1310 PERKIN AND HORSFALL: GENISTEIN. PART 11. CXXV. -Genistein. Part 11. By ARTHUR GEORGE PERKIN, F.R.S.E., and LOUIS HUBERT HORSFALL. IT was shown (Trans., 1899, ’75, 830) that dyer’s broom (Genista tinctoria) contains, in addition t o luteolin, a new substance genistein, which possesses feeble dyeing properties. When decomposed, the latter gave phloroglucinol and an acid resembling p-hydroxyphenylacetic acid, and this reaction, together with its other properties, indicated i t t o be a trihydroxyphenylketocumaran. 0 Further investigation, however, was desirable in order t o establish the identity of the acid and the nature of the products of methylation. These points could not be dealt with previously owing to the small quantity of genistein which the plant contains and the laborious nature of the operations for its extraction and complete purification. For the apparently meagre results of this paper, 25 grams of genistein were employed, the preparation of which extended throughout five months.Methylation products of Genistein. It has been shown (Zoc. cit.) t h a t by methylation genistein yields t w o dimethyl ethers, (a) the main product melting at 13’7--139O, and ( b ) present in minute quantity and characterised by its sparing solubility in alcohol. This substance, to which the formula C1,H80,(O*CH,), was assigned, yielded a monoacetyl compound, C,,H70,(0*CH,),* C,H,O, and that the latter had t h e composition assigned to i t was evident from a study of its decomposition with an acid. The ether (a) was first examined.PERKIN AND HORSFALL: GENISTEIN.PART 11. 1311 0.6710 dissolved in 15 C.C. of boiling acetic acid and treated with 2 C.C. of sulphuric acid gave 0,5815 C,,H,,O,. Found 86.66 per cent, Calculated S7-19 per cent. Decomposition w i t h AZcoholic Potaslb.-The dimethyl ether ( a ) was digested with strong alcoholic potash a t 150-160O for 3 hours. The resulting solution was evaporated to dryness, the residue dissolved in water, neutralised with acid, treated with excess of sodium hydro- gen carbonate, and extracted with ether. On evaporation, a small quantity of colourless residue remained which did not crystallise on long standing, and was similar in character to that yielded by luteolin trimethyl ether (this vol., p. 1317) by this method. To identify it, the residue mas dissolved in dilute sodium carbonate solution, diazo- benzene sulphate added, and the red precipitate thus produced collected, washed, dried, and purified by extraction with alcohol and crystal- lisation from a mixture of alcohol and acetic acid, It formed a glisten- ing mass of orange-red needles melting a t 251-252', did not dissolve in cold aqueous alkalis, and was identical with disaxobenxenephloi.o- glucinol mononzethyl ether, previously prepared from the methyl ethers of quercetin (Proc., 1900,16, 1Sl) and luteolin.The sodium hydrogen carbonate solution from which this phenol had been removed mas neutralised with acid, extracted with ether, and the extract evapor- ated. The resulting oily residue gradually solidified and mas then digested with boiling light petroleum, filtered, and the crystals which had separated overnight recrystallised from the same solvent with the aid of animal charcoal.0.1088 gave 0.2594 CO, and 0.0595 H,O. 0.1365 ,, 0,1940 AgI. CH,=9*07. C = 65.02 ; I1 = 6.07. C,H,,O, requires C = 65.06 ; H = 6-02 ; CH, = 9.03 per cent. This acid consisted of colourless leaflets melting a t 85-S6', and was identical with the methoxyphenylacetic acid, CH,*O*C,H,* CH; CO,H, prepared by Cannizzaro (Annulen, 1861, 117,243). The decomposition products of genistein dimethyl ether ( a ) are thus phloroglucinol mono- methyl et?Ler and p-methoxyphenylacetic m i d , consequently this com- pound is the nornzd dimethyl ether of the colouring matter. The sparingly soluble Dinzethyl Ethw @).-The yield of this product was exceedingly small and little more than one gram was available for investigation.The melting point, previously given as 1 87-189", is foo lorn, and should be 200-202°. I t has been shown (loc. cit.) that this substance contains two methoxy-groups, and that by the action of hydriodic acid genistein or a very similar compound is formed. To account for its production, it seemed possible that during methylation a methyl group had entered the ring, the product being consequently a dimethyl ether of methylgeiiistein (loc. cit.). 4 u 21312 PERKIN AND HORSFALL : (XENISTEIN. PART 11. The acetyl derivative prepared in the usual manner forms colourless It needles, dissolves sparingly in alcohol, and melts a t 212-214O. contains only one acetyl group. 0.6548 gave 0.5768 regenerated ether.Found 88.08. CI4H70,(0*CH3),* C,H,O requires Cl6HI4O5 = 87.1 9 per cent. CH,* G14H603(0*CH3)2* C2H30 ,, C&,,O, = 87.72 ,, The regenerated dimethyl ether melting at 200-202° was 0.1148 gave 0.2886 GO, and 0.0555 H,O. By the action of alcoholic potash at 150-160°, this ether yielded a n acid which crystallised in colourless leaflets, melted at 85-86', and was identical with p-methoxyphenyhcetic acid. The phenolic pro- duct of the decomposition became crystalline on standing, and was identical in appearance with that yielded by the analogous trimethyl ether of high melting point from luteolin (Zoc. cit.). That it was the same substance was confirmed by the preparation of its disazobenzene derivative, for this formed orange-red needles melting at 198-2019 As previously indicated, this is most probably disaxo6enxenernethdyZ- phloroglucinol monomethyl ether, and consequently the ether obtained from genistein melting at 200-202° is considered to be rnethylgenistein dimethyl ether.anal y sed. C=68.56 ; H=5*37. C17H,,0, requires C = 