Changes in the concentration of inorganic phosphate (Pi) sugar phosphate, phosphoenol pyruvate (PEP), the adenylate charge and pH were followed by 31P-NMR and assay of cell extract during glucose fermentation in Brochothrix thermosphacta. These changes provide an estimation of glycolytic flux via pyruvate kinase as opposed to cycling of PEP through the phosphotransferase system. There appears to be a strong causal relationship between the reduced pH, the high NTP level and the apparent reduction in pyruvate kinase activity, which support the contention that synergistic effect of pH and ATP on pyruvate kinase is an important regulatory factor in these cells.