Generation of 5,5-Dimethyl-1-pyrrolineN-Oxide Hydroxyl and Scavenger Radical Adducts from Copper/H2O2Mixtures: Effects of Metal Ion Chelation and the Search for High-Valent Metal–Oxygen Intermediates

Abstract

A metal-catalyzed nucleophilic addition mechanism for the formation of radical adducts of the spin trap 5,5-dimethyl-1-pyrrolineN-oxide (DMPO) has been described recently (K. Makino, T. Hagiwara, A. Hagi, M. Nishi, and A. Murakami, 1990,Biochem. Biophys. Res. Commun.172, 1073–1080; P. M. Hanna, W. Chamulitrat, and R. P. Mason, 1992,Arch. Biochem. Biophys.296, 640–644). In the present investigation, we have demonstrated that the recently reported inhibition of copper-dependent hydroxyl radical formation by the complexing agent 1,10-phenanthroline (OP), which appears to contradict the well-known chemical nuclease properties of CuI(OP)2, is an artifact resulting from an inhibition of the nucleophilic addition of water to DMPO by OP (A. C. Mello-Filho and R. Meneghini, 1991,Mutat. Res.251, 109–113). Copper bound to OP was found to be a good catalyst of hydroxyl radical formation: the CuII(OP)2complex can be reduced by H2O2and the CuI(OP)2generated reacts with the peroxide to form[formula]In contrast, no evidence could be obtained for oxidant formation from the[formula]reaction system, despite the detection of a prominent signal from the DMPO hydroxyl radical adduct (DMPO/[formula](the formation of which was due solely to the nucleophilic addition of water to DMPO). The failure to generate an oxidant in this reaction mixture was attributed to the failure of hydrogen peroxide to reduce[formula]as hydroxyl radical formation did occur when[formula]was added directly to H2O2. However, in order to account for the high concentration of α-hydroxyethanol radi cals detected when ethanol was included in the[formula]reaction, the possibility that an oxidant in addition to[formula](e.g., CuO+) is generated is discussed.