LESSON vii TOXINE REACTIONS 159 



(6) Inoculate two flasks, containing 100 cc. each, with 

 Bacillus diphtherise. 



(7) After twenty-four to thirty-two days filter the cul- 

 tures through a Berkefeld filter. 



(8) Evaporate the filtrate down to a small bulk at 40 C. 



Reactions 



(a) Millon's reaction : more or less typical. 



(b) Xanthoproteic reaction : more or less typical. 



(c) Alcohol : precipitate. 



(d) Acetic acid and ferrocyanide of potassium : tur- 



bidity after some time. 



(e) Phospho-molybdic acid : slight precipitate. 



(/) Ammonium sulphate : no precipitate ; hence no 

 albumoses present. 



(g) No biuret reaction; hence no albumoses or 

 peptones present. 



To obtain the toxine in a dry condition, pour the filtrate 

 into an excess absolute alcohol after it has been evaporated 

 down to a small bulk. 



Allow the precipitate to settle and to stand under 

 alcohol for a few days. 



Separate off the alcohol, and dry the residue at 40 C. 



The readiest method, for all practical purposes, of 

 obtaining diphtheria toxine in solution is to filter 

 virulent broth cultures 

 as described on p. 141. 



