252 XXIX.-The Poisonous Princ+le of Uyechites Suberecta. By JAMES JOHN BOWREP, Analytical Chemist to the Government of Jamaica. THE plant which has yielded the substances to be described in this paper is very common in Jamaica, occurring sometimes in clumps standing by itself, but far more commonly climbing over fences, or clinging to low growing shrubs. Its dark-green leaves and large bright yellow flowers render it very couspicuous in the hot, dry dis- tricts in which it flourishes. In the island it is called “ Nightshade,” in consequence of its deadly character, but Professor Daniel Oliver has kindly identified it for me as Urechites suberectn, .Muell. Arg. (Echites Neriandm Griseb). My attention was specially directed to this plant by the wonderful tales told of its poisonous powers in the hands of those acquainted with its properties, it being stated that they could so administer it as to cause the very speedy death of the victim, or so as to kill only after the lapse of weeks or months.It is believed to have been used with comparative frequency in the time of slavery, as a means of getting rid of obnoxious masters, and is supposed to be occasionally used at the present time as an inshment of crime. The leaves are intensely hitter and very acid, producing a sensation on the lips and tongue of being swollen and blistered, though their aspect is quite unchanged. The powder of the leaves applied to the nostrils causes violent sneezing, hence it was necessary to take pre- cautions in powdering them, or unpleasant effects followed. These active properties are possessed by all the green parts of the plant and by the flowers, but exist very slightly, if aC all, in the woody portions.My researches have been confined to the leaves and green tips of the plant, from which I have obtained three substances, characterised in a high degree by their physiologically active qualities. These active bodies I have named Urochitoxin, Amorphous Urechitoxin, and Urechitin. Preparation of Urechitoxin and Amorphous Urechitoxin. The active principle of the plant is dissolved out by water, but the extraction is very incomplete, even when large quantities of water are used, while a great amouiit of iuactive matter is also taken up which complicates the subsequent purification. Alcohol easily and com- pletely removes the bitter substance, but unless it be nearly absolute, it also takes up much extractive matter; on the other hand, if veryBOWREY ON THE POISONOUS PRIKCIPLE, ETC.253 strong, it dissolves green resinous matter, which is, however, easily got lid of by dilution. These observations led me to the following process :-The leaves and green tips were first air-dried, and then at 100" C., as long as they gave off moisture, air also heated to 100" C. being passed over them to hasten the drying. The now very crisp and brittle leaves were finely powdered as rapidly as possible, so as to avoid absorption of moisture from the atmosphere, and the powder was extracted with spirit of at least 98 per cent. alcoholic strength. If maceration was continued €or a sufficiently long time, a comparatively small volume of spirit rendered the extraction practically complete. The dark-green tincture obtained was distilled to recover part of the spirit; but if it was well made, and therefore very strong, distillation was advantageously dispensed with ; whether or no, the next step was t o add about an equal bulk of boiling water, pouring it into the tinc- ture gradually, and stirring vigorously meanwhile ; this caused a green coloured resinous precipitate, which was removed by filtration, and after washing with spirit of 50 per cent. alcoholic strength, was found to be devoid of bitter taste.The clear filtrate, now of a yellow-brown colour, was mixed with the washings, and a strong solution of basic acetate of lead added as long as a yeEZow precipitate fell, care being taken not to add more than just enough for this purpose, or a loss of urechitoxin resulted from its partial precipitation together with the coloured precipitate.This loss is due simply to dilution of the spirit by the lead solution. The yellow precipitate was filtered off, and washed once or twice with spirit of 50 or 60 per cent.; prolonged washing would remove all bitter matter from it, but is not economical. The mixed clear yellow filtrate and washings were next freed from lead by means of snl- phuretted hydrogen and filtration, and as much urechitoxin is carried down by the lead sulphide, it must be well washed with hot spirit of 50 or 60 per cent. On evaporation in a vacuum over sulphuric acid, the acid liquors deposited small white crystals ; when their quantity ceased to increase, the mother-liquor was removed by means of the filter- pump, and evaporated to dryness over oil of vitriol in a vacuum, yielding a resin-like mass of crude amorphous urechitoxin.The crystals were washed on the filter with spirit of 20 per cent., and then dissolved off in a little boiling 30 per cent. spirit. The solution was quite colourless, and on cooling microscopic crystals of urechitoxin separated out in such quantity as to render the whole mass pansty ; they were collected on a Bunsen's filter, washed with a little cold 30 per cent. spirit, and dried in a vacuum. The mother-liquor and washings evapomted in a vacuum over oil of vitriol yielded a further crop of urechitoxin, and a little amorphous urechitoxin.The crude amorphous urechitoxin mentioned above was dissolved in POL. XXXIII. U254 BOWREY ON THE POISONOUS PRINCIPLE a little absolute alcohol, to get rid of any inert extractive matter present, the solution evaporated at a gentle heat until syrupy, and while it was still-warm, hot water was added gradually, the mixture being vigorously stirred all the time, so as to bring the water into contact with every part of the resulting soft resin. After removing the water, the resinous mass, if nearly