An unexpected new pathway for nitroxide radical production via more reactve nitrogen-centered amidyl radical intermediate during detoxification of the carcinogenic halogenated quinones by N-alkyl hydroxamic acids

Abstract

We found previously that nitroxide radical of desferrioxamine (DFO•) could be produced from the interaction between the classic iron chelating agent desferrioxamine (DFO, an N-alkyl trihydroxamic acid) and tetrachlorohydroquinone (TCHQ), one of the carconogenic quinoind metabolites of the widely used wood preservative pentachlorophenol. However, the underlying molecular mechanism remains unclear. Here N-methylacetohydroxamic acid (N-MeAHA) was synthesized and used as a simple model compound of DFO for further mechanistic study. As expected, direct ESR studies showed that nitroxide radical of N-MeAHA (Ac-(CH3)NO•) can be produced from N-MeAHA/TCHQ. Interestingly and unexpectedly, when TCHQ was substituted by its oxidation product tetrachloro-1,4-benzoquinone (TCBQ), although Ac-(CH3)NO• could also be produced, no concurrent formation of tetrachlorosemiquinone radical (TCSQ•) and TCHQ was detected, suggesting that Ac-(CH3)NO• did not result from direct oxidation of N-MeAHA by TCSQ• or TCBQ as proposed previously. To our surprise, a new nitrogen-centered amidyl radical was found to be generated from N-MeAHA/TCBQ, which was observed by ESR with the spin-trapping agents and further unequivacally identified as Ac-(CH3)N• by HPLC-MS. The final product of amidyl radical was isolated and identified as its corresponding amine. Analogous radical homolysis mechanism was observed with other halogenated quinoid compounds and N-alkyl hydroxamic acids including DFO. Interestingly, amidyl radicals were found to induce both DNA strand breaks and DNA adduct formation, suggesting that N-alkyl hydroxamic acids may exert their potential side-toxic effects via forming the reactive amidyl radical species. This study represents the first report of an unexpected new pathway for nitroxide radical production via hydrogen abstration reaction of a more reactive amidyl radical intermediate during the detoxification of the carcinogenic polyhalogenated quinones by N-alkyl hydroxamic acids, which provides more direct experimental evidence to better explain not only our previous finding that excess DFO can provide effective but only partial protection against TCHQ (or TCBQ)-induced biological damage, and also the potential side-toxic effects induced by DFO and other N-alkyl hydroxamic acid drugs.