Assay of Rat Plasma Pyruvate Kinase Activity with Luciferin-Luciferase

Abstract

An assay method for pyruvate kinase in rat plasma is described. Plasma samples were incubated with ADP and phosphoenolpy-ruvate in Tris buffer solution. The ATP produced by pyruvate kinase was measured by photocounting after the addition of a commercially available luciferin-luciferase preparation. Interference by ATP or adenylate kinase originally present in the sample was removed by a high degree of dilution. The assay is sensitive, reproducible, and rapid, especially when used for large numbers of samples. By this method, pyruvate kinase activity in normal rats was determined to be 0.51±0.05 (n=6) U/ml plasma. In rats fed a vitamin E-deficient basal diet for 7, 10, or 14 weeks, pyruvate kinase activities were 0.70±0.11, 1.64±0.51, and 4.28±0.85 (n=6) U/ml plasma, respectively. This method appears to be useful for the determina-tion of pyruvate kinase activity in nutritional or pharmacological studies.