220 BACTERTOLOGTCAL CHEMISTRY 



produce proteolytic enzymes, can never utilise proteins 

 or proteoses and their nitrogen must be supplied already 

 partially broken down to amino -acids or as ammonium 

 salts. Peptone (which is protein that has been sub- 

 jected to mild acid hydrolysis) in a medium serves 

 this purpose. 



Nitrogen Fixation 

 The phenomenon of nitrogen fixation, so important 

 from an agricultural standpoint, has been known from 

 time immemorial, but that it is due to micro-organisms 

 in the soil was only established comparatively recently. 

 Jodin in 1862 showed that certain micro-organisms (which 

 he called " mycoderms ") could grow in solutions con- 

 taining sugar or tartaric acid but no organic nitrogen, 

 and that if the cultures were allowed to stand in sealed 

 vessels, nitrogen as well as oxygen was removed from the 

 enclosed space. Berthelot later showed that soil which 

 was allowed to stand underwent an increase in nitrogen 

 content, but that this increase did not occur if the soil 

 were first sterilised by heat. Later still he isolated organ- 

 isms which could grow at the expense of atmospheric 

 nitrogen. Much of our knowledge of nitrogen fixation is, 

 however, due to Winogradsky, who began to study the 

 problem in 1893 ; he showed that an anaerobe, CL 

 pastoriaiium, very closely related to CL butyricum which 

 also fixes atmospheric nitrogen, was capable of 

 growth on synthetic media and could derive its 

 nitrogen from the atmosphere. The fixation of nitrogen 

 was proportional to the amount of glucose fermented ; for 

 each 1 gram of glucose destroyed (with formation of 

 butyric and acetic acids together with carbon dioxide 

 and hydrogen) approximately 2-5 mg. of nitrogen was 

 fixed. If other sources of nitrogen, such as ammonium 

 salts, were present fixation of atmospheric nitrogen 

 ceased and the nitrogen of the ammonium salts was used 

 preferentially. Winogradsky suggested that the nitrogen 

 was reduced with formation of ammonia by nascent 