colourless, was heated on the water-bath for a few minutes, then placed over oil of vitriol, and a vacuum produced as quickly as possible ; this caused it to swell and froth up, and speedily lose all moisture, the product being a porous, easily powdered, light amber-coloured mass of amorphous urechitoxin. If, however, the resin precipitated by the hot water was dark in colour, it was dissolved in spirit of 50 or 60 per cent., and purified animal charcoal added until most of the colour was removed, the charcoal separated by filtration, and the filtrate evaporated nearly to dryness at a gentle temperature, and finally dried in a vacuum as de- scribed above.I found it impossible entirely to bleach solutions of amorphous urechitoxin by means of animal cbarcoal, and even sufficient charcoal to remove most of the colour abstracts a large proportion of the poison ; it is therefore necessary to wash the charcoal with boiling spirit of about 60 per cent., in order to avoid a considerable loss of amorphous ui*echitoxin.Urechitoxin aud urechitin are also removed, more or less, from their solutions by animal charcoal. Much of the urechitoxin and amorphous urechitoxin I have prepared has been obtained by the foregoing method, but samples have also been made by the following modified and less troublesome process. After the removal of the lead by sulphuretted hydrogen, the liquid was distilled in a vacuum, nearly to dryness, at a temperature not exceed- ing 35", whereby a soft resinous residue, perfectly homogeneous and free from crystals, was obtained ; it was mixed with several times its bulk of spirit of 15 per cent., and allowed to stand for several days, when its appearance was found greatly changed ; it now consisted of a mas8 of microscopic crystals of urechitoxin, mixed with more or less ;imorphous urechitoxin in the form of minute oily drops.The weak spirit was of a dark colour, being charged with the more highly- coloured portions of the residue ; it was drawn off by the filter-pump, and the crystals freed from amorphous urechitoxin by washing with successive small quantities of 20 per cent. spirit, and finally with a little 30 per cent. The final operations to obtain the two forms of the poisonous principle in a pure state were the same as those already described. Prom the perfectly dried leaves of the ZTrechites suberecta I have obtained as much as 1 per cent. of urechitoxin, and nearly 3 per cent. of mixed urechitoxin and amorphous urechitoxin. On an average theOF URECHITES SUBERECTA. 255 fresh leaves lose four-fifths of their weight on being completely dried; they must therefore contain rather more than one-half per cent.of their active principle. In one operation a much larger proportion of water than that re- commended above was mixed with the tincture, and no care was taken to conduct the evaporation at a low temperature, or to prevent access of air, the large amount of liquid present being driven off by the heat of an ordinary water-bath. The result was an unsatisfactory yield of amorphous urechitoxin, while no crystalline product whatever was obtained. After a satisfactory process for the preparation of urechitoxin had been worked out, the various samples obtained were mixed and re- crystallised from 30 per cent. spirit, the yield being about two ounces of dry crystals : Sample A.A small portion of A was recrystallised three times from 80 per cent. spirit, the crystals being freed from their mother-liquor each time by washing with spirit of the same strength: the result was Sample B. Another sample, C, was prepared by slow crystdlisation from a solution of a portion of Sample A in 60 per cent. spirit. When but a fraction of the amount in solution had sepamted the crystals were collected, washed, and dried. Samples A and B consisted of microscopic crystals perfectly clear and transparent singly, but snow-white in mass. The crystals in C were much larger, being nearly half-an-inch long. Preparation of Dechitin. The fact that occasionally no urechitoxin and but little amorphous urechitoxin was obtained from the dried leaves, coupled with some ob- servations made in the preliminary experiments of this research, led me to suspect that the poisonous principle of tohe plant underwent some change either in the process of drying or in the subsequent manipulations.If not, why was it that occasionally nothing but the amorphous form was obtained ? The experiments now to be detailed shed some little light on this question, and led to the discovery of the form of the poison which I have called urechitin. Five pounds of fresh leaves were finely minced and then exhausted in a displacement apparatus with successive quantities of nearly abso- lute alcohol, in all fifteen pounds of spirit being used. The tincture which came through first was dark brown, then followed some of a dark green, while the portions which came through as the exhaustion approached completion were light green : these were put on one side.On mixing the brown and dark green tinctures, much green matter n 2256 BOmTREY ON THE POISONOUS PRINCIPLE separated, which, after being washed with 50 per cent. spirit, was found to be free from bitter taste ; it was removed by filtration, and the clear yellow-green filtrate run into a bolt-head connected with another and larger one immersed in ice. The air was next pumped out of the vessels, and the smaller one surrounded with warm water ; distillation now proceeded briskly at temperatures ranging from 28" to 38". As the alcohol passed over, crystals contaminated with a little green resinous matter separated. The distillation was continued until the liquid began to froth over into the condensing vessel; the crystals were then collected on a filter, and the mother-liquor re- turned to the bolt-head and distillation in a vacuum again started, froth- ing being prevented by gradually running in the light-green tincture