[Vor. 1 
180 ANNALS OF THE MISSOURI BOTANICAL GARDEN 
than on any of the organic-compound-containing media. Glu- 
cose, вассһатове, and mannite were chosen as energy-furnishing 
compounds because of their general usefulness in this capacity 
among free-nitrogen-fixing bacteria, and also because they are 
representatives from three great classes of carbon compounds. 
Certain unpublished experiments, carried out by B. M. Dug- 
gar on nitrogen fixation in the fungi, indicate that, whereas no 
fixation takes place at ordinary temperatures, it does take place 
at elevated temperatures. It was thought desirable, therefore, 
to investigate the effect of elevated temperature on the process 
of elementary-nitrogen fixation by alge in the absence of 
combined nitrogen. However, the results tabulated in table 
пт show clearly that not only did no growth on any nitro- 
gen-free medium appear at the higher temperature, but also 
that that appearing on nitrogen-containing media was, in many 
cases, poorer than that obtained in cultures kept at ordinary 
temperatures. It should further be noted that growth was in 
some cases entirely suppressed. It would appear, therefore, that 
in the species investigated, growth at elevated temperatures is 
less vigorous than at ordinary temperatures and that, in all 
probability, no favorable effect on free-nitrogen fixation is to be 
expected by growing these species at the higher temperature 
maintained in the experiment. 
The incipient, ephemeral growth which was observed in a 
few cases where combined nitrogen was not furnished is be- 
lieved to be due to the minute quantity of combined nitrogen 
which was unavoidably introduced in the inoculation process. 
The inoculating material was, of necessity, derived from agar 
containing ammonium nitrate, and while no agar was trans- 
ferred it is altogether probable that enough combined nitrogen 
was carried over in the water adhering to the cells to account for 
the trace of growth. It should be emphasized again that in 
every case this growth was so slight as to have escaped detec- 
tion had not a comparison been made with a flask absolutely 
free from growth. 
In table ту the results of the two experiments are combined 
and show that in seven species complete results have been ob- 
tained. These results indicate with perfect uniformity that 
growth, under the conditions realized in the experiments, is 
impossible in the absence of combined nitrogen, even when readily 
