Generation of Free Radicals from Model Lipid Hydroperoxides and H₂O₂ by Co(II) in the Presence of Cysteinyl and Histidyl Chelators

Electron spin resonance spin trapping was utilized to investigate the generation of free radicals from cumene hydroperoxide (cumene-OOH), tert-butyl hydroperoxide (tert-butyl-OOH), and H₂O₂ at pH 7.2 by Co(II) in the presence of cysteinyl and histidyl chelating agents. The spin trap used was 5,5-dimethyl-1-pyrroline N-oxide. Incubation of Co(II) with cumene-OOH or tert-butyl-OOH did not generate any detectable amounts of free radicals. However, in the presence of glutathione, cysteine, penicillamine, or N-acetylcysteine, Co(II) generated cumene-OOH-derived carbon-centered radicals, cumene alkoxyl radicals, and hydroxyl (^•OH) radicals. Oxidized glutathione and cystine used instead of reduced glutathione or cysteine did not generate any free radical, indicating an important role of the -SH group in radical generation. While the addition of diethylenetriaminepentaacetic acid (DTPA) prevented radical generation, deferoxamine had only a slightly inhibitory effect. Similar results to those obtained using cumene-OOH were obtained utilizing tert-butyl-OOH in place of cumene-OOH. The yields of free radicals were in the order of glutathione > cysteine > penicillamine > N-acetylcysteine. Incubation of Co(II) with cumene-OOH or t-butyl-OOH in the presence of the histidyl oligopeptide Gly-Gly-His also generated lipid hydroperoxide-derived free radicals, with the yield being comparable to that obtained using thiols. In contrast, histidine, anserine, homocarnosine, or carnosine did not cause any free radical generation from Co(II) and lipid hydroperoxides. Incubation of Co(II) with H₂O₂ produced only a small amount of ^•OH radicals. Addition of glutathione to the mixture of Co(II) and H₂O₂ resulted in generation of both glutathionyl (GS^•) and ^•OH radicals, which could be inhibited by DTPA and deferoxamine. Deferoxamine nitroxide radical was produced from deferoxamine incubated with Co(II) and H₂O₂. Under the same experimental conditions, cysteine, penicillamine, and N-acetylcysteine inhibited free radical generation from the reaction of Co(II) with H₂O₂. Histidine and histidyl oligopeptides, homocarnosine, and carnosine did not have a significant effect. However, anserine enhanced the ^•OH radical generation from this reaction. The results indicate that Co(II) is capable of generating free radicals from lipid hydroperoxides and H₂O₂ in the presence of proper chelating agents, which may be relevant to the mechanism(s) of Co(II)-related toxicity and carcinogenicity.