which had been put on one side ; this weak tiucture was run in so that the vacuum was not injured, and at such a rate as just to avoid frothing.By this manipulation the distillation was carried on to dry- ness at a temperature not exceeding 38". The dry residue was treated with nearly absolute alcohol in considerable quantity, whereby all bitter substance was dissolved, and much brown gummy matter left behind. The alcoholic solution was diluted with enough hot water to reduce its alcoholic strength t o about 40 per cent. ; this caused the precipitation of much inert green matter, which was removed by filtration, and was washed with a little hot 40 per cent.Rpirit. The washings and filtrate were returned to the bolt-head (from which of course the inert gummy matter had been cleaned out), and on standing deposited some white crystals, the quantity of which was greatly increased by distilling off the alcohol in a vacuum, as already described. When all the spirit had passed over, the crystals, which were now of a dirty green colour in mass, were collected on a filter and washed with water. The filtrate was of a brownish-yellow colour and very bitter; on evaporation it left a, not very considerable residue giving the colour reaction of the poisons (to he described further on), but as if much foreign matter was present.The crystals were dissolved in 80 per cent. spirit, and an alcoholic solution of basic acetate of lead added as long as it caused a precipi- tate, which was then removed by filtration and washed with strong spirit. The mixed filtrate and washings were next freed from lead by a stream of snlphuretted hydrogen and filtration, then distilled in a vacuum until most of the alcohol had passed over. A quantity of nearly colourless crystals of urechitin having separated during the distillation, they were collected and put on one side for further purification. The mother-liquor, on evaporation to dryness, yielded a comparatively small amount of what appeared to be amorphous urechitoxin containing a few crystals of urechitin.The purification of the crystals mentioned above was effected in the It has not yet been further examined.OF URECHITES SUBERECTA. 257 following manner ; they were ground in a mortar with barely enough nearly absolute alcohol to form a thin paste, and the mixture, after standing in a closed vessel for several hours, was thrown on a Bunsen'a filter, the alcoholic solution drawn through as completely as possible, and the powder washed with successive small volumes of the same alcohol as long as it came through coloured. The mixed alcoholic fil- trates had a light yellow colour, and on slight dilution with water, deposited some pure white crystnls, which were collected and added to the powder on the filter. On further dilution, more crystals were precipitated, but now mixed with much resinous matter (? amorphous urechitoxiu), from which I did not sccceed in separating them.The mass of white powder and crystals were dissolved at a tempera- ture of 60" in barely su5cient 70 per cent. spirit, and the clear solution allowed to cool very slowly ; by next morning a fine crop of perfectly colourless crystals had formed, which were collected, washed withnlittle 60per cent. spirit, dried in a vacuum over quick lime, and bottled. The mother-liquor was slightly coloured, a Eittle animal charcoal served to decolorise it, after which it was distilled in a vacuum as long as alcohol came over ; the crystals which formed as the alcohol passed off were collected (the watery mother-liquor held hardly anything in solu- tion), dissolved in strong hot spirit; and warm water was added so as to reduce the alcoholic strength of the solution to about 40 per cent.Crystals began to form at once and increased in quantity as the solu- tion cooled; they were filtered off and washed. Their mother-liquor, on still further dilutiou, deposited more crystals having the same ap- pearance and properties as those first foriued; the two crops were therefore mixed, dried over quick-lime in a vacuum, and labelled, " Urechitin, Sample B." On farther treatment, the liquors from '8 yielded a small amount of a bitter resin resembling amorphous urechitoxin, and also a minute quantity of a crystallisable body like urechitoxin, but the amount obtained was too little to enable me to determine if the two were really identical.The water in the fresh leaves complicating the extraction of the urechitin, I made an experiment to ascertain whether drying at ordi- nary temperatures would alter the result obtained from the fresh leaves. Five pounds of leaves were allowed to dry in the air as long as the? lost weight, their weight ultimately falling to 22+ ounces ; an experiment on a few grains of these air-dried leaves proved that at a temperature of 100" they woald lose nearly three ounces more. The 22+ ounces were reduced to a fine powder, and exhausted with nearly absolute alcohol, seven pounds of which served to remove all soluble matter. The tincture was treated much in the Bame manner The bottle was labelled, '' Urechitin, Sample A."258 BOWREY ON THE POISONOUS PRIXCIPLE as that from the five pounds of fresh leaves, but no basic acetate of lead was used in this experiment ; in fact, nothing but alcohol, water, and a little animal charcoal were employed to obtain the urechitin in the pure state.A small sample of fine bold crystals, pro- duced by a very partial and slow evaporation, in the cold, of a solu- tion in 80 per cent. spirit of the total quantity of crystals obtained. Procured from the mother-liquor of Sample C by diluting it with hot water until its alcoholic strength was reduced about one half. The solution was frequently shaken as it cooled, and deposited the crystals more massive, though somewhat irregular, than any other sample I have prepared excepting C. A small sample of minute crystals separated from the mother-liquor, and washings