395

XXXIIL-COMMUNICATIONS FROM THE RESEARCH LABORATORY 0 F THE PHARMACEUTICAL SOCIETY O F GREAT BRITAIN.

Contdmtions to our Knowledge of the Aconite Alkaloids. Part 111. The Formation and Properties of Aconine and its Conversion into Aconitine.

By WYNDHAM R. DUNSTAN, M.A., and F. W. PASSMORE, Ph.D.

IN aprevious paper (Part I, Trans., 1891,59,271), one of us, in con- junction with Dr. W. H. Ince, has described the principal properties of crystalline aconitine derived from Aconiturn napellus. The differ- ences between the chemical properties of the alkaloyd described by us and those recorded by previous workers prove that the substance we obtained was purer than any specimen hitherto described. Ex- periments then recorded showed that, on hydrolysis, aconitine breaks up, apparently, in accordance with the equation C33H&NO12 + H,O = C26H~lNOll + C7H602, benzoic acid being formed, together with an amorphous base, first obtained by Wright and Luff, and named by them aconine. Much uncertainty, however, still exists with yeference to the mode of formation and properties of this alkaloid, and since these points are of fundamental importance in connection with the question as to the constitution of aconitine, we determined to in- vestigate the nature of the base or bases resulting from the hydrolysis of the pure alkaloid. Wright and Luff (Trans., 1878, 33, 318) assert that aconine results from the action of water at a high tempe- rature, or of a mineral acid, or of alkalis on aconitine, the change occurring in accordance with the equation given above. They describe the base as being a slightly-coloured, friable, hygroscopic mass, readily soluble in water and alcohol, but nearly insoluble in ether. The aqueous solution is stated t o be bitter, but to produce no tingling sensation. It melts near B O O , and its composition is repre- sented by the formula C26H39NOll. Neither the base nor its salts could be crystallised. Wright and LufT further state that aconine acts as a reducing agent on salts of gold and silver. The pro- ducts of the hydrolysis of aconitine point to the conclusion that this alkaloid is benzoylaconine, and although Wright and Luff failed to reconvert aconine into aconitine by heating it with benzoic anhydride, &hey obtained what was probably benzoylapoaconitine.

Dragendoi-ff and Jiirgens" (J. Phnrm. [3], 18,277) have questioned

9 We have not been able to refer to the original paper, and are therefore 2 F 2

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396 DUNSTAN AND PASSMORE : FORMATIOX AND PROPERTIES

the accuracy of the account of the hydrolysis of aconitine given by Wright and Luff. They assert that the hydrolysis occurs in two stages. I n the first stage one molecular proportion of benzoic acid is separated with €ormation of a base which they state is identical with the picraconitine found by Wright and Luff in one commercial sample of roots, reputed to be those of A. napellus. I n the second stage, picraconitine undergoes hydrolysie, losing a molecular propor- tion of benzoic acid, together with methyl alcohol, and furnishing aconine, which is the final product of the change. If these facts are correct, aconitine must be regarded as dibenzoylmethoxyaconine.

Hydrolysis of dconitine.

The main purpose of our experiments was to ascertain whether any base, other than aconine, can be found at any stage of the hydrolysis of aconitine.

Pure aconitine (m. p. 188.5", corr.) was heated for several hours with water in a closed tube at 150". Water was used in preference t,o alkali or acid, so that the change might take place gradually, and without the resinification which always results from the use of alkali. The details of an experiment are as follows :-1 gram of aconitine was heated with 25 C.C. of water in a closed tube at 150-155" for 10 hours. After this treatment, the liquid still produced the tingling sensation characteristic of aconitine, and, therefore, presumably the whole of the latter had not suffered decomposition. On distillation, the liquid afforded no methyl alcohol or other volatile product. The solution was acidified with dilute sulphuric acid and extracted with ether, until nothing further was removed. The ethereal solution wa8 washed, dried, and distilled ; the residue of benzoic acid weighed nearly 0.1 gram, that is, about 10 per cent. of the aconitine taken ; the quantity of benzoic acid calculated from the equation given above is 18.8 per cent. ; rather more than one-half of the aconitine had, therefore, undergone hydrolysis, if the equation is correct. The acid solution from which the benzoic acid had been removed was made alkaline with ammonia, and completely extracted with ether.. The ethereal solution left, on evaporation, an amorphous residue, weighing 0.35 gram. This residue gave rise to the tingling sensation. I ts properties suggested that it was it mixture of aconine and acon- itine, and it was found that although dry aconine is insoluble in anhydrous ether, it may be extracted to some extent by ether from an aqueous solution, especially when aconitine is also present.

