178 Nitrogen Fixation by Yeasts 



acid used in the nitrogen determinations, we find a much higher 

 blank here, but several blanks were analyzed and the close 

 agreement between them showed the results above given to be 

 trustworthy. It should be mentioned here that the sterile blanks 

 in all these series were not merely analyzed from the original solu- 

 tion but were inoculated like the cultures, then sterilized and incu- 

 bated side by side with the cultures for the same length of time. It 

 is interesting to note here that the amount of visible growth cannot 

 be directly correlated with the amount of nitrogen fixed, for sev- 

 eral of the cultures which appeared to have made only a small 

 amount of growth showed quite a considerable fixation of nitrogen. 

 I presume that this has been observed by other investigators work- 

 ing on this problem and is probably due to the fact that some 

 of the nitrogen compounds produced are soluble and diffusible and 

 therefore give no visible evidence of their presence. It is hardly 

 necessary to add here that despite the favorable constitution of 

 the medium employed it is not nearly so favorable for the growth 

 of the organisms tested as the beer wort in which the stock cul- 

 tures were kept as can be noticed particularly in the cultures of 

 Aspergillus niger and Penicillium glaucum where the membranes 

 formed in the dextrose solution are very thin and light in color and 

 the spore production much smaller, than in the beer wort cultures. 



Series III. 



As explained above the culture solutions employed in the pre- 

 ceding series were prepared with tap water and the necessary sugar 

 and salts added. It appeared to the writer that a more rigid 

 test of the power to fix nitrogen possessed by the organisms in 

 question should be made. It seemed desirable to see if, like the 

 nitrogen fixing bacteria of the Azotobacter group, they had the 

 power to fix nitrogen in nitrogen-free solutions or solutions which 

 are practically nitrogen-free. It was also thought desirable to 

 test the comparative values of mannite and four of the sugars in 

 such nitrogen-free solutions as sources of energy for the organisms. 

 To that end solutions like the one described above were prepared 

 but distilled water, free from ammonia, was substituted for tap 

 water. The salts employed being chemically pure and used only 

 in small quantities as noted, could not contain more than a trace 



