Electrochemical study of hydrogen peroxide formation in isolated mitochondria

Mitochondrial respiration generates reactive oxygen species that are involved in physiological and pathological processes. The majority of methods, with exception of electron paramagnetic resonance, used to evaluate the identity, the rate and the conditions of the reactive oxygen species produced by mitochondria, are mainly based on oxidation sensitive markers. Following latest electrochemical methodology, we implemented a novel electrochemical assay for the investigation of aerobic metabolism in preparations of isolated mitochondria through simultaneous measurement of O₂ consumption and reactive species production. This electrochemical assay reveals active H₂O₂ production by respiring mouse liver mitochondria, and shows that ATP synthase activation and moderate depolarization increase the rate of H₂O₂ formation, suggesting that ATP synthesizing (state 3) mitochondria might contribute to oxidative stress or signaling.