We examined hydroxyl free radical (∙OH) production in the mixture of H2O2 and N-methyl-N′-nitro-N-nitrosoguanidine (MNNG) without exposure to light using the electron spin resonance spin-trapping technique. When the mixtures were protected from exposure to light, ∙OH was formed at pH 6.5 and above; it was not formed at pH 5.0 and below, consistent with our previous report. The amount of ∙OH trapped depended on the concentrations of MNNG and H2O2 and the pH. Nitrite ion was also detected colorimetrically at pH 6.5 and above, but not detected at pH 5.0 and below in the mixtures without exposure to light. Moreover, its production depended on the concentrations of MNNG and H2O2 and the pH. The formation of N-methyl-N′-nitroguanidine in the mixture at pH 7.8 was confirmed by thin-layer chromatography and melting point. These results suggest that nucleophilic attack by H2O2 on the nitroso nitrogen of MNNG results in the formation of N-methyl-N′-nitroguanidine and peroxynitrous acid, which degrades homolytically to yield ∙OH and nitrogen dioxide, resulting in the production of nitrite ion, at pH 6.5 and above without exposure to light.Key words: N-methyl-N′-nitro-N-nitrosoguanidine, hydrogen peroxide, hydroxyl free radical, peroxynitrous acid.
Study of the mechanisms of hydrogen peroxide and hydroxyl free radical-induced cellular injury and calcium overload in cardiac myocytes