of D, by distilling off the alco- hol in a vacuum.As this sample proved by its reaction and also by the numbers it gave on combustion, to be contaminated with a small quantity of some other substance, I shall not again mention it. These three samples weighed 167 grains, equal to a yield of 0.48 per cent. of the weight of the fresh leaves used. As the loss of urechitin in the process of purification plainly fhr more than equalled the weight of impurity present in Sample B, it is safe to conclude that the fresh leaves contain at least one-half per cent. of urechitin. This operation gave me three samples of crystals, viz. :- Urechitin, Sample C. Urechitin, Sample D. Urechitin, Sample E. Urec7iitoxim contains no nitrogen. The carbon and hydrogen were determined in each of the three samples after drying at 100" C., with the following results :- Obtained.Percentage. Taken. co,. HfO. C. H. A. 0.2370 gram 0.5914 gram 0.1682 gram 61-15 7.89 0.2266 ,, 0.5088 ,, 0.1610 ,, 61.24 7-89 B. 0,2039 ,, 0.4580 ,, 0,143 ,, 61.26 7.86 0.2876 ,, 0.6478 ,, 0.2044 ,, 61.43 7.90 C. 0.1650 ,, 0.3709 ,, 0.1165 ,, 61.31 7.85 These figures prove the three samples to be practically identical, and give a mean of C 61.28, H 7-88, 0 30.84, as the percentage composi- tion of urechitoxin. The formula, C13HZoO6, is the simplest which fairly well represents these percentages :- C 60.94 C 61.28 C13H2005 requires H 7.81 . Analysis gives{H , 7188 { 0 31.25 0 3084 From spirit of about 30 per cent. alcoholic strength nrechitoxin usually crystallises in thin four-sided prisms, bounded at each end byOF URECHITES SUBERECTA.259 two planes, from very weak spirit or water in sharp pointed needles ; it does not crystdlise readily from strong spirit, indeed I doubt if crystals can be obtained from alcohol approaching absoluteness. The crystals break across their length very readily. After being dried between folds of bibulous paper, urechitoxin still retains abont 8 per cent. of water, which is lost over oil of vitriol or more rapidly at loo", no further loss results from heating to 120'. If the dry crystals be exposed to the atmosphere for a few hours, they gain from 8 to 9 per cent. on their dry weight. The exact amount of gain seems to depend on the t.emperature, the increase of weight in the laboradtory at Kingston, Jamaica, with an average temperature of 29", being one-ninth less than that noticed in some experiments made in the South Kensington Research Laboratory, with an average temperature, during the period of exposure, of 16.5".The gain in an atmosphere saturated with water was practically the same as that in the ordinary atmosphere of the laboratory. Mechanically dry urechitoxin is readily deprived of dl water at 200" without its crystalline appearance being alteted ; but if it little more moisture be present than it naturally takes up from the air, that temperature will cause it to melt into a clear gum-like mass, from which it is very difficult to remove all moisture, unless it be first cooled and finely powdered. Urechitoxin and its solutions are intensely bitter; half a C.C.of a water solution containing one part in 40,000 is strongly bitter, and a similar quantity of a solution of one in 100,000 hits a distinct, some- what astringent taste, but is hardly bitter. If left in contact with the lips or tongue for a short time, the poison produces a tingling, smarting sensation, and the affected parts feel swollen. If the smallest particle enters the nostrils it causes violent sneezing. It is very poisonous, as the facts given in my paper on its physiological action plainly prove ; in one experiment less than one milligram of it subcutaneously injected proved fatal to a cat in 16 hours ; in another an injection of three milligrams was fatal to a strong cat in one hour and a quarter. Urechitoxin in excess, left in contact with water for several days, gives a solution containing one part of the poison in about 1,500 of the solvent ; it is much more soluble in boiling water, in which the excess melts to a clear soft resin ; the solution, on cooling and standing for a few hours, deposits tt rather considerable crop of crystals, but permanently retains in solution about one-thousand th of its weight of urechitoxin. It may be well to mention here that whenever I speak of definite weighta or proportions of " urechitoxin," the dry substance is referred t o ; the loose combination which it forms with about 9 per cent.of water, I distinguished as " Hydrated Urechitoxin."260 BOWR.EY ON THE POISONOUS PRINCIPLE The solubility of iirechitoxin in spirit of various strengths, was de- termined by placing excess of the hydrated substance with the spirit in a corked flask which was frequently agitated.After standing for a day and two nights, the saturated solution was filtered off and a weighed quantity evaporated in a warm place, and finally heated in a water-oven as long as weight was lost. The temperature of the labo- ratory was about 15' during the preparation of the solutions. The following are the results obtained, the numbers being parts by weight of spirit required to dissolve one part of dry urechitoxin :- 965.0 spirit containing 10 volumes per cent. of alcohol. 875.0 ,7 7, 15 71 ,? 9 , 585.0 77 Y 1 20 99 9 9 :? 198.0 ,, 7, 30 99 9 , 9 , 65.0 ,, 99 4?0 9 , 97 9 , 140 ,, Y, 50 9 9 ,? 9 , 4.3 9 , ,> 60 9 , ? ? ?7 2.0 3) ,, 70 ? 9 7, 97 1.4 ?? ,? 80 9 9 ,Y 9 , The saturated solutions in 60, 70, and 80 per cent.spirit were clear, but thick and syrupy. The poison is taken up to saturation by 70 and 80 per cent. spirit in a few minutes. The stronger alcoholic solutions deposit urechitoxin in a pseudo-amorphons form on addition of water, or even on exposure to the air. The solution formed by placing excess of dry urechitoxin in contact Kith spirit is much stronger than that resulting from the use of the hrdrated substance, thus : 123 parts of 30 per cent. spirit dissolved 1 part of urechitoxin. 