604 T. J. MURRAY 



NITROGEN FIXATION 



The ability of microorganisms to fix atmospheric nitrogen was 

 first definitely demonstrated to be due to an anaerobic bacillus, 

 B. Pasteurianus, in 1893 by Winogradski. It remained for 

 Beyerinck in 1901 to demonstrate an aerobic organism that also 

 assimilated free nitrogen. Non-symbiotic fixation of nitrogen 

 in the soil is due to both types of organisms but at times may 

 be due only to one type. Lipman and Burgess (1915) found 

 two-thirds of the soils examined by them free from Azotobacter. 

 Yet these soils were capable of fixing nitrogen when inoculated 

 into solutions. They ascribed the nitrogen fixation to Clostri- 

 dium forms. Haselhoff and Bredemann (1906) investigated an- 

 aerobic nitrogen-collecting bacteria and found results approxi- 

 mating those of Winogradski. The amount of nitrogen fixed 

 varies with the amount of carbonaceous matter present, the more 

 carbon the higher the assimilation. Working with pure and 

 mixed cultures, they found from 0.42 to 2.74 mgm. of nitrogen 

 fixed per gram of mannite. Lipman (1908), working with pure 

 cultures of Azotobacter, found from 0.39 to 10.45 mgm. of nitrogen 

 per gram of mannite formed in four weeks in mannite solution. 

 In this work mass cultures were used, either by inoculating solu- 

 tion with soil or by adding the source of carbon to the soil. 



In my work nitrogen fixation was carried out in soils and in 

 solution. For the solution work, 100 cc. of a nitrogen poor 

 medium was inoculated with from 2 to 5 grams of soil. The 

 following solution (N. J. 1908) was used: 



H2O 1000 . grams 



K2HPO4 0.2 gram 



MgS04 0.2 gram 



NaCl 0.5 gram 



Mannite 20.0 grams 



FeCla 1 drop of 10 per cent solution 



The solution was neutralized with KOH using phenolphthalein 

 as an indicator. Anaerobic conditions were obtained by adding 

 about an inch of sterile paraffin oil or by placing the material 

 in the anaerobic apparatus described under the ammonification 

 experiments. 



