Identification of Digitaline, Picrotoxine and Colchicine
It has already been remarked that in exhausting the first acid solution with ether—previous to the neutralization, according to Otto’s method—colchicine, a weak alkaloid, digitaline, an indefinite mixture, picrotoxine (which appears to possess the properties of an acid), and traces of atropine, pass into solution.
The ether is evaporated on a water-bath to dryness, the residuary mass treated with slightly warmed water and the solution filtered from the undissolved resinous matter. The aqueous solution is next rendered feebly alkaline by addition of soda lye, and then well agitated with ether, until this fluid ceases to leave a residue upon evaporation. The ethereal solution is now decanted, and the water present removed by means of chloride of calcium. If it is evaporated, a residue containing the colchicine, digitaline and traces of atropine (mixed possibly with a minute quantity of picrotoxine, which is here left out of consideration) is obtained.
a. The alkaline solution, from which the ether has been removed, is acidulated with hydrochloric acid and again shaken with ether. The picrotoxine present is now dissolved, and upon dehydrating (by means of fused chloride of sodium) and evaporating the ethereal solution can be obtained in crystals. The crystals of picrotoxine are easily recognized by their forming in feathery tufts as well as by their length and silky brilliancy. Should crystals fail to form in a short time, it is advisable to take up the residue, left by the evaporations of the ether, with slightly warmed alcohol, and to allow the latter to spontaneously evaporate on a watch-glass, or, if the quantity of substance is exceedingly minute, on the slide of a microscope. After determining the form of the crystals, it should be ascertained that they possess an intense bitter taste and exhibit the other characteristic properties of picrotoxine. The following reaction is distinctive: If the crystals are dissolved in an aqueous solution of soda and a few drops of “Fehling’s solution”O added, a reddish precipitate of cuprous oxide is formed.
b. Provided picrotoxine has not been found, the ethereal solution obtained by agitating the alkaline fluid with ether is to be examined for colchicine and digitaline. To this end, the residue obtained upon evaporating the solution to dryness is taken up with water, and the filtered fluid tested as follows: 1. It is ascertained if a drop of the solution possesses the bitter taste of digitaline. 2. Another drop is treated with solution of tannin; if either alkaloid be present, a precipitate is formed. 3. Two drops of the solution are next tested: one with tincture of iodine, the other with chloride of gold. These reagents precipitate colchicine, but do not affect solutions of digitaline or picrotoxine. Unfortunately traces of atropine, possibly present, would cause the same reaction; the test therefore fails to be conclusive. 4. Several portions of the solution are evaporated on watch crystals. Concentrated nitric acid is added to one portion: if colchicine be present, an evanescent violet coloration is produced, which changes to a light yellow upon addition of water, and to a pure yellow or reddish-orange color, if the mixture is saturated with a slight excess of caustic alkali. 5. Another portion of the residue is dissolved in a few drops of concentrated sulphuric acid, and the solution stirred with a glass rod moistened with bromine water: in presence of digitaline a violet-red color is produced. This coloration is more distinct when a small quantity of the alkaloid and an excess of sulphuric acid are present. 6. If a large amount of substance is at hand, the residue can be boiled with hydrochloric acid, and the green or brownish color and characteristic odor of digitaline produced, in case this body be present: this, however, is not a very delicate test. 7. Finally; it is advisable when the presence of digitaline is suspected to ascertain its physiological action. For this purpose, a minute quantity of the substance is placed upon the heart of a frog: in presence of the alkaloid, the pulsations are immediately retarded, or even arrested.
Although by means of the tests given above the existence of a special alkaloid, or of one of the ill-defined substances just mentioned, may be justly regarded as probable, its presence has not yet with certainty been demonstrated. This is especially true in cases where the compound possesses but few characteristic properties. When possible, the suspected substance should be obtained in a crystaline form, and then compared by aid of the microscope—if the small quantity present permits of no other examination—with crystals of the pure alkaloid, prepared under the same conditions.
In case 20 or even 10 centigrammes of substance are at hand, it is best to convert the alkaloid into its hydrochlorate, and evaporate the solution of this salt to dryness. The residue, after being weighed, is dissolved in water, and a solution of sulphate of silver added. The precipitate of chloride of silver formed is collected and carefully weighed, in order to calculate the weight of the chlorine contained in the hydrochlorate and consequently the molecular weight of the alkaloid. The filtrate from the chloride of silver, which contains the alkaloid in the state of sulphate, is treated with hydrochloric acid, to remove the excess of silver present and the fluid then filtered. The filtrate is next shaken with potassa and ether. Upon decanting and evaporating the ethereal solution, a residue consisting of the alkaloid present is obtained, which is then purified by crystallization from alcohol. An elementary analysis of the alkaloid is now executed. Certainty as to the presence of an individual alkaloid is attainable only when the execution of this confirmatory test is possible. The reactions previously described can be performed with fifteen centigrammes of substance, and this amount is sometimes contained in a cadaver. If but one or two centigrammes are at hand, it is still possible to detect the presence of an alkaloid; a conclusion, however, as to which cannot be arrived at, especially if the substance found is a liquid or an amorphous body, and one that presents few distinctive properties.