9.5 ,7 50 79 9 , 9 7 It is much more soluble in hot spirit; some of 30 per cent. was heated to its boiling point,, and dry urechitoxin gradually added. It seemed as if i t would be taken up indefinitely.As each portion was dropped into the spirit, it appeared to melt and blend rather than to dissolve. All the urechitoxin at hand waa used, and a clear solution like strong gum-water obtained, which cooled without crystallising, but on standing a short time, crystals began to form, a stream of light liquid ascending from each one. In the course of a few hours the solution was converted into a magma of minute crystals, in which but little liquid was visible, and the tube might be inverted without any- thing escaping. In this experiment 1 part of urechitoxin was dissolved by 1.6 parts of the hot 30 per cent,. spirit. When dry urechitoxin in excess was left in contact with dry ether of 0.720 sp. gr., 1 part was dissolved by 565 of the solvent : but when the hydrated crystah were mixed with the same ether saturated withOF URECHITES SUBERECTA.261 water, 2,230 parts were needed for solution. Dry urechitoxin placed in benzene required 3,820 parts for solution. By chloroform the dry substance is taken np indefinitely, forming a soft resin-like mass ; the hydrated poison is not so readily soluble, but is all but miscible. Amylic alcohol takes up dry urechitoxin as freely as does chloroform, but not so rapidly. When the hydrated crystals were mixed in excess with amylic alcohol, and the solution was filtered after standing for 24 hours, 1 part of urechitoxin was found to have dissolved in 3.7 parts of amylic alcohol. Urechitoxin is exceedingly soluble in glacial acettic acid. If a strong solution be diluted with water, the poison is to a great extent imme- diately precipitated in a pseudo-amorphous condition ; on standing, the dilute acid gradually deposits a further quantity in the crystalline form.The psetido-amorphous urechitoxin left in contact with the weak acid, or with water or dilute spirit, gradually and entirely changes into ordinary urechitoxin, apparently without undergoing solution. I apply the term pseudo-amorphous to the non-crystalline urechitoxin obtained by fusing the crystals in water, or by evaporation of their solution in chloroform, &c., indeed to any urechitoxin in a, non-crystalline state which by appropriate means can be changed into crystals, while I re- strict the term amorphous to a substance resembling nrechitoxin in many respects, but which I have entire13 failed to convert into a crys- talliqe form.Pseudo-amorphous nrechitoxin is reconverted into the ordinary urechitoxin by crystallisation from spirit, or by the prolonged contact of water or weak spirit in quantities far too small for its solution. Strong solutions of urechitoxin are very apt to deposit the pseudo- amorphous body, and i t is so deposited from all solutions evaporated at it high temperature. Sometimes a drop or two of a solution oE urechitoxin evaporated in the cold, for examination under the micro- scope, will give a residue in which no crystals can be detected ; but by allowing a little water or very weak spirit to evaporate in contact with it, the residue will eventually be wholly converted into crystals. Pseudo-amorphous urechitoxin is easily powdered ; the powder mixed with cold water becomes soft and aggregates together.I f the mixture be well incorporated by vigorous stirring for a few minutes, and then filtered, a cold supersaturated solution is obtained, which in a few hours throws down the excess in the crystalline form. Rapidly heated to between 170" and 180", nrechitoxin fuses with- out change; but if its temperature is gradually raised, it loses its crystalline appearance at about 150°, and at 160' is fully fused. By this slow heating it becomes slightly coloured, and its weight is diminished about three-quarters of a per cent. ; it is, moreover, per- manently changed into an amorphous form of the poison. A similar2 62 BOWREY ON THE POISONOUS PRINCIPLE change takes place to a small extent when its solutions are heated.This observation was made in recovering the urechitoxin used in deter- mining its solubility, &c. This was done by crystallisation from hot 30 per cent. spirit, the mother-liquor from one operation being used to recover a subsequent quantity, and so on. Finally the mother- liquor was partially evaporated on a water-bath. On cooling, horn- ever, very few crystals formed ; on still further evaporation and again cooling, no crystals were deposited. The liquor was now practically free from alcohol, yet very bitter. It was evaporated to dryness in a vacuum, leaving a resinous mass which refused to yield crystals to any treatment, and which perfectly resembled amorphous urechitoxin. In strong cold aqueous hydrochloric acid urechitoxin dissolves freely fo a colourless solution, which very speedily changes to yellowish- green, then becomes opalescent, and deposits crystals and more or less amorphous colouring matter.The reaction is complete in a few hours ; if heat be applied it takes place immediately, but at the same time some other action is induced, in consequence of which a much smaller yield of crystals, or even none at all, is obtained. All bitterness is de- stroyed by this reaction. The crystals are much less soluble in spirit than urechitoxin ; they crystallise in microscopic prisms and plates of a very light yellow colour, are tasteless, and give the urechitoxin colour reaction with greater brilliancy than the pure poieon. I have not yet obtained it pure enough in quantity suficient for analysis ; but by using fuming aqueous hydrochloric acid, diluted with one volume of water and two volumes of absolute alcohol, as the solvent for the urechitoxin, and allowing sufficient time, a quantity of a crystalline substance uncontaminated with amorphous matter was obtained.This