260 AZOFICA TION 



agin, and casein to culture media had only a slight influence on the 

 fixation of nitrogen by Azotobacter. 



Caffeine, alloxan, betain, trimethylamin, legumin, cinnamic acid, 

 aspartic acid, asparagin, hippuric acid, creatin, creatinin, xanthin, 

 and hypoxanthin, are all toxic to Azotobacter even in small quantities. 

 Only the first two have been tested in concentrations dilute enough 

 to stimulate, which is remarkable, as many of these compounds 

 stimulate the higher plants and some can be. utilized directly by 

 the plant. 



Esculin, vanillin, daphnetin, cumarin, pyrocatechin, heliotropin, 

 arbutin, resorcin, pyrogallol, phloroglucin, hydroquinon, salicylic 

 aldehyd, oxalic acid, quinic acid, dihydrostearic acid, rhamnose 

 and borneol, on the other hand, do not stimulate in any concentra- 

 tion. Nor are they toxic until fairly large quantities have been 

 added. In this regard the nitrogen-fixing organisms appear to differ 

 widely from the nitrifying bacteria and higher plants. The resist- 

 ance of the nitrogen-fixers to various chemicals has likewise beer- 

 called to our attention by Lipman in his study of the influence of 

 alkalies on nitrogen-fixation. 



Influence of Colloids. It was recognized early in the study of 

 nitrogen-fixation that when sterilized soil is added to a nutritive 

 medium it greatly increased the quantity of nitrogen fixed. This 

 condition is due to several factors and is partly explained by 

 Krzemieniewski's results wherein he found that nitrogen-fixation is 

 decidedly increased by the addition of soil humus, either as free 

 humic acid or as salts of potassium, sodium or calcium. Kaserer 

 maintains that this is due to the inorganic nutrients, especially to 

 aluminum and silicic acid supplied to the microorganisms through 

 the humus. This is probably true in part, for the fixation varies with 

 the humus derived from different sources. Moreover, artificial 

 humus, prepared by boiling sugar with acids, fails to stimulate. 



That much of the beneficial effect is due to the constituents in the 

 humus appears likely from the results obtained by Sohngen who 

 found that colloidal iron oxid, aluminum oxid, and silicon oxid all 

 greatly stimulated the nitrogen-fixing powers of Azotobacter chroo- 

 coccum. This he attributed to the absorption of oxygen and nitrogen 

 by the colloid, which he maintains would make them more readily 

 available to the organisms. The boiling of natural humus with 

 hydrochloric acid would either remove the foreign material or change 

 it from the colloidal form, and thus, as has been found to be the 

 case, render it inert. Lohnis and Green take exception to this 

 explanation, for they found no adsorptive action exerted by humus 

 on either the nitrogen or the oxygen. Furthermore, Rosing found 

 that he could stimulate just as effectively with iron as with humic 

 acids. But much larger quantities of colloidal iron are required 

 when it is used singly than when used in conjunction with an organic 