The liquid which had been extracted with ether was now shaken obliged to depend upon the abstract in English, which gives no experimental details.

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OF ACONINE AND ITS CONVERSION INTO ACONITINE. 397

with chloroform, which removed aconine with a trace of resin ; the residue left on evaporating the chloroform solution weighed 0.2 gram.

Excess of baryta-water was now added to the alkaline solution, which was evaporated to expel ammonia, and filtered from barium snlphate ; the excess of baryta was then precipitated by the requisite quantity of dilute sulphuric acid. The filtrate, when evaporated to dryness, left a residue weighing 0.43 gram, which was almost enti~ely soluble in a~lcohol, and consisted of nearly pure aconine. The resi- dues from the solutions in ether and chloroform were found to be mixtures of aconitine and aconine, and on heating them with water in a closed tube as before, more benzoic acid was formed, and sub- stances soluble in ether, chloroform, and water were again obtained. The tingling sensation produced by the original liquid proved that aconitine was still present. By a repetition of this process, nearly the whole of the original alkalo'id was converted into aconine. No methyl .alcohol was found at any stage in the process, and no trace of an alkalo'id having the properties of picraconitine could be detected.

The hydrolysis of aconitine was then repeated in the manner above described with several quantities of aconitine, varying from 1 to 5 grams, in every instance, with the same result: only benzoic acid and aconine were formed and neither methyl alcohol nor picr- aconitine could be isolated. We are, therefore, justified in concluding that the hydrolysis of aconitine inio aconine and benzoic acid takes place in accordance with the equatiou given above.

The amorphous base, not aconine, which Dragendorff and Jurgens thought to be picraconitine, may have been napelline, arising from impure aconitine having been employed. The circumstance that aconine, though insoluble in ether when dry, is nevertheless extracted from aqueous solution both by ether and chloroform, especially when aconitine and napelline are present, may also have led to the con- clusion that some alkaloid other than aconine had been formed. We regard it as probable that the methyl alcohol, supposed by these observers to be formed during the hydrolysis of aconitine, also took its origin in the use of impure aconitine, since we have noticed tha t organic solvents obstinately cling to the imperfectly crystallised alkalo'id and are set free when it is boiled with water. No such effect is observed when pure aconitine is employed.

Properties of Crystalline Aconine Hydrochloride.

Up to the present time, aconine has been described as an amorphous base, which forms amorphous salts. Wright and Luff speak of it as an amorphous, resinous alkaloid (m. p. 130°), readily soluble in water and alcohol, soluble also in chloroform, but insoluble in ether. Its

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398 DUWSTAN AND PASSMORE : FORMATION AND PROPERTIES

aqueous solution reduces salts of gold and silver, and also Fehling's solution. The salts formed from it were amorphous, and were not obtained pure.

In the course of the experiments relating to the hydrolysis of pure aconitine, a few grams of aconine were obtained. With this material we have studied the properties of the alkalo'id. This specimen of aconine was purer than any we had previously obtained, being almost if not entirely free from the resin which is formed along with i t when aconitine is acted on by alkalis. I ts aqueous and alcoholic solutions did not ci-ystallise when slowly evaporated in a desiccator, but finally dried up to a nearly colourless, friable, resinoid mass. The aqueous solution was decidedly alkaline, and had powerful reducing pro- perties. I ts taste was slightly bitter, but the tingling sensation pro- duced by aconitine was quite absent.

Since the base could not be crystallised, an attempt was made to crystallise its salts. The aqueous solution was exactly neutralised with dilute hydrochloric acid, and evaporated to a very small volume on the water-bath. The syrupy liquid, which at first showed no signs of crystallising, was allowed to stand for several days i n a vacuous desiccator over calcium chloride. It slowly became crystalline. The minute crystals were white, but the syrup which surrounded them was slightly coloured. The mixture was, therefore, drained on a porous tile in a desiccator, by which means the crystals were separate& almost entirely from the coloured syrup. This plan of purification was found to be less wasteful and more effective than recrystallisation from water or alcohol ; the salt absorbed by the tile was washed out, and more crystals were obtained from the solution. The white, dry mass removed from the tile consisted of minute crystals. Before their properties were examined, they were recrystallised by the addi- tion of ether to an alcoholic solution. The crystals were dissolved in a small quantity of strong alcohol a-nd anhydrous ether was then added until a faint turbidity appeared; on standing in a stoppered bottle, this solution deposited colourless rosettes of needle-shaped crystals, a further quantity being obtained by the addition of more ether to tbe remaining liquid.

