatedunder diminished pressure to 1 litre, and allowed to remain over-night. Strychnine phytate separated out in small, colourless,needle-shaped crystals, melting a t 203-204O (uncorr.).This saltwas crystallised many times from water, in which it was onlysparingly soluble. The melting point remained unchanged.Strychnine phytate contains water of crystallisation, which it losesvery slowly on exposure t o air, rapidly when dried a t 115O. Itsaqueous solution gave an acid reaction with blue litmus.Separate fractions of strychnine phytate dried a t 115O gave, onanalysis, the following results (the anhydrous salt was intimatelymixed with the finest powdered copper oxide f o r the estimation ofcarbon and hydrogen) :0*2208 gave 0.4776 CO, and 0.1188 H20. C=58.99; H=5.98.0.2407 ,, 0.5198 C02 ,, 0.1284 H20. C=58.89; H=5.92.0.3510 ,, 0.7560 C02 ,, 0.1832 H2O. C=58.74; H=5*79.0.1762 ,, 0.3816 CO, ,, 0.0950 G O .C =59*05 ; H =5*98.0.1888 ,, 0.4092 C02 ,, 0.0970 H2O. C=59*11; H=5.70.0.2750 ,, 0.5930 CO2 ,, 0.1440 H2O. C=58*80; H=5.81.Mean = 58.92 ; = 5.86.0.3936 gave 0.0934 Mg2P207. P = 6-59.0.4896 ,, 0.1158 MgZP207. P=6.58.0.6090 ,, 0.1440 Mg2P207. P = 6.58.0'5354 ,, 0.1300 Mg2P207. P=6.75.0.4574 ,, 0.1138 Mg2P207. P = 6.92.0.3874 ,, 0*0960 Mg2P207. P = 6-89,Nean = 6-72.0.4176, air-dried salt, lost 0.0302 H,O. H20 = 7-23.0.6916 ,, ,, ,, 0.0480 H2O. H20=6*94.1-2360 ,, ,, ,, 0.0846 H20. H20=6.84.Mean = 7-00.The composition of the strychnine salt agreed with the strychninesalts of several inositolphosphoric acids in which strychnine andphosphorus are in the ratio of 1 molecule of strychnine to 1 atoinof phosphorus ;. for example 544 CLARKE: PHYTIN AND PHYTIC ACID.I.C6H802(HP04)2,2C2,H,02N2 requires C = 59.25 ; H = 5-55 ;P=6.37 per cent.C6H802(HP04)2,2C21H,02N,,4H20 requires H20 = 6.90 per cent.11. C,H802(~P04)4,4C2,H2202~2 requires C = 58.87 ; H = 5-66 ;P = 6-75 per cent.C6H,02(H2P04)4,4@21H220a;N2,8H20 requires H20 = 7.27 per cent.111. C",H,(H2P04)6,6C2,H2202N2 requires c = 59-45 ; H = 5.63 ;P= 6.98 per cent.@6H,(~P04)8,6CZ1H2202N2,12H20 requires H20 = 7.50 per cent.I1 and I11 are acid salts. The acids in formulze I and I1 arecapable of yielding a complex calcium magnesium salt with phos-phoric acid of the composition C,2H,0,,Pl,Ca7Mg, which woulddecompose on removal of the bases and liberation of the free acidinto inositolphosphoric acids and phosphoric acid.The solution from which strychnine phytate (m.p. 203-204O)had separated was further evaporated under diminished pressure.After the separation of a small additional quantity of salt, meltinga t 203-204O, the mother liquors, on concentrating to small bulk,deposited a large quantity of a readily soluble salt (m. p. 252-253O).The amount of this salt was approximately equal to the weight ofstrychnine phytate obtained. It was easily separated in a state ofpurity from the sparingly soluble strychnine phytate, and proved,on examination, t o be strychnine dihydrogen phosphate.0.3130 gave 0-0824 Mg2Pz07. P = 7.32.0.2962 ,, 0.0754 Mg2P207. P=7.08.C21H,20zN2,H,P04 requires P = 7.17 per cent.Phosphoric acid was prepared from the strychnine dihydrogenphosphate described above by decomposing an aqueous solution withexcess of sodium carbonate solution, separating the strychnine, pre-cipitating the phosphate as lead phosphate in the presence of diluteacetic acid, and decomposing the lead salt by means of hydrogensulphide. An aqueous solution of the phosphoric acid was heatedwith sufficient benzylamine t o form benzylamine dihydrogen phos-phate, and the latter salt purified by recrystallisation from water.When dried a t l l O o it gave the following results on analysis:0.3030 gave 0.1642 Mg2P20,.C,H,N,H,PO, requires P = 15-12 per cent.Additional proof that the acid liberated from phytin containsmuch phosphoric acid, in addition t o the organic phosphoric acidalready described, was obtained during attempts to prepare theZ-ment h ylamine salt.Nineteen grams of Z-menthylamine were neutralised by a solutionof the mixed acids prepared from phytin, and an equal volume ofthe same solution was added.This strongly acid solution on slowP = 15.08CLARKE: PHYTIN AND PHYTIC ACID. 545evaporation in a vacuum over sulphuric acid deposited, in the formof rhombic prisms, 15 grams of I-menthylamine dihydrogen phos-phate.After purification and drying a t looo it gave, on analysis, thefollowing result :0.2268 gave 0.0984 Mg,P,O,. P = 12.07.0'3334 ,, 0.1442 Mg,P,O,. P = 12.04.C,,H2,N,H,P0, requires P = 12.21 per cent.The solution from which Z-menthylamine dihydrogen phosphatehad been separated, on further evaporation in a vacuum oversulphuric acid, deposited Z-menthylamine phytate in the form of anuncrystallisable oil.Preparation of Inositol from Strychnine Phytate.Nineteen grams of pure air-dried strychnine phytate (m.p.203-204O) were dissolved in the least possible quantity of boilingwater, and decomposed by a slight excess of aqueous ammonia.Strychnine was separated after remaining overnight, and the solu-tion of ammonium phytate evaporated to dryness and dried in adesiccator over sulphuric acid. Ammonium phytate is a non-crystal-lisable gum, very readily soluble in water. In the presence of afew drops of dilute nitric acid it gives no precipitate with acidammonium molybdate solution on warming t o 60°, but if allowed toremain for several hours a t that temperature a very slight yellowprecipitate is formed, owing to the slow hydrolysis of phytic acid.Five grams of dry ammonium phytate were dissolved in 25 C.C. of30 per cent. sulphuric acid and heated in a sealed tube a t 120-130°for eight hours. The dark-coloured solution was diluted withwater and filtered from a small deposit of carbon. Sulphuric acidwas removed by treatment with barium carbonate, and the solutioncontaining inositol evaporated to small bulk, acidified with a fewdrops of nitric acid, and poured into alcohol. Inositol separated asa solid, crystalline mass (1.5 grams). The hexa-acetyl derivative,prepared by boiling with acetic anhydride and a trace of zincchloride, was purified by crystallisation from ethyl alcohol, andmelted sharply a t 211O (uncorr.). It was dried a t l l O o for analysis.(Found, C = 50.27 ; H = 5-47. C,,H,OI2 requires C= 50.00 ; H = 5-55per cent.).The author desires to express his thanks t o his assistant, Mr.S. C. Banerjee, for valuable help in the preparation of the phytinused in this investigation.THE CEEMICAL LABORATORY,UNITED PROVINCES, DEPARTMENT OF AGRICULTURE,INDIA