483 



to l'/ï cm. The flasks were then left standing in an incubator 

 at 36° C. Duplicate cultures were made for each experiment. 



When the bacteria were fairly developed (which was the case 

 after two days) one of the cultures was examined for the presence 

 of the amino-acids, alluded to above; the other remained in the 

 ncubator. Moreover a new culture-medium was inoculated with it. 

 We used Uschinsky's solution, unmodified or modified as indicated 

 below. ') 



Since the /;^ of the paunch contents was about 7,4, we took 

 care to let the pfj of our culture media be the same. 



In order to demonstrate the presence of tryptopiiane we applied 

 the reactions of Voisenet (with HCl, formaldehyde and nitrite) and 

 of HoPKiNS-CoLE (with HjSO^ and glyoxylic acid). Millon's reagent 

 was used for ascertaining the presence of tyrosine. Voisenet's rea- 

 gent stains differently with indole and with tryptophane. Indole after 

 shaking out with ether was reacted on with dimethylpara amidoben- 

 zaldehyde. 



UscHiNSKi's solution, whether modified or not, but invariably 

 without an aromatic or heterocyclic compound, inoculated with a 

 small quantum of the paunch-contents, always gave in the sediment 

 (obtained by centrifugation after the addition of alcohol) after a 

 sojourn at 36° C. in an incubator, a very distinct tryptophane, and 

 tyrosine- reaction, whereas initially the i-eactions were negative. 



A better growth and more powerful reactions were obtained by 

 mixing 10 c.c. of the fresh paunch fluid with 25 c.c of Uschinski's 

 solution. 



Whereas the reactions in the sediment were invariably positive, 

 the supernatant fluid displayed negative reactions. 



In order to make sure that the tryptophane and the tyrosine 

 reactions were not due to other indole or phenol-derivatives, the 

 sediment was, in a few cases, centrifuged anew with diluted alcohol 

 and once more with ether (indole). The reactions of the sediment 

 were as distinct as before. The cultures themselves were also shaken 

 out with ethei' some times. With the above-named aromatic 

 aldehyde the ether gave a negative indole-reaction. It was evident, 

 therefore, that neither free tryptophane, nor other free indole-deriva- 

 tives, nor free phenol-like bodies were present. The positive reactions 

 may, therefore, be attributed to the body-protein of the bacteria. 



On inoculation of new Uschinsky solutions with the cultures an 



1) The ordinary UscHiNSKY-solution contains K, Na, Ca, Mg, PO4, CI and SO4; 

 besides glycerol, ammonium -lactate and sodium aspartate. 



