86 



FISHERY BULLETIN OF THE FISH AND WILDLIFE SERVICE 



due to contamination from the observed cell- 

 activity count. Of the total radioactive phos- 

 phorus in the medium, about 2.5 percent was 

 exchanged with the cells. There was no detect- 

 able change in the inorganic-phosphorus concen- 

 tration of the medium. 



Table 6. — Uptake of radioactive phosphorus by cells from 

 medium when kept in the dark 



[The medium contained 9,887 counts per minute] 



Exchange determined with radioactive phosphorus 

 in the cells. — Since a tendency for the cells to 

 clump in the previous experiment possibly caused 

 one determination to be low, this experiment was 

 devised with the active phosphorus inside the 

 cells so that determination could be made on the 

 medium. To test for the loss of radioactive 

 phosphorus from the cells through exchange, a 

 culture of 40 X 10 7 cells per liter was prepaied 

 with 1 microcurie of radioactive phosphorus per 

 100 cc. of medium and less than 0.5^gAP/L. 

 After 16 hours the culture was centrifuged, and 

 the cells containing radioactive phosphorus were 

 suspended in new medium containing 56 MgAP/L 

 of nonradioactive phosphorus. This culture was 

 placed in the dark and samples were removed 

 periodically. There was a loss by exchange of 

 radioactive phosphorus from the cells to the 

 medium (table 7). Of the total radioactive 

 phosphorus in the cells, 3.26 percent was exchanged 

 with the medium. 



Table 7. — Loss of radioactive phosphorus to medium by cells 

 kept in the dark 



[The cells from 25 cc. of medium contained 22,570 counts per minute] 



DISCUSSION 



Some investigators have found it necessary to 

 wash the cells before using them for chemical 

 analysis. According to Gest and Kamen (1948), 

 the washing liquid is usually distilled water, 

 0.85 percent saline, or less frequently, fresh 

 culture medium. Washing with a saline solution 

 is less drastic than with distilled water. Even 

 so, many organisms have been found to lose 

 phosphorus when washed with saline solutions. 

 A young culture of Bacterium coli, after two wash- 

 ings with a saline solution, showed a rapid release 

 of inorganic phosphorus in the absence of glucose 

 (Macfarlane 1939). Also an appreciable quantity 

 of the original phosphorus of Trypanosoma equip- 

 erdum is lost when the organisms are washed 

 with saline solution (Moraczewski and Kelsey 

 1948). It must be pointed out, however, that this 

 ,type of experiment does not demonstrate exchange. 

 For phosphorus exchange to occur, there must be a 

 movement of equal numbers of phosphorus atoms 

 both into and out of the cells. This naturally 

 precludes interpreting the above observations as 

 being exchange, since neither distilled water nor 

 saline solutions ordinarily contain phosphorus. 



Fresh-water algae grown in high-phosphate 

 media have been shown to store an appreciable 

 quantity of soluble phosphate which is readily 

 lost when the cells are washed with medium 

 (Gest and Kamen 1948). Also marine species 

 of algae exchange phosphate to a nutrient medium 

 when filter- washed, as has been shown by Gold- 

 berg, et al. (1951) and in the experiments reported 

 in this paper. Even with the use of radioactive 

 phosphorus which makes possible an altering of 

 the isotopic distribution of phosphorus in either 

 the medium or the cells, exchange is still difficult 

 to demonstrate, especially in rapidly dividing 

 unicellular forms. This is because the amount 

 of phosphorus absorbed by the cells is many times 

 greater than that leaving the cells through 

 exchange. 



Goldberg, et al. (1951) stated that as much as 

 50 percent of the phosphorus is removed from the 

 marine diatom Asterionella when it is washed 

 with sea water; but the factors controlling this 

 removal of radioactive phosphorus from the cells 

 were not discussed. In the filter-washing ex- 

 periments conducted in this investigation, 

 Nitzschia cells were always washed with a culture 

 medium containing the same concentration of 



