1919] 



ALLEN — AZOTOBACTER CHROOCOCCUM 9 



trality at incubation temperatures. 1 The addition of calcium 

 carbonate, therefore, to Beijerinck's solution introduces two 

 variables : It changes its reaction and it precipitates the phos- 

 phates. It is not surprising, therefore, that the tendency has 

 been to omit calcium carbonate in Beijerinck's solution, es- 

 pecially when mannite is used as the energy source. 2 In Ash- 

 by 's solution more soluble calcium (as sulphate) is added than 

 in Beijerinck's solution, hence more of the phosphate is car- 

 ried down, with the result that the solution is very poorly 

 buffered and hence unsuited to the growth of Azotobacter 

 (especially when dextrose is used), unless calcium carbonate 

 be supplied to each culture vessel. Although direct compari- 

 sons are few or wanting entirely, it seems from the literature 



that Beijerinck's solution is preferable to Ashby's. In the 

 light of the above reasoning it may easily be that this is be- 

 cause it contains more soluble phosphate than does Ashby's 

 and that this, besides being more available as a nutrient, acts 

 as a soluble buffer and as such is more effective than the solid 

 calcium carbonate. 



Now it was conceivable to us that the phosphorus nutrition 

 and possibly the maintenance of proper hydrogen ion concen- 

 tration separately or in conjunction were operative in all the 

 above-cited cases of growth stimulation of Azotobacter chroo- 

 coccum. For instance, in Krzemieniewski 's experiments the 

 organic combinations of calcium may have supplied this ele- 

 ment in an assimilable form and organic phosphates in the 

 humus may also have been beneficial. The colloids which 

 Sohngen used might have acted as protective colloids, i.e., pre- 

 vented the complete flocking out of the tricalcium phosphate 

 after it was once formed. The properties of soil grains which 

 he believed was due to adsorption of gases may have been due 

 to their adsorbed phosphate ions, or possibly to the difficultly 

 soluble phosphate compounds precipitated as a thin film as a 



1 The exact P H value is influenced by the mannite or the dextrose present. 



3 When dextrose is added it, of course, shifts the reaction slightly toward 

 the acid side, because of its properties as a weak acid. (See Mathews, A. P. 

 Physiological Chemistry, p. 32. 1916.) 