body I have named urechitoxetin ; a description of it is given fur- ther on. The strongly acid filtrates from the completed action of hydrochloric acid on urechitoxin contain nothing bitter, but hold in solution a body which immediately reduces boiling alkaline cupric solutions, causing precipitation of red cuprous oxide. By removal of the acid and sub- sequent evaporation in a vacuum, this body or mixture of bodies was obtained as a non-crystallisable treacly fluid, of a dark colour, and possessing no marked taste, but smelling of burnt sugar. It awaits further investigation.A change somewhat similar to that produced by hydrochloric acid has been noticed to occur in water solutions of the poison left €or a long time in closed vessels. For instance, a solution containing 1 part of urechitoxin in 10,000 of water remained unchanged f o r several weeks, but in the course of months a substance resembling the mould which so frequently forms in solutions of tartaric acid, was de- veloped, and fine crystals precipitated. Neither the liquid nor the crys-OF URECHITES SUBERECTA. 263 tals now had any bitter taste ; the latter were comparatively insoluble in spirit, but gave the urechitoxin colour-reaction with great bril- liancy.Solutions of caustic potash and soda have no immediate effect on urechitoxin, but I have noticed that strong solutions of the poison in spirit which have been allowed to remain in an alkaline state for a considerable time lose their bitterness. In strong nitric acid it dissolves readily and quietly to a light yellow solution, which gives off red fumes on being heated, at the same time becoming lighter in tint. Colowr-reaction. Urechitoxin dissolves readily in strong pure sulphuric acid to a light orange-yellow solution having a tint of brown in it. If this SO- lution be diluted with water, a dirty light yellow precipitate is thrown down. If, however, it is allowed to stand in a closed vessel, it gradii- ally becomes of a, redder and redder tint, passing through carmine to mauve, and finally into purple, These changes are much hastened by heat, and if the heat be continued the colour passes into a brown of great depth.Water added to the purple solution causes a dirty orange-coloured precipitate. The colour changes are also produced by the addition of a small quan- tity of either of the following substances : nitrates, nitrites, chlorates, bleaching powder, bromine, and iodine. Ferrocpanide of potassium does not cause the colour changes, while ferricyanide brings the colours out slowly and without the brightness of tint produced by the reagents mentioned above. Chromates and bichromates change the yellow of the sulphuric acid solution to a very light yellow-green, the green gradually becoming stronger as the mixture stands, no doubt from the reduction of chromic acid.Permanganate of potash, added cautiously, first bleaches the solu- tion almost completely, then brings out a fine but not strong violet tint, which finally passes into a dirty brown colour. I prefer nitric acid for the production of the colour changes, and a8 excess destroys all colour, I am in the habit of using an oxidising agent prepared by adding about a drop of nitric acid to 100 C.C. of sul- phuric. acid. If a drop of sulphuric acid is placed on 0.01 milligram of urechitoxin lying in a white basin, a yellow solution is produced, in which the changes of colour to mauve are distinctly seen on adding a minute drop of the oxidising liquid. Even 0.001 milligram will with careful manipulation give the reaction distinctly.Tunic acid causes a white precipitate in a water-solution of urechi- toxin (1 part in 1,500), which is redissolved on addition of potash2 64 BOWREY ON THE POISONOUS PRINCIPLE or acetic acid; but no precipitate falls if the solution be slightly more dilute. The tannate is very soluble in spirit. The various forms of the poisonous principle of the Urechites smberectn are much more soluble in water when contaminated with extractive matters derived from the plant than when in a pure state. These impure solutions give dense curdy precipitates with tannic acid. Every bittes. water so- lution obtained from the plant, whether immediately or after the action of selective solvents, &c., I have found to give a precipitate with tannic acid, while no precipitate was formed in solutions devoid of bitterness.Urechit oxet in. The urechitoxetin prepared, as already described, by the action of alcoholic hydrochloric acid on urechitoxin, was carefully purified by repeated crystallisation from spirit, the final product being a quantity of light dull-yellow microscopic crystals with a silky lustre. They are readily and completely dried at loo", and on exposing the dry crystals to the air for 48 hours, they take up but 0.2 per cent. of mois- ture. On analysis the purified and perfectly dry urechitoxetin gave the following results :- Obtained. Percentage. Taken. coz. H2O. C. H. 02147 gram 0.6091 gram 0.1642 gram 77.37 8.50 02580 ,, 0.7338 ,, 0.1959 ,, 77.57 8-44 0.1920 ,, 0.5423 ,, 0.1464 ,, 77.45 8-32 The mean percentage composition obtained is C 77.46, H 8.49, 0 14.05, while the formula Cd4H5,0s requires C 77.42, H 8.50, 0 14.08.Urechitoxetin has no marked taste, and is physiologically inactive : hence I have not investigated its properties to any great extent, and will merely mention a few of the more marked. It is practically in- soluble in water and in weak spirit, nor does it dissolve to any large amount in spirit of 80 per cent., unless it be heated ; much crystal- lisev out as the solution cools. It appears to resemble urechitoxin in a certain proneness to pass into an amorphous modification. It dissolves in sulphuric acid to a light orange-yellow solution ; if the solution is very strong, the colour is orange-red, but the increase in depth of tint is by no means proportioned to the amount dissolved : for after the redder tint is once reached, no marked gain in colour is seen on adding very much more urechitoxetin.On adding a minute quantity of nitric acid to the yellow solution, the same colour-changes are produced aa urechitoxin similarly treated exhibits, but they are much clearer and brighter. If the oxidising agent be added to a strong solution, the increase of colonr is very remarkable, the oxidised solution appearingOF URECHITES SUBERECTA. 