FIXATION OF ATMOSPHERIC NITROGEN 575 



tation. That complex is later used partly for energy and partly for 

 synthesis of protoplasm. It was actually demonstrated 64 that a large 

 percentage of the total nitrogen fixed in the first few days of growth 

 of Azotobacter consists of amino acid nitrogen. This proves that the 

 elementary nitrogen goes through the simple organic forms before it is 

 changed into protein. It also tends to prove that nitrogen is fixed by 

 combination with hydrogen and not with oxygen, thus insuring much 

 greater economy of energy. 



Influence of available nitrogen compounds upon nitrogen fixation. The 

 nitrogen fixing bacteria do not depend entirely upon atmospheric nitro- 

 gen for their need, but are capable of utilizing combined nitrogen present 

 in the medium. This is a direct result of the distinct difference in growth 

 and nitrogen-fixation pointed out above. Nitrates are readily utilized 

 by Azotobacter as sources of nitrogen; 65 accordingly the presence of 

 nitrates in the medium inhibits the fixation of atmospheric nitrogen. 

 Ammonium sulfate and peptone are also available nitrogen compounds. 66 



The injurious action of the nitrate upon the fixation of nitrogen 

 was considered to be due either to (a) a direct toxic action of the salt 

 upon the growth of the organism, (b) stimulation by nitrate of other 

 organisms in a mixed culture which are antagonistic to Azotobacter, 

 and (c) competition by such organisms with Azotobacter for the energy 

 supply. 



By taking the number of organisms developing and the nitrogen fixed 

 in the nitrate-free cultures as 100, it was found 67 that the addition of 50 

 mgm. of NaN0 3 brought about an increase in the number of organisms 

 from 100 to 3150, while the nitrogen fixed was increased 342 per cent in 

 sterilized soil and 500 per cent in unsterilized soil. The presence of ni- 

 trate was thus found to stimulate greatly the multiplication of the Azo- 

 tobacter organism, while it reduced its physiological efficiency. Bon- 

 azzi, 68 therefore, suggested that Az. chroococcum may fix nitrogen in the 



64 Waynick, D. D., and Woodhouse. By what steps does Azotobacter fix 

 nitrogen? Cal. Agr. Exp. Sta. Ann. Rpt. 1918-19, p. 62-63. 



65 Lipman, 1903 (p. 115); Pringsheim, H. Zur Stickstoffassimilation in Gegen- 

 wart von Saltpeter. Centrbl. Bakt. II, 40: 21-24. 1914; Hanzawa, J. Einige 

 Beobachtungen fiber Stickstoffbindung durch Azotobacter in stickstoffarmen 

 und in stickstoffreichen Substanzen. Centrbl. Bakt. II, 41: 573-576. 1913. 



66 Krainsky, 190S-1912 (p. 566); Prazmowski, A. Azotobacter-Studien. II. 

 Physiologie und Biologic Bull. Intern. Acad. Sci. Cracovie, Math. Nat. CI. Sc. 

 B, No. 7: 855-950. 1912. 



67 Hills, T. L. Influence of nitrates on nitrogen assimilating bacteria. Jour. 

 Agr. Res., 12: 183-230. 1918. 



68 Bonazzi, 1921 (p. 567). 



