Lactate-mediated enzymatic reduction of metmyoglobin in vitro

This study examined the lactate-mediated enzymatic reduction of equine metmyoglobin (MMb) by using various lactate salts (sodium, potassium and calcium) and bovine m. triceps brachi extracts in vitro. The equine MMb was reduced by the assay mixture containing lactate, NAD+, and the muscle extract at pH 5.7. In the absence of the bovine muscle extract, NAD+ or l-lactate in the assay mixture, MMb reduction was not observed. Replacing l-lactate with d-lactate or adding oxalate (lactate dehydrogenase inhibitor) to the assay mixture resulted in no MMb reduction, suggesting that lactate dehydrogenase is involved in the MMb reduction, interacting with l-lactate and NAD+. An increase (P < 0.05) in the rate of MMb reduction was also observed with an increase of the amount of bovine muscle extract, NAD+ or lactate concentration in the assay mixture. There was no difference (P > 0.05) in the rate of MMb reduction when potassium lactate was added to the assay mixture instead of sodium lactate. The MMb reduction through the lactate–NAD+-muscle extract system was increased (P < 0.05) with an elevation in pH from 5.1 to 6.1. The results of this research confirm that the inclusion of l-lactate can induce the MMb reduction through the lactate– NAD+-enzyme coupling interaction.