265 black and opaque except in a very thin stratum. Heat brings out the colour changes, and they also appear if the solution in sulphuric acid be allowed to stand in a closed vessel in the cold, but several days pass before the purple tint is reached. Water added to a fresh solution in pure sulphuric acid throws down a light canary-yellow precipitate, while if added after the purple colour has developed, the precipitate is dirty orange.Excess of oxidising agent quickly bleaches a sulphuric acid solution of urechitoxetin without showing the various colours with any clearness. Amorp how Ur ec hit oxin. In the preparation of urechitoxin from leaves of Urechites euberectcc dried at loo", several samples of amorphous urechitoxin were ob- tained, great care was taken to remove all inactive matter, and as in their physiological and general chemical properties they closely resembled each other, I hoped they would prove to be of definite composition ; analysis has, however, disappointed this expectation, the percentage of hydrogen found in different samples varjing from 7.85 to 8.19, and the carbon fram 62.02 to 64.46.Some recent ex- periments lead me to suspect that several bodies exist having the physiological activity of urechitoxin, some of them, no doubt, being ready formed in the plant, while others are produced by chemical changes. If this suspicion is correct, the samples of amorphous ure- chitoxin I have prepared are uncertain mixtures of two or more of these bodies. Amorphous urechitoxin is, in appearance, intermediate between a resin and a gum; it has a light, dull yellow colour, and is readily powdered. It resembles urechitoxin in its toxic action, and is nearly, if not quite as poisonous; introduced into the mouth or nostrils it seems even more active, but this results simply from its greater solubility.In cold water it becomes soft ; excess placed in weak spirit gradually melts into an oil-like liquid ; it is miscible with strong spirit. It reacts with reagents in the same manner as does urechitoxin, but gives no cystaZZine product when acted on by hydrochloric acid, and perhaps its colour reaction is less vivid. Urec hitin. This substance usually crystallises in long fonr-sided prisms, bounded at each end by two planes, but frequently more complicated forms appear. The crystals are transparent and colourless ; they break very easily at right angles to their longest axis. As deposited from alcoholic solutions, the crystals contain 6 per cent, of water, which is readily driven off at loo", no further loss resulting from266 BOWREY ON THE POISONOUS PRINCIPLE coz.0.5132 gram 0.6465 )) 0.3892 ,, 0.3952 ,, 0%02 ,, 0.7044 ,, 0-6004 ,, 0'5277 ,, 0.5263 ,, exposure to a temperature of 120" C. The anhydrous poison does not recover more than 1 per cent. of water if left in contact with cold damp air. Analysis of the crystals after drying at 100" gave the following results :- Urechitin contains no nitrogen, &O. ----- 0 -1592 gram 0'2002 ), 0.1123 ), 0'1235 ), 0.2176 02182 ,) 0.1850 ), 0.1633 ,, 0.1639 ,, Taken. Sample A. 0 ,2113 gram 0,2671 ,, 0*1480 )) Sample B. 0.1622 ,, Sample C. 0.2887 ,, 0.2905 ,, 0.2474 ,, Sample D. 0.2169 ,, 0.2167 ,, ~~ - Obtained. Percentage. C. 66 -24 66 -01 66 *19 66 4 5 66 *15 66 -13 66 -19 66 *35 66 -24 -- H. 8 -37 8 *33 8 43 8 -46 8 -38 8 -35 8 '31 8 *36 a -38 These figures show the four samples to be identical, and give a mean of C 66.22, H 8.38, and 0 25-44), as the centesimal composition of urechitin.The formula, C2&0t1, closdy represents the analytical results :- C 66.22. { 0 25.40. C,H4,02 requires H 8-30 . Obtained H 8.38. { 0 25.30 C 66.40 Urechitin in the solid form is tasteless, but its alcoholic solutions are equal t o those of urechitoxin in bitterness, and it also resembles this substance in its toxic action and power. In cold water and cold spirit of less than 40 per cent. alcoholic strength, it is practically in- soluble. Boiled in water it shows no sign of fusion; the liquid filtered hot and allowed to stand for a considerable time deposits a very small quantity of the poison in the crystalline state.Its solubility in various liquids was determined in the same manner as that of urechitoxin, with the subjoined results, the numbers being parts by weight of the solvent required to hold one part of urechifin (weighed after drying a t 100") in solution. The determinations were made at a temperature of about 16.5O. 1,580 spirit containing 50 per cent. of its volume of alcohol. 610 I9 60 99 ?? 190 ?9 70 9 , 9 , 88 ?9 80 $9 ?? 35 very nearly absolute alcohol.OF URECHITES SUBERECTA. 267 227 dry ether, sp. gr. 0.720. 420 same ether saturated with water. 140 amylic alcohol. 466 benzene. 2.7 chloroform. 