Requirement of a diperoxovanadate-derived intermediate for the interdependent oxidation of vanadyl and NADH

Oxygen release accompanying oxidation of vanadyl by diperoxovanadate was suppressed on addition of NADH. The added NADH was rapidly oxidized, oxygen in the medium was consumed, and the reaction terminated on exhaustion of either NADH or vanadyl. The consumption of oxygen and disappearance of NADH needed small concentrations of diperoxovanadate to initiate and increased with increase in the concentration of vanadyl and NADH or decrease of pH. The products of the reaction were found to be NAD+ from NADH and vanadate oligomers from vanadyl and oxygen. The reaction was insensitive to catalase and was not dependent on H2O2. The reaction was inhibited by superoxide dismutase, cytochrome c, EDTA, Mn2+, histidine, and DMPO, but not by hydroxyl radical scavengers such as ethanol and benzoate. The ESR spectrum of the reaction mixture showed the presence of the 1:2:2:1 quartet signal typical of a DMPO-OH adduct, but this was not modified by ethanol. This oxygen radical species, possibly of .OV type derived from diperoxovanadate, is proposed to have a role in the reactions of oxygen release and NADH oxidation.