When dried at loo", these crystals of aconine hydrochloride lost water without darkening ; 0.25025 gram of salt lost 0.0077 gram of water a t 100" ; that is, 2-92 per cent. When the temperature was raised to 120°, and the heating continued for a few hours, there wm a further loss of 0.0158 gram, after which the weight of the salt remained constant. The total loss in weight amounted, therefore, to 631 per cent.

It appears from these results that the crystalline hydrochloride contains two molecular proportions of water, and is represented by

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O F ACONINE AND ITS CONVERSION INTO ACONITINE. 399

the formula C~~H~,NO~~,HCl ,2HZO. water is lost at 100" and the other a t 180" ; thus :-

One molecular proportion of the

Loss calculated from the formula

3.08 per cent. C26H41NOil,HCl,SH20. Found.

lH,O.. . . . . . . 2H,O .... .... 5.95 ,, 6-31 ,,

2.92 per cent.

The dry salt is very hygroscopic and when exposed to air gradually absorbs the equivalent of one molecular proportion of water. I n order t o confirm the accuracy of the formula given above, the amount of chlorine in the dried hydrochloride was determined with the fol- lowing results :-

I. 0.1550 gram of salt gave 0.037 gram of silver chloride ; 5.93 per cent.

11. 0.3920 gram of salt gave 0.0975 gram of silver chloride; 6-15 per cent.

111. 0.3678 gram of salt gave 0.0917 gram of silver chloride; 6-22 per cent.

The mean of these three determinations of the chlorine is 6.13 per cent., whilst the amount calculated from the formula CZ6H41NOll,HC1 is also 6-13 per cent.

Anhydrous aconine hydrochloride melts at 175.5" (corr.). It readily dissolves in water and alcohol, and is slightly soluble in chloroform, but is nearly insoluble in ether and light petroleum. The aqueous solu- tion has at first a bitter-sweet and afterwards a burning taste. Like aconitine hydrochloride, it exerts a lzevorotatory action on polarised light, but its effect is much feebler. The anhydrous salt was employed in determining the specific rotation of an aqueous solution with the following result :-

a[15"] = 0.9" ; I = 2 dm. ; p = 5.75; d 15"/4" = 1.015,

whence 100 x -0.9 - - -7.71". = 2 x 1.015 x 5-75

The aqueous solution of aconine hydrochloride is neutral to litmus and is uot precipitated by alkalis even when concentrated. Picric acid does not precipitate the solution, neither does gold chloride, ex- cept in strong solutions, when an amorphous, pale-yellow precipitate is produced.

Other Crystalline Sal ts of Aconine,

Besides the hydrochloride, the hydriodide, hydrobromide, and sulph- All these salts are very soluble in water, ate were crystallised.

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400 DUNSTAN AND PASSMORE : FORMATION AND PROPERTIES

and do not crystallise 'as readily as the hydrochloride. The crys- talline sulphate is especially difficult t o obtain ; the aqueous solution when exposed in a desiccator, evaporates to a thick syrup without crystallising. On standing, very fine needle-shaped crystals appear i n the viscolns liquid, and are most difficult to separate from it.

Properties of Pure Bconzine.

Aconine was prepared from the pure hydrochloride by adding silver sulphate to the aqueous solution, slightly more silver salt being added than was required to precipitate the chloride. The precipitate of silver chloride was filtered off and the small excess of silver sulph- ate removed as aulphide. The solution of aconine sulphate thus ob- tained was precipitated by the addition of exactly the requisite quantity of baryta-water, the barium sulphate filtered off, and the filtrate evaporated to a small volume on the water-bath. The syrupy liquid was exposed in a, desiccator, when i t slowly dried up, without crystallising, to a colourless, hygroscopic, gum-like mass, which could not be crystallised from any of its solutions.

The amorphous base melts at 132" (corr.) without decomposition. On combustion, it afforded the following data :-

I. Weight of base, 0.1245 gram ; weight of carbon dioxide, 0,2614

11. Weight of base, 0.30610 gram ; weight of carbon dioxide, 0.6937 gram ; weight of water, 0.08985 gram.

gram ; weight of water, 0.22105 gram.

Calculated for I. 11. C,,H,,NOU.

C .......... 57.27 57-35 57-46 H.. ........ 8.02 8.02 7.55

The fact that the hydrogen is rather higher than the calculated quantity is to be accounted for by the extremely hygroscopic charac- ter of the base, which makes it difficult to manipulate without absorp- tion of water.

Aconine is very soluble in water or alcohol, and slightly in chloro- form, but insoluble in ether and light petroleum. The aqueous s o h - fion reduces salts of gold and silver and also Fehling's solution. It is precipitated by the general alkaldidal reagents, and mercuric chloride produces a copious yellowish-white precipitate, which darkens on standing.

