This study demonstrates that aerobic bacteria generate superoxide radicals (O2•−) during normal metabolic respiration, similar to mammalian mitochondria. The researchers tested seven bacterial species by measuring superoxide production using three independent methods: reduction of ferricytochrome c, auto-oxidation of adrenaline, and luminol chemiluminescence. When sucrose or acetate was added to washed bacterial suspensions, all three assays showed evidence of superoxide formation, and exogenous superoxide dismutase inhibited these reactions. Antimycin A, an electron transport chain inhibitor, increased the superoxide production rate approximately 3.4-fold while reducing oxygen consumption, indicating superoxide is generated during initial electron transfer reactions before the antimycin inhibition site. Fractionation studies showed the highest superoxide-generating activity in membrane fractions with high cytochrome content. The superoxide production rate varied 60-fold among bacterial species (0.11 to 6.22 nmol/mg/min), though importantly, bacterial membrane fractions produced superoxide approximately 15-24 times faster than mammalian mitochondria. These findings suggest bacteria produce superoxide as a byproduct of respiratory chain activity and may potentially use it as a defense mechanism.

Key findings

• Multiple aerobic bacterial species generate superoxide radicals during substrate oxidation, detectable by cytochrome c reduction, adrenaline oxidation, and luminol chemiluminescence assays
• Superoxide production is stimulated by antimycin A and correlates with cytochrome content, indicating generation during initial electron transfer chain reactions before the antimycin inhibition site
• Bacterial membrane fractions produce superoxide 15-24 times faster than mammalian mitochondria despite having similar production mechanisms
• Superoxide production rates vary 60-fold across bacterial species, with no significant difference between pigmented and non-pigmented variants of the same species
• Exogenous superoxide dismutase partially inhibits bacterial NADH-dependent oxygen consumption, suggesting superoxide may be released extracellularly or function as a defense mechanism