Optimum reaction conditions for human lactate dehydrogenase isoenzymes as they affect total lactate dehydrogenase activity.

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° (pH 30 ) substrate concentration, and coenzyme concentration for the human LDH isoenzymes. For the P → L procedure, broad pH 30 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 LDH 1 to 1.7 x 10 -3 M for LDH 5 at pH 30 7.30. For the L → P procedure, the pH 30 optima were within the range of 8.30-8.88 for the LDH 5 through LDH 1 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 LDH 1 to at least 7.2 x 10 -2 M for LDH 5 . 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.

[1]  R. Mccomb,et al.  A comparison of reduced-NAD preparations from four commercial sources. , 1968, Clinical chemistry.

[2]  Arthur F. Krieg,et al.  Lactate dehydrogenase isoenzymes a comparison of pyruvate-to-lactate and lactate-to-pyruvate assays. , 1967, Clinical chemistry.

[3]  E. Vesell.  pH Dependence of Lactate Dehydrogenase Isozyme Inhibition by Substrate , 1966, Nature.

[4]  H. Barnett.  The staining of lactic dehydrogenase isoenzymes after electrophoretic separation on cellulose acetate , 1964, Journal of clinical pathology.

[5]  E. Amador,et al.  SERUM LACTIC DEHYDROGENASE ACTIVITY: AN ANALYTICAL ASSESSMENT OF CURRENT ASSAYS. , 1963, Clinical chemistry.

[6]  R. Bartlett.  Rapid cellulose acetate electrophoresis. I. Serum proteins. , 1963, Clinical chemistry.

[7]  C. Artom.  An introduction to diagnostic enzymology. , 1963 .

[8]  D. Amelung.  [Serum enzyme determinations in pernicious anemia]. , 1960, Deutsche medizinische Wochenschrift.

[9]  J. Dreyfus,et al.  SERUM ENZYMES IN THE PHYSIOPATHOLOGY OF MUSCLE , 1958, Annals of the New York Academy of Sciences.

[10]  G. Weber,et al.  [Lactic dehydrogenase activity in blood & blister fluid in dermatoses]. , 1958, Dermatologische Wochenschrift.

[11]  E. Vesell,et al.  Localization of Lactic Acid Dehydrogenase Activity in Serum Fractions , 1957, Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine.

[12]  B. Hill,et al.  Elevation of a serum component in neoplastic disease. , 1954, Cancer research.

[13]  P. Strandjord,et al.  An automated method for the determination of total lactate dehydrogenase, heat-stable lactate dehydrogenase isozyme-1, and hydroxybutyrate dehydrogenase. , 1966, The Journal of laboratory and clinical medicine.

[14]  U. Moeser.  LACTIC DEHYDROGENASE. , 1964, The Journal of the Maine Medical Association.

[15]  R. J. Henry,et al.  Revised spectrophotometric methods for the determination of glutamic-oxalacetic transaminase, glutamic-pyruvic transaminase, and lactic acid dehydrogenase. , 1960, American journal of clinical pathology.