Characteristics of the active oxygen in covalent binding of the pesticide methoxychlor to hepatic microsomal proteins.
This study examined the characteristics of the active oxygen species involved in generation of the reactive intermediate of methoxychlor which covalently binds to liver microsomal proteins. The possibility that the active oxygen participating in the above reaction is the superoxide anion (O2-) or a species generated from O2- was examined with the help of superoxide dismutase (SOD) and with an SOD-mimetic agent, CuDIPS [Cu2+(3,5-diisopropylsalicylic acid)2]. It was observed that, whereas CuDIPS inhibited covalent binding of methoxychlor metabolite(s), SOD did not. However, ZnDIPS [Zn2+(3,5-diisopropylsalicylic acid)2], which exhibits no SOD-mimetic activity, did not inhibit covalent binding. Furthermore, both CuDIPS and ZnDIPS had little or no effect on the formation of demethylated (polar) metabolites of methoxychlor, demonstrating that the inhibition of covalent binding by CuDIPS was not merely due to a general inhibition of the hepatic monooxygenase system. These findings suggested that O2- was involved in covalent binding, but was not accessible to SOD. Additional support for O2- involvement stems from the observation that alpha-tocopheryl acid succinate markedly inhibited covalent binding of methoxychlor. The possibility that hydrogen peroxide (H2O2) was involved in covalent binding of methoxychlor appears unlikely. Catalase had no effect on covalent binding when NADPH was the cofactor, and the use of H2O2 in place of NADPH did not yield covalent binding. Certain scavengers of hydroxyl radical (ethanol, t-butanol and benzoate) inhibited, and other known scavengers (DMSO and mannitol) did not inhibit, covalent binding. EDTA stimulated binding, desferal (desferrioxamine) exhibited no effect on binding, and diethylenetriaminepentaacetic acid (DETAPAC) inhibited binding. A possible explanation for this observation is that the Fe2+ needed for generation of X OH is much more easily obtained from Fe3+-EDTA than from Fe3+-desferal, which resists reduction. The inhibitory effect by DETAPAC may be due to chelation of another metal which is needed for the reaction. Lastly, certain scavengers of singlet oxygen inhibited covalent binding with little effect on the formation of polar metabolites of methoxychlor. In conclusion, these studies support the involvement of X OH and singlet oxygen, possibly derived from O2-, in the formation of the reactive methoxychlor intermediate.(ABSTRACT TRUNCATED AT 400 WORDS)
D001565:Benzoates / Q000494:pharmacology; D019817:Benzoic Acid; D000440:Butanols / Q000494:pharmacology; D002338:Carotenoids / Q000494:pharmacology; D002374:Catalase / Q000378:metabolism; D003676:Deferoxamine / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D000431:Ethanol / Q000494:pharmacology; D006639:Histidine / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000378:metabolism; D008353:Mannitol / Q000494:pharmacology; D008731:Methoxychlor / Q000378:metabolism*; D008862:Microsomes, Liver / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism*; D004369:Pentetic Acid / Q000494:pharmacology; D011506:Proteins / Q000378:metabolism*; D012459:Salicylates / Q000378:metabolism; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism; D014364:Tryptophan / Q000494:pharmacology; D015032:Zinc / Q000378:metabolism; D019207:beta Carotene; D020002:tert-Butyl Alcohol
D001565:Benzoates; D000440:Butanols; D011506:Proteins; D012459:Salicylates; D019207:beta Carotene; D013481:Superoxides; C030865:copper bis(3,5-diisopropylsalicylate); D002338:Carotenoids; D000431:Ethanol; D008353:Mannitol; D006639:Histidine; D009249:NADP; D004369:Pentetic Acid; D014364:Tryptophan; D019817:Benzoic Acid; D004492:Edetic Acid; D006861:Hydrogen Peroxide; D002374:Catalase; D013482:Superoxide Dismutase; D003676:Deferoxamine; D015032:Zinc; D020002:tert-Butyl Alcohol; D008731:Methoxychlor; D010100:Oxygen