TETANIN. 321 



patient, identical in its physiological and chemical reactions with that 

 isolated from cultures of Rosenbach's germs on beef-broth. The 

 presence of tetanin during life in tetanus patients has thus been 

 demonstrated. It has not been found in the brain and nerve tissue 

 of persons dead from tetanus. A portion of the jelly-like mass taken 

 from the amputated arm was found to contain tetanus bacilli as well 

 as staphylococci and streptococci, and when planted on beef-broth 

 tetanin was formed, but no tetanotoxin or spasmotoxin. 



Kitasato and Weyl (1890), employing pure cultures of the tetanus 

 bacillus, obtained from 1 J kilograms of beef used as culture medium 

 1.7118 gram of tetanin hydrochlorid (0.137 per cent.). Tetano- 

 toxin was also present. 



For its isolation Brieger employed the following method : The 

 cultures were slightly apidulated with hydrochloric acid, heated and 

 filtered ; the filtrate was then treated with lead acetate and with alco- 

 holic mercuric chlorid in the manner described under mytilotoxin 

 (page 313). Kitasato and Weyl digest the cultures with 0.25 per 

 cent, hydrochloric acid for some hours at 60°, then render slightly 

 alkaline, filter, and distil in vacuo at 60°. The residue in the retort 

 is worked for tetanin by Brieger's method, while the distillate con- 

 tains tetanotoxin, ammonia, indol, hydrogen sulphid, phenol, and 

 butyric acid. The filtrate from the above mercuric chlorid precipitate 

 contains the greater part of the active principle, provided the precipi- 

 tate has been thoroughly washed. After the removal of the mercury 

 by hydrogen sulphid the liquid is evaporated, and the residue is re- 

 peatedly extracted with absolute alcohol, in which the tetanus poison 

 readily dissolves and can thus be separated from the insoluble am- 

 monium chlorid. The alcoholic solution is treated with alcoholic 

 platinum chlorid which precipitates the ammonium and creatinin 

 platinochlorids, whilst the platinochlorid of the poison remains in 

 solution. The filtrate from this precipitate gives, on the addition of 

 ether, a flocculent precipitate possessing exceedingly deliquescent 

 properties. The precipitate is, therefore, rapidly filtered off by means 

 of a pump, and dried in vacuo. It can then be recrystallized from 

 hot 96 per cent, alcohol, and the beautiful clear-yellow plates thus 

 obtained, if dried again in vacuo, become rather difficultly soluble in 

 water, from which it can then be recrystallized and obtained in a per- 

 fectly pure condition. If boiled with boneblack, it decomposes 

 yielding a non-poisonous crystalline compound. 



Phosphomolybdic acid cannot be used in the separation of tetanin 

 inasmuch as it destroys the poison (Brieger). Bocklisch has also 

 observed that this reagent destroys the poison formed in the putre- 

 faction offish. 



Tetanin obtained by treating the hydrochlorid with freshly pre- 

 cipitated moist silver oxid forms a strongly alkaline yellow syrup. 

 With alkaloidal reagents it gives the same reactions as the hydro- 

 21 