The aqueous solution is decidedly alkaline and dextrorotatory. The determination of the specific rotation afforded the following results :--

4151 = +1*65; I = 2 dm. ; p = 3.534; d 15"/4" = 1.01,

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OF ACONINE AND ITS CONVERSION INTO ACONITINE. 401

whence

Aconine thus possesses the same peculiarity as was shown in Part I of this paper bo belong to aconitine, the base being dextrorotatory and the saltJs lzvorotatory.

When boiled with alkalis, aconine darkens and slowly resinifies. The physiological action of aconine is being investigated; it does not :appear to be poisonous in small doses.

Formation of Benzoylaconine.

The question as to the exact relationship of aconine to aconitine has been t o a certain extent elucidated by the experiments already described. It has been shown that pure aconitine yields only aconine and benzoic acid when hydrolysed and that the molecular formula of aconine is C,,H,,NOlr. I t is, therefore, almost certain that aconitine is benzoylaconine. I f this conclusion is correct, it should be possible t o prepare aconitine from aconine by introducing a benzoyl group. By the action of benzoic anhydride 011 aconine, Wright and Luff ob- tained, not aconitine, but a base probably identical with benzoylapo- aconitine which did not, apparently, exert the characteristic physio- logical action of aconitine. We have repeated this experiment with pure aconine, which was heated wi th benzoic anhydride in a closed tube at 100--130" for about three hours. No aconitine could be de- tected in the product. Some aconine was recovered unchanged, together with an amorphous base, which did not give rise to the tingling sensation and was not further examined. It appears to be the game substance as that isolated by Wright and Luff.

Since aconitine is so readily hydrolysed by acids and alkalis, we did not attempt to introduce the benzoyl group by means of benzoyl chlor- ide and alkali after the manner proposed by Baumann and Udransky.

The strongly basic properties of aconine suggested the probability ;that it might be able to decompose ethyl benzoate with the formation of ethyl alcohol and aconitine, or if the temperature of reaction were high enough to cause dehydration of the aconitine, anhydro-aconitine might be the final product. The reaction may be expressed by the equations

Conversion of Aconiwe in to Aconitine.

(Aconitine). {ii.) C,H,,NOI2 = HzO + C33H43NOI0.

(Aconitine.) (An hydro- aconitine).

It was found that aconine did not readily react with ethyl benzoate

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402 DUNSTAN AND PASSMORE : FORMATION AND PROPERTIES

under ordinary pressure, and the experiment was therefore made in a closed tube.

Half a gram of pure aconine was dissolved in 10 C.C. of alcohol, and 0.2 gram of ethyl benzoate added to the mixture, which was heated in a closed tube at 130" for three hours. Some ethyl benzoate re- mained unattacked a t the close of the experiment. The liquid was heated in an open flask on the water-bath until all the alcohol had evaporated. Water was then added and the liquid acidified with dilute hydrochloric acid ; a drop of this solution produced the charac- teristic tingling sensation on the tongue I Unaltered ethyl benzoate and benzoic acid were now removed by extracting the liquid with ether, the solution was rendered alkaline with dilute ammonia, and the alkalojid extracted with ether. The ethereal solution, on evapora- tion, left a resinojid mass which did not crystallise. By neutralisa- tion with dilute hydrobromic acid and evaporation of the solution, a colourless, crystalline hydrobromide was obtained. From this salt the alkalo'id was regenerated, dissolved in dilute hydrochloric acid, the solution precipitated with gold chloride, and the pale-yellow pre- cipitate of aurochloride washed and dissolved in alcohol. By the gradual addition of water to the alcoholic solution, the salt was de- posited in crystals which melted at 141" (corr.). The aurochloride of anhydro-aconitine melts at exactly this temperature (Part I).

By the reaction of aconine wit)h ethyl benzoate, we had thus obtained the anhydride of aconitine, and since this compound may be converted into aconitine, a partial synthesis of the natural alkaloid has been, effected, and the relationship of aconine and aconitine demonstrated beyond question.

It is intended to make further experiments on the conversion of aconine into aconitine and also to prepare other compounds of aconine containing various acid radicles. The examination of the physiological. action of these aconine derivatives is likely to throw light on t h e cause of the intensely toxic property with which the non-poisonous aconine becomes endowed when 8 benzoyl group is grafted on to its molecule.

Decomposition Products of Aconine.

The behaviour of aconine when attacked by various reagents has been closely investigated, but so far without much success, as i t has .not been possible to crystallise the principal decomposition products.

