20 



ELECTROLYTES IX BIOLOGICAL SYSTEMS 



the supernatant fluid was measured. The pellet of cells was resuspended in 

 20 ml NaCl, and two i.o-ml samples were removed to measure the total radio- 

 activity taken up by the cells. After the suspension was recentrifuged, the 

 pellet was resuspended in NaCl and aliquots were taken for radioactivity 

 determination. This determination measures the radio-activity retained by 

 the cells after washing. The procedure was repeated at intervals as shown in 

 table 16. 



The radioactivity measured for unwashed cells at lime zero gave an apparent 

 water space of 209%. An equally absurd result would be obtained from the 

 supernatant analysis method since binding of the S'^^ had occurred. This is 

 indicated by the fact that the cells retained radioactivity after washing. Esti- 

 mation of the water space by wash loss measurements are also meaningless 

 since the cystine is degraded to H2S (9). 



Table 16. Uptake of s'*-labeled cystine by e. coli 



Cells briefly immersed in a complete medium containing radiocystine gave 

 more satisfactory water space determinations. One-half ml of cells in the sta- 

 tionary growth phase were suspended in 2 ml of synthetic medium at 3°C. 

 The results obtained are shown in table 17. Very little metabolic uptake was 

 noted and the measured water space was 69%. Ninety-seven per cent of the 

 labeled cystine taken up was removed by the first 5 ml NaCl wash solution. 



b) Uptake of S^^-labeled methionine. These difficulties due to metabolic 

 alteration are not as serious when radiomethionine is used to measure the water 

 space of E. coli since methionine is not as metabolically reactive as cystine 

 under these conditions. The data of table 18 demonstrate that methionine 

 penetrates the E. coli cell boundary by passive diffusion. The water space 

 measured by the supernatant radioactivity gave a value of ^^i^c when 0.56 ml 

 of cells were suspended briefly in 2 ml of complete medium at 2o°C. After sub- 

 tracting the metabolically incorporated radioactivity, the water space calcu- 

 lated was 67% of the cell volume. The wash loss determination of water space 

 was 64%. 



Permeability of Escherichia Coli to C'^-Glutamic Acid. Rapid metabolism of 

 glutamic acid occurs when washed E. coli cells are added to chilled saline solu- 



