[669] Comptes Rend., cvi. 858, 861; cviii. 58-59, 809-810; cvi. 1604-1605.
§ 679. Poisons connected with Tetanus.—Brieger, in 1887, isolated a base of unknown composition, to which he gave the name of “spasmotoxine.” It was produced in cultures of the tetanus bacillus in beef broth.
Two more definite substances have also been discovered, viz., tetanine and tetanotoxine.
Tetanine, C13H30N2O4, is best isolated by the method of Kitasato and Weyl.[670] Their method of treating broth cultures of the tetanus bacillus is as follows:—
[670] Zeit. f. Hygiene, viii. 404.
The broth is digested with 0·25 per cent. HCl for some hours at 460°, then rendered feebly alkaline, and distilled in a vacuum. The residue in the retort is then worked up for tetanine by Brieger’s method; the distillate contains tetanotoxine, ammonia, indol, hydrogen sulphide, phenol, and butyric acid. On treating the contents of the retort by Brieger’s mercury chloride method, the filtrate contains most of the poison. The mercury is removed by SH2, the filtered solution evaporated and exhausted by absolute alcohol, in which the tetanine dissolves. Any ammonium chloride is thus separated, ammonium chloride being insoluble in absolute alcohol. The alcoholic solution, filtered from any insoluble substance, is next treated with an alcoholic solution of platinum chloride, which precipitates creatinine (and any ammonium salts), but does not precipitate tetanine. The platinum salt of tetanine may, however, be precipitated by the addition of ether to the alcoholic solution. The platinum salt, as obtained by precipitation from ether, is very deliquescent; it has, therefore, to be rapidly filtered off and dried in a vacuum. It can then be recrystallised from hot 96 per cent. alcohol, forming clear yellow plates; these plates, if dried in a vacuum, become with difficulty soluble in water.
Tetanine may be obtained as a free base by treating the hydrochloride with freshly precipitated moist silver oxide. It forms a strongly alkaline yellow syrup, and is easily decomposed in acid solution, but is permanent in alkaline solutions.