1/6 FIXATION OR ASSIMILATION OF NITROGEN 



As in the case of Clostridium, these organisms obtain 

 the energy they need to bring the inert free nitrogen 

 into chemical combination from the combustion of carbon 

 compounds in the soil. The presence of phosphates and 

 lime is necessary for their growth, and possibly potash 

 and other inorganic salts are useful also ; according to 

 Fischer's investigations they die out or become inert in 

 soils containing less than .1 per cent, of lime. All the 

 species of Azotobacter are strongly aerobic and need an 

 abundant supply of air. An adequate amount of 

 moisture is also essential for their welfare, the best results 

 being obtained when the soil contains between 5 to I 5 

 per cent. ; more than this leads to imperfect aeration. 

 They only thrive satisfactorily when nitrogen compounds 

 are present in small amount or absent altogether. t jftiey 

 carry out their work most effectively at a temperature 

 between 25 C. and 30 C., but growth is active at 20 C, 

 and A. Koch states that he founcK nitrogen fixation going 

 on in the soil during winter. At temperatures above 

 30 or 35 C. they produce involution forms and become 

 weakened in vitality and nitrogen-fixing power. To 

 avoid complications with Clostridium pastorianum present 

 in the soil upon which he experimented, Beijerinck 

 used mannite and calcium, sodium or potassium salts of 

 propionic acid as the necessary carbon food for these 

 organisms, since these substances do not readily undergo 

 the " butyric " fermentation. He found Azotobacter also 

 capable of utilizing glucose, levulose, galactose, cane 

 sugar, and salts of several organic acids as sources of 

 carbon. 



The nutrient medium in which the cultures are made 

 is placed in small amounts in wide-bottomed flasks, so as 

 to allow a free access of air to a large surface of the 



