234 



FISHERY BULLETIN OF THE FISH AND WILDLIFE SERVICE 



— i 1 



NITZSCHIA 



~[ r~ 



2 3 4 



TIME IN DAYS 



3 4 



TIME IN DAYS 



Figure 8. — Comparison of pH changes in unbuffered, buf- 

 fered, and aerated culture media. Dots represent pH 

 changes in unbuffered culture medium ; circles, pH 

 changes in medium buffered with .001 molar KH 2 POj 

 and RHPO,; X's, pH changes in aerated medium. 



procedure used in previous experiments for the 

 preparation of cultures was followed. At the end 

 of each day for 7 days, cell counts and a pH de- 

 termination were made. 



The greatest pH concentration reached in any 

 of the buffered and aerated cultures during the 

 first 4 days of the experiment was 7.7 (fig. 8, A, 

 B, and C) and it was not exceeded during the fifth, 

 sixth, and seventh days in the Nitzschia culture 

 v fig. 8, B). In the buffered flasks a pH of 9 

 was reached on the seventh day in the Ohlorella 

 culture, while in the mixed cultures the pH in- 

 creased to only 8.5 on the sixth and seventh days. 

 ChloreUa cells growing alone had a higher rate 

 of growth than the ChloreUa cells in mixed cul- 

 ture. When compared with the results of the 



previous experiments, there was no significant dif- 

 ference in the total population sizes or the division 

 rates for either ChloreUa or Nitzschia when grown 

 alone, and a similar inhibition of ChloreUa oc- 

 curred in the mixed cultures as in the previous 

 experiments. Therefore pH can be eliminated as 

 the factor responsible for the observed inhibition 

 of growth of ChloreUa in the mixed cultures. 



In aerated medium : 



Another experiment was run in which the pH 

 was prevented from fluctuating as widely as in 

 unbuffered medium by bubbling air through the 

 culture medium during the period of illumination. 

 In addition to preventing as great a fluctuation 

 in pH, this method also furnished a larger supplv 

 of carbon dioxide for photosynthesis. 



In this experiment, seven flasks were eacu 

 fitted with a rolled cotton-wool plug through 

 which an air inlet tube extended to the bottom 

 of the flask so that air could be bubbled through 

 the culture medium. A cotton filter, consisting of 

 a calcium chloride drying tube filled with cotton- 

 wool, was attached to the air inlet tube of each 

 flask and the entire apparatus was autoclaved. 

 ChloreUa cultures were prepared in each flask 

 with a concentration of 70 million cells per liter. 

 The air from an air pump was run into a jar 

 fitted with a rubber stopper containing seven out- 

 let tubes arranged in a circle around the inlet 

 tube. This apparatus was placed on the illumi- 

 nated shelves and one flask of ChloreUa was at- 

 tached to each outlet tube. At the end of each 

 clay for 7 days one culture was removed. A flask 

 containing 100 cc. of distilled water was used to 

 replace the culture in order that the remaining 

 cultures would continue to receive a constant 

 supply of air. Cell counts and a pH determina- 

 tion were made each day for the culture removed 

 from the apparatus. This experiment was re- 

 peated using Nitzschia cultures with an initial 

 concentration of 10 million cells per liter. The 

 experiment was run a third time using a mixed 

 culture of ChloreUa and Nitzschia in the same 

 respective initial concentrations as before. 



ChloreUa grown alone under the conditions of 

 this experiment reached a population size approx- 

 imately twice that obtained in previous experi- 

 ments. While the ChloreUa population showed a 



