118 C. C. WARDEN, J. T. CONNELL AND L. E. HOLLY 



carried on to the extent one would desire seem clearly to indi- 

 cate a protective influence, specific or otherwise, but propor- 

 tionate to dosage, on the part of the antitoxin. 



Further observations upon the production of artificial toxin 

 mixtures have suggested the availability of emulsifying sub- 

 stances other than proteins, such, for instance, as the dyes, of 

 which Congo red has thus far alone been tried. This question 

 is of interest in connection with the coloring matter of broths 

 noted earlier. 



B. MEGATHERIUM. THE ANTIGEN 



The strains of B. megatherium were two in number, one of our 

 own, and that known as No. 7 kindly sent us from the Museum 

 of Natural History, the latter having been used and commended 

 by Rous, Robertson and Oliver. ^^ 



Heavy cultures of the organism were grown for twenty-four 

 to thii'ty-six hours at 35°C. in Roux flasks on beef-peptone 1 

 per cent agar, the mass removed in small amounts of water and 

 saponified in the manner previously described. The collected 

 fatty acid complex was then examined and found to consist of 

 approximately 56 per cent of insoluble unsaturated fatty acid 

 and 44 per cent of volatile fatty acid. The saturated fraction 

 obtained by steam distillation had a melting point of 30°C. a 

 neutralization value of 233 mgm. NaOH, and a calculated M.W. 

 of 175, showing that it probably consisted wholly of capric acid. 

 The insoluble residue from the distillation, after conversion into 

 the Pb salts and extraction with ether, showed the absence of 

 further saturated acids, and the fluid acid obtained by conver- 

 sion of the Pb salts gave an I.V. of 90, a neutralization value of 

 142 mgm. NaOH and a calculated M.W. of 283, data quite in 

 agreement with an oleic acid. The tentative formula for the 

 Megatherium antigen, then, consisted of oleic acid 56 per cent 

 and capric acid 44 per cent. This was different from any pre- 

 vious complex studied but resembled in physical characters 

 most closely that for B. anthracis. 



12 Jour. Exp. Med., 1919, 29, 283. 



