56 PROTEIN POISONS 



sand, mixed with water, 20 per cent, glycerin, or physio- 

 logical salt solution, to a dough consistency, and subjected 

 the mass to a gradually increased pressure of from 400 to 

 500 atmospheres. The clear, expressed fluid contained a 

 large quantity of coagulable protein, which decomposed 

 hydrogen peroxide, but lost this property on being heated. 

 It gave the protein tests and behaved like a nucleoprotein. 

 Ruppel 1 obtained a nuclein containing 9.42 per cent, of 

 phosphorus from the residue left after the preparation of 

 his tuberculosamin. This he called tuberculinic acid, and 

 he believed that it exists in the cell partly combined with 

 the tuberculosamin and partly free. Levene 2 prepared 

 three proteins from tubercle bacilli grown on protein-free 

 medium. The dried bacilli were ground for two or three 

 days in a porcelain mill, then extracted repeatedly for two 

 days with an 8 per cent, solution of ammonium chloride. 

 These proteins coagulated at from 50 to 64, 72 to 75, 

 and 94 to 95 respectively. Ammonium sulphate pre- 

 cipitated all of them; sodium chloride only the first; 50 

 per cent, magnesium sulphate the first; magnesium sul- 

 phate to saturation the second, but not the third. It 

 required less acid to precipitate the first, but 0.2 per cent, 

 hydrochloric acid precipitated all three. The third was 

 richer in phosphorus than the others, and Levene con- 

 cluded that the tubercle bacillus consists principally of 

 nucleoproteins, one of which differs from the others in 

 that it is not precipitated by magnesium sulphate and does 

 not give the biuret reaction. lie called attention to the 

 coincidence between the coagulation temperature of the 

 first protein and that necessary for the sterilization of the 

 bacillus. He believed that tuberculin is a specific substance 

 having the constitution of a nucleoprotein. He also made 

 a study of tuberculinic acid, finding but little of this free 

 in mannite synthetic cultures, but considerable in beef- 

 broth cultures. Samples differ in composition, and experi- 

 ments suggest that tuberculinic acid is less stabile than 



1 Loc. cit. 



2 Jour. Med. Research, July, 1901, 135; Medical Record, IS'.KS, liv, 873. 