68.00 ; H = 5.33 per cent, The following would appear t o be its constitution : 0 To determine whether any characteristic distinction could be noted between genistein and the presumed methylgenistein, a trace of each was prepared by the decomposition of the respective methyl ethers with hydriodic acid. I n appearance and general reactiocs, both pro- ducts appeared to be identical, but a distinction in the melting point was'noted, as the genistein melted at 287-289',* and tho methylgenistein at 276-27S0.From these results, it is unlikely that methylgenistein or its glucoside exists in the Genista tinctoria, and the substance is almost certainly produced in the form of its ether during methylation. Owing to lack of material, experiments could not be made t o determine if it could be formed by digesting genistein dimethyl ether with methyl alcoholic potash and methyl * A sample of genistein from the dyer's broom, the purity of which could not be doubted, melted a t 291-293", slightly higher than that from the ether, a fact readily explained, as the quantity of the latter did not admit of recrystallisation.PERKIN AND HORSFALL: GENTSTEIN. PART 11. 1313 iodide, but by acting a t first with a somewhat larger quantity of the alkali and iodide upon genistein, it was observed that the yield of tbe methylgenistein compound was somewhat increased.Owing to the difficulty in preparing it, we do not propose to examine it fur- ther, for i t s analogy to the rnethylluteolin ether is evident. Genistein Diethyl Ether. Genistein dissolved in a solution of potassium hydroxide (5 mols.) in ethyl alcohol was digested a t the boiling point with excess of ethyl iodide for 3 days. From the resulting solution, which on cooling formed a semi-opaque, gelatinous mass, unaltered ethyl iodide was removed by distillation; the alcoholic liquid was then poured into much ether, and the product washed with water and evaporated to dryness. The amorphous, horn-like mass was dissolved in boiling alcohol, allowed to partially cool, and a small quantity of an indefinitely crystalline substance ( c ) which separated, was rapidly removed by filtration through calico.This operation had for its ob- ject the removal of a less soluble ether if such should be present, as is the case with the methylation product. The filtrate became gelatin- ous on cooling, and as other solvents did not yield a crystalline pro- duct, it was evaporated to dryness and acetplated in the hope that the resulting compound would be easier to manipulate. This was found to be the case, for the nzonoacetyl derivative, Cl,H70,(0*C,H,),*C,H30, crystallised in colourless needles melting at 168-1 70". 0.1120 gave 0.2780 CO, and 0.0575 H,O. C,OH,oO, requires C = 67.41 ; H = 5.62 ; CI8Hl8O5 = 88.20 per cent. The genistein diethpl ether regenerated from the above substance could now be crystallised from alcohol, in which it is sparingly soluble, and was thus obtained in colourless needles melting a t 132-134'.The mother liquor should be removed while still warm, as the later separations are deposited in the gelatinous condition. C=67*68; H=5*70. 0.4165 ,, 0.3670 C18H180,. C,,H1,05 = 88.11. 0.1087 gave 0.2752 CO, and 0.0573 H,O. C=69*04 ; H=5.85. C18Hl,0, requires C = 68-79 ; H = 5.73 ; C,H, = 18.47 per cent. The ether was decomposed with alcoholic potash and the products separated in the same manner as those yielded by the dimethpl ether. The acid thus obtained formed colourless leaflets melted at 88", and had the properties of p-ethoxyphenylacetic acid, C,H,*O*C,H,*CH,*CO,H. C,oH120, requires C = 66-66 ; H = 6.66 per cent.0.0645 ,, 0.1005 AgI. C,H, = 19-22, 0.1051 gave 0.2546 CO, and 0.0593 H,O. C = 66.06 ; H = 6-27,1314 PERKIN AND HORSFALL: LUTEOLIN. PART 111. The accompanying phenol is most probably phloroglucinol mono- ethyl ether, but owing to lack of data this cannot yet be identified. When treated with disazobenzene sulphate in the usual manner, an orange-red compound resulted, which crystallised from alcohol in needles, and was insoluble in cold aqueous alkalis. The product (c) obtained during the purification of the above substance melted a t 127--129", and did not appear to contain a second ethyl ether, although, as a mere trace was available, it could not be properly examined. Experiments were carried out in the hope of preparing a pure azo- benzene derivative of genistein, but these, as well as those for 'the production of nitro-derivatives, were unsuccessful ; they indicated, however, that if sufficient material were available for the determin- ation of the necessary conditions, such derivatives could be produced, a s has been the case with the somewhat closely allied colouring matter apigenin.Szcmrnarg of Results. When methylated, genistein, like luteolin (Zoc. cit.), gives two ethers which are insoluble in aqueous alkalis, and capable of forming monoacetyl derivatives. That melting at 137-139" is the normal dimethoxy-compound, whereas the second, which melts a t 200-202", is considered to be methylgenistein dimethyl ether. It is probable that the formation of the latter is due t o a methyl group entering the ring of the phloroglucinol nucleus during methylation, for there is no evidence to show that methylgenistein (or its glucoside) is contained in Genistcl t i n c t o r i a . The production of p-methoxy- and p-ethoxy-phenylacetic acids from the corresponding ethers of gen istein proves that the acid previously obtained from this colouring matter is p-hydroxyphenylacetic acid; and the results, as a whole, are in har- mony with the constitution which was previously assigned to genistein. CLOTHWORKERS' RESEARCH LABORATORY, DYEING DEPARTMENT, YORKSHIRE COLLEGE,