Optimum Reaction Conditions for Human Lactate Dehydrogenase Isoenzymes as They Affect Total Lactate Dehydrogenase Activity

[摘要] Optimum reaction conditions at 30° ± 0.5° for two continuous spectrophotometric assay procedures, lactate to pyruvate (L → P) and pyruvate to lactate (P → L), were determined with respect to pH at 30° (pH30) substrate concentration, and coenzyme concentration for the human LDH isoenzymes.For the P → L procedure, broad pH30 optima were within the range of 7.20-7.40 for all the LDH isoenzymes. The coenzyme optima were identical for all of the isoenzymes tested at a reduced NAD concentration of 1.5 x 10-4 M. Pyruvate substrate optima ranged from 7.5 x 10-4 M for LDH1 to 1.7 x 10-3 M for LDH5 at pH30 7.30.For the L → P procedure, the pH30 optima were within the range of 8.30-8.88 for the LDH5 through LDH1 isoenzymes, respectively. Optimum activity was obtained at a NAD concentration of 6.0 x 10-3 M and remained constant at least to 1.8 x 10-2 M for each of the isoenzymes tested. L-Lactate substrate optima ranged from 4.0 x 10-2 M for LDH1 to at least 7.2 x 10-2 M for LDH5.From the isoenzyme studies, the degree of variation possibly involved in either method due to variations in isoenzyme distribution was calculated for total LDH samples. These calculations showed that both methods, P → L and L → P, were essentially equivalent. This equivalency was verified by a comparative study of the two methods on human serum samples.

[学科分类] 过敏症与临床免疫学