The purpose of the present study was to evaluate the mechanism of microbial resistance to oxidative stress induced by photolysis of hydrogen peroxide (H2O2) in relation to microbial catalase activity. In microbicidal tests, Staphylococcus aureus and Candida albicans were killed and this was accompanied by production of hydroxyl radicals. C. albicans was more resistant to hydroxyl radicals generated by photolysis of H2O2 than was S. aureus. A catalase activity assay demonstrated that C. albicans had stronger catalase activity; accordingly, catalase activity could be one of the reasons for the resistance of the fungus to photolysis of H2O2. Indeed, it was demonstrated that C. albicans with strong catalase activity was more resistant to photolysis of H2O2 than that with weak catalase activity. Kinetic analysis using a modified Lineweaver-Burk plot also demonstrated that the microorganisms reacted directly with hydroxyl radicals and that this was accompanied by decomposition of H2O2. The results of the present study suggest that the microbicidal effects of hydroxyl radicals generated by photolysis of H2O2 can be alleviated by decomposition of H2O2 by catalase in microorganisms.