THE EFFECTS OF PHOSPHATE-FREE AND SUBSTRATE-FREE INCUBATION CONDITIONS ON GLYCOLYSIS IN EHRLICH ASCITES TUMOR-CELLS

The influence of phosphate-free medium buffered with synthetic organic buffers, and of a preliminary incubation of cells in medium lacking a dded substrate ('pre-incubation') was investigated with mouse-cultured Ehrlich ascites tumour cells. In comparison to phosphate-containing b icarbonate-buffered balanced-salts medium, organic-buffered medium, wi thout a preliminary substrate-free pre-incubation, was associated with 20-30% reduction in the rate of glycolysis, the 3- to 4-fold accumula tion of fructose 1,6-bisphosphate and the halving of both ATP and tota l adenine nucleotide levels. These perturbations were reversed by the inclusion of 5 mM sodium phosphate in the organic-buffered medium. Pre -incubation for up to 90 min, before inclusion of glucose, resulted in greater depression of the glycolytic rate and concentrations of adeni ne nucleotides. This: occurred in both the balanced-salts medium and t he organic-buffered medium. During pre-incubation cells were lysed, re leasing lactate dehydrogenase. when physically agitated too vigorously . It was concluded that the use of phosphate-free medium and pre-incub ation are not advisable procedures for routine metabolic investigation s with this cell line.