Dand L-Lactate dehydrogenase reactions increase pH to lactate production and decrease pH to pyruvate production

Abstract

The traditional explanation of metabolic acidosis in extrahepatic tissues is the increased cellular production of lactic acid. We tested whether the LDH reaction produces lactate and pyruvate anions, or acids, and measured the pH changes that occur for either direction of the reaction. In-vitro reagents (25 to 40 mL) of the Dand L-LDH reaction were prepared for lactate and pyruvate production at known substrate concentrations ([Substrate]), no added H+ buffers, at ~25 °C. The pH of the reagent was measured using a glass polymer pH electrode. The initial reagent pH was adjusted to a target pH value by addition of HCl or KOH (0.02 to 0.1 N). The baseline pH was recorded for 2 min, followed by the addition of LDH. The pH response was recorded continuously at varied intervals for 20 min. Reagent samples (100 to 200 μL) were obtained intermittently for measurements of product. The H+ consumption of pyruvate reduction to lactate increased pH (6.3 to 10.5), and the H+ release of lactate oxidation to pyruvate decreased pH (8.8 to 6.9). The [Product] was small for all [Substrate] and LDH activity conditions (~0.3 – 0.8 mmol·L-1), and lowest for pyruvate production. The pH and [Product] results were similar for both Dand L-LDH reactions. The pH to [Product] profiles for lactate vs. pyruvate equated to 9.215 vs. -9.074 pH units.mmol-1.L-1, respectively. Scientists and clinicians are encouraged to use these results to further understand the function of the LDH reaction during perturbations of pH in biological systems.