Nitrous acid does not appear to react with aconine. 0.7 gram of pure aconine was dissolved in 5 C.C. of water and mixed with a solution of 0.1 gram of sodium nitrite in 20 C.C. of water. The requisite quantit.y of dilute hydrochloric acid was then gradually added to the well-cooled mixture, After standing for some time, until the euolu-

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OF ACONINE AND ITS CONVERSION INTO ACONIT'INE. 403

tion of nitrous fumes had ceased, the solution was made alkaline and extracted with ether, which, however, removed nothing appreci- able. The aqueous solution was exactly neutralised with dilute hydrochloric acid and evaporated to dryness ; the residue extracted with boiling alcohol, and the solid left on the evaporation of the solvent again extracted with cold alcohol. Neither of these solutions contained any substance giving Liebermann's reaction for a nitroso- derivative and after evaporation in a desiccator and long standing, the latter solution furnished ci=ystals of aconine hydrochloride.

The oxidatiotL of aconine was effected with potassium permanganate in the following manner. About 1 gram of aconine dissolved in water was oxidised by the gradual addition of 3 grams of potassium permanganate, solution of potassium carbonate being added from time to time so that the liquid was always alkaline. The mixture was heated on the water-bath to complete the oxidation, then cooled and filtered. The precipitate of manganese oxide was extracted with alcohol, which removed a little resin. The filtrate was acidified with hydrochloric acid and extracted with ether, which removed very little. After it had been rendered alkaline with ammonia, the solution was extracted w-ith chloroform and subsequently with ether but nothing appreciable was dissolved from the aqueous solution by either sol- vent. The solution was now evaporated to a small volume on the water-bath and precipitated with lead nitrate. The precipitate, which contained some lead chloride, was collected, washed, and de- composed with hydrogen sulphide. The filtrate from the lead sulphide, when evaporated, deposited crystals of oxalic acid ; these were converted into the silver salt, which was analysed. It contained 71-24! per cent. of silver ; silver oxalate contains 71.05 per cent.

The filtrate from the precipitate produced by lead nitrate was evaporated, and the lead removed by hydrogen sulphide. No crys- talline acid or other product could be obtained from the solution.

Act ion of N e t h y l Iod ide o n dcon ine and Aconitine. Formation of Aconitine Methiodide and Methhydroxide.

The action of methyl iodide on aconine and on aconitine has been studied under various conditions. So far, no crystalline additive compound has been obtained from aconine, although it appears that when this alkaloid is heated with methyl iodide for several hours in a closed tube at loo", a base is formed which neither crystallises itself nor yields crystalline salts. This substance has not yet been further examined.

A crystalline additive compound of nconitine with methyl iodide is readily obtained.

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404 FORNATION AND PROPERTIES O F ACONINE, ETC.

Half a gram of pure aconitine was dissdved in a considerable ex- cess of methyl iodide. On heating the mixture a t 100" in a closed tube, nearly colourless crystals soon began to separate, and gradually increased in number until a t last the mixture became semi-solid. The excess of methyl iodide was drained from the crystals and found t o contain scarcely any of the new CompouIld, which seems to be only slightly soluble in this liquid. The crystals were dissolved in methyl alcohol and crystallised from the solution by the gradual addition of ether. By this means, aconitine nzethiodide was obtained in rosettes of crystals having a faint yellow t i n t ; these, after drying at 100", melted at 819.5" (corr.). The methiodide dissolves in alcohol, water, and chloroform, less readily in light petroleum and in ether. The aqueous solution is neutral to litmus. On analysis, the meth- iodide furnished the following result :-

0.2567 gram dissolved in 20 C.C. of water and precipitated with a slight excess of silver sulphate gave 0.0762 gram of silver iodide. The salt thus contains 16.04 per cent. of iodine. The formula C33H45N012,CH31 requires 16.09 per cent.

From the filtrate obtained in this experiment, the silver was re- moved by hydrogen sulphide, and then the free sulphuric acid was exactly precipitated with baryta-water. This solution of the sulphate deposited no crystals when slowly evaporated in a desiccator, and eventually dried up to a varnish.

The sulphate was redissolved in water, the aqueous solution made alkaline with ammonia and several times extracted with ether. The ethereal solution of aconitine rnethh ydroaide, C,,H,,NO,,, CH,*OH, on evaporation furnished a resinous residue which could not be crystallised. The aurochhide appears as a pale-yellow, amorphous precipitate when a solution of nuric chloride is added to an aqueous solution of the hydrochloride of the base. This salt did not crystal- lise fro= its alcoholic solution.

The properties of aconitine methhydroxide and its salts will be further examined, aud it is intended to investigate the physiological action of these compounds.

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