THE CLEAVAGE OF PROTEINS WITH ALKALI 97 



as we could judge by the development and intensity of 

 the symptoms, in similar amounts. That the poison could 

 be extracted by alkaline alcohol was proved. ' However, 

 the yield was not satisfactory, and a second test was made, 

 and in this the strength of the alkali was doubled. These 

 were treated as before, and the pigs that received the 

 injections developed the characteristic symptoms and 

 died. The one that had the alcoholic extract died within 

 six, and the other within eight minutes. This confirmed 

 the hope that the alcoholic alkali was quite as efficient as 

 the aqueous in the extraction of the poisonous group. 

 While the aqueous extract contained a large amount of 

 the carbohydrate group, it was found that the alcoholic 

 extract, after evaporation to dryness and solution in water, 

 gave the biuret, Millon, and xanthoproteic tests, but failed 

 wholly to give the Molisch reaction. The carbohydrate 

 group had been split off in both samples, but being insoluble 

 in absolute alcohol, it remained with the insoluble portion 

 of the cellular substance. 



The above and many other experiments have demon- 

 strated that the best method, so far devised, for extracting 

 the poisonous group from the cell protein, or, as subsequent 

 work has shown, from any protein, is by means of a 2 per 

 cent, solution of sodium' hydroxide in absolute alcohol. 

 If satisfactory results are obtained, the alcohol used in the 

 extraction must be absolute. If it is not, more or less of 

 the carbohydrate will be mixed with the poison; a sticky 

 mass will be obtained, and the patience of the experimenter 

 will be taxed severely. Previous thorough extraction of 

 the protein with alcohol and ether for the removal of fats, 

 waxes, and other substances soluble in these agents, is 

 also essential to satisfactory work. 



The method for preparing the bacterial cellular substance 

 has been given, but it may be well to give here some details 

 for the preparation of egg-white before splitting it up into 

 poisonous and non-poisonous proteins. 



Fresh eggs (we have usually taken twenty dozen at a 

 time) are broken and the whites dropped into a beaker or 

 7 



