Lactic Acid Assay with L(+)lactic Acid Dehydrogenase from Rabbit Muscle

Abstract

Abstract L undholm , L., E. M ohme ‐L undholm and N. V amos. Lactic acid assay with L(+)lactic acid dehydrogenase from rabbit muscle. Acta physiol. scand. 1963. 58. 243–249. — Lactate assay with L(+)lactic acid dehydrogenase from rabbit muscle and with DPN involved the problem of quantitatively reducing lactate to pyruvate. The problem was overcome by use of a solution with a high pH, by an excess of DPN, and by addition of a carbonyl reagent e.g. semicarbazide or hydrazine, which trapped the formed pyruvate. Hydrazine was found to be preferable to semicarbazide since it appeared to bind pyruvate more effectively. It was thus possible to carry out the reaction, with a 100 per cent recovery, at pH = 9.0. This was advantageous because, at pH > 9.0, hydrazine and semicarbazide rapidly formed products with DPN that absorbed at 340 mμ, giving a high blank value that varied greatly with the pH of the solution and seriously affected the precision of the method. Unlike hydrazine, moreover, semicarbazide reacted with formed DPNH in the presence of atmospheric oxygen, so that the extinction of DPNH decreased.