167 boiling 30 per cent. spirit, the solution beginning to deposit crystals immediately on its temperature falling. Urechitin differs from urechitoxin in not taking a pseudo-amorphous form 4 its solutions i n the various menstrua already mentioned de- posit it in the crystalline form wbether they be evaporated slowly and at low temperatures, or on the water-bath.It is also precipitated in crystals on adding water to its solution in strong spirit or glacial acetic acid. It is freely soluble in this acid, though not to the same extent as urechitoxin, but it does not dissolve in dilute mineral acids. When a small quantity, in a thin test-tube, was suddenly immersed in oil already heated to 220" it changed in appearance, but did not absolutely fuse ; the tube was withdrawn, and when the temperature had risen to 260" it was again immersed; the poison now fused immediately, and on cooling was like colourless glass; it dissolved readily in hot strong spirit, and on evaporation the usual crystals of nrechitin were obtained.Another portion was gradually heated ; no alteration was visible until the temperature was over 180°, when the crystals began to colour here and there, but showed no signs of melt- ing. When the bath was nearly 195" points of fusion were ob- served where the colouring had become most marked; the tempe- rature was kept about 1 9 7 O , and the change gradually progressed, while fumes were evolved which condensed on a cold surface to clear drops having a strong acid reaction. Ultimately the whole fused to a dark-brown mass, which was disposed to froth up as if gas were being generated in it. It was now removed from the oil-bath, and was found to have lost one-tenth of its weight.The cold fused mass was next treated with a few drops of cold strong spirit, in which it dis- solved readily, with the exception of some crystals, which proved to be urechitin still unchanged by the heat. The alcoholic solution failed to yield crystals to any method ofatreatment, but on evaporation v gave a substance closely resembling some of my samples of so-called amorphous urechitoxin; it was intensely bitter, and gave the colour- reaction well, but was less soluble than any other amorphous sample of the poison I have prepared. Urechitin is acted on by strong aqueous hydrochloric acid much in the same way as urechitoxin; it is, however, not so freely soluble, and the solution on standing becomes much more strongly green. Crystals268 BOWRET OS THE POISONOUS PRIhXIPLE, ETC.are deposited which, as far as I have examined them, resemble those resulting froin the action of the acid on urechitoxin; but the liquid from which the crystals have been deposited does not, after neutralim- tion, so readily reduce alkaline cupiic solutions as does that €om ure- cliitoxin. With sulphuric acid and oxidising agents, urechitin gives the same series of colours as urechitoxin, but they are hardly so bright, and are certainly inferior to those given by urechitoxetin. The difference is most marked in the fresh solution in pure sulphuric acid, which is of ft brownish-yellow instead of the reddish-yellow d o u r of the mechitox- etin solution ; the brown is very decided if the solution is strong. On diluting the freshly-made solution with water, a dirty green precipitate falls; if the water be added after the colour-changes have been developed by nitric acid, a precipitate similar in colour to that given by urechitoxin or urechitoxetin, viz., dirty orange, is thrown down.I have noticed that, by mixing sugar with urechitoxetin before trying its colour reaction, results very much like those yielded by urechitoxin and nrechitin can be obtained. That the \.rtrious bitter poisonous substances described in this paper are glucosides I think is certain, but I do not consider sufficient data have been accumulated to allow a molecular formula to be assigned to either urechitin, urechitoxin, or urechitoxetin. The formulse given in this paper are merely the simplest that express the numbers obtained by analysis, and it may well be that none of them represent the mole- cule, bnt that some much heavier formulae differing but slightly in percentage numbers are correct.The facts here recorded, together with others observed in the course of the research, lead me to think that if sufficient time were devoted to experimenting on Urechites suberecta, it might yield as many sub- stances, and as confusing rksults as Digitalis purpurea. I have con- fined myself in this paper mainly to two definite crystalline bodies, and have left uuenumerated several less definite though interesting substances, which have been noticed during the research. For instance, in one experiment on the preparation of urechitoxin, I got a small quantity of a substance as soluble in pure water as gum arabic, and possessing the toxic powers of the plant in a high degree; but having failed hitherto to obtain it in such a condition as would leave no reason to doubt that it was a definite compound and not a mixture, I do not think that honouring it with a name and detailed descrip- tion, would tend to anything except confusion. That the poisonous properties of Urech,ites suberecta are mainly due to urechitin is, I consider, established by the mode of its preparation, as it is unlikely that chemical change would be induced by tho use of nothing more than water, alcohol, and a temperature never exceedingHANNAY ON A NEW MANGANESE REACTION. 269 38'. On the other hand, I have no doubt that urechitoxin is a product of chemical change from urechitin, a change wrought by the much more violent method of its preparation. To guard against mistake, it may be well to add that whenever I speak of a substance as devoid of bitkerness, tasteless, and so on, without mentioning whether it was in the solid form or in solution, it is to be understood that it was experimented with in both con- ditions. I have succeeded in detecting these poisons when present in very minute quantities in complex mixtures by processes which their solubilities will suggest to any one familiar with the ordinary methods of detecting vegetable poisons in toxicological investigations ; but as my experiments on the best methods of discovering them are not completed, I will not refer to their detection here. In conclusion, I wish to state that the preparation of the various substances described was carried out in the Government Laboratory, Kingston, Jamaica, where most of the qualitative results were also worked out, while nearly all of the quantitative experiments were performed in the Research Laboratory, South Rensington, a.nd that my best thanks are due to Dr. Frankland for kindly permitting me fo work there.