Pathak D N, Roy D
Department of Environmental Health Sciences, School of Public Health, University of Alabama, Birmingham 35294.
Carcinogenesis. 1992 Sep;13(9):1593-7. doi: 10.1093/carcin/13.9.1593.
It has been previously reported that the reactive metabolites phenylsemiquinone and phenylbenzoquinone are generated during microsomal cytochrome P450-catalyzed redox cycling of o-phenylphenol (OPP). However, covalent modification of DNA by OPP-reactive metabolites has yet not been demonstrated. In the present study we have investigated the covalent binding in DNA by OPP-reactive metabolites using 32P-postlabeling. Analysis of adducts by 32P-postlabeling in products of chemical reaction of DNA with phenylbenzoquinone revealed four major and several minor adducts. The chemical reaction of deoxyguanosine 3'-phosphate with phenylbenzoquinone also showed four major adducts. The chromatographic mobility of major adducts of deoxyguanosine 3'-phosphate-phenylbenzoquinone was identical to that of major adducts of DNA-phenylbenzoquinone. The major adducts are demonstrated to be stable. The total covalent binding in deoxyguanosine 3'-phosphate by phenylbenzoquinone (686,000-687,000 amol/nmol nucleotide) was higher than that observed in DNA (26,500-28,000 amol/nmol nucleotides). Reaction of DNA with OPP or a hydroxylated metabolite of OPP, phenylhydroquinone, in the presence of microsomes and NADPH or cumene hydroperoxide showed four major adducts. Adduct formation in DNA by OPP or phenylhydroquinone in the presence of the microsomal activation system was drastically decreased by known inhibitors of cytochrome P450. The chromatographic mobility of major adducts in DNA by OPP or phenylhydroquinone in the presence of microsomal activation system matched with those major adducts observed in deoxyguanosine 3'-phosphate or DNA reacted with pure phenylbenzoquinone. These data demonstrate that OPP or phenylhydroquinone, a hydroxylated metabolite of OPP, is able to bind covalently to DNA in the presence of a microsomal cytochrome P450 activation system. Phenylbenzoquinone is one of the DNA-binding metabolite(s) of OPP. It is concluded that OPP is genotoxic in an in vitro system and genotoxicity produced by OPP-reactive metabolites may play a role in OPP-induced cellular toxicity or cancer.
此前已有报道称，在微粒体细胞色素P450催化的邻苯基苯酚（OPP）氧化还原循环过程中会生成反应性代谢产物苯半醌和苯醌。然而，尚未证实OPP反应性代谢产物对DNA的共价修饰。在本研究中，我们使用32P后标记法研究了OPP反应性代谢产物与DNA的共价结合。通过32P后标记法分析DNA与苯醌化学反应产物中的加合物，发现了四种主要加合物和几种次要加合物。脱氧鸟苷3'-磷酸与苯醌的化学反应也显示出四种主要加合物。脱氧鸟苷3'-磷酸 - 苯醌主要加合物的色谱迁移率与DNA - 苯醌主要加合物的色谱迁移率相同。已证明主要加合物是稳定的。苯醌与脱氧鸟苷3'-磷酸的总共价结合（686,000 - 687,000 amol/ nmol核苷酸）高于在DNA中观察到的结合（26,500 - 28,000 amol/ nmol核苷酸）。在微粒体和NADPH或异丙苯过氧化氢存在下，DNA与OPP或OPP的羟基化代谢产物苯氢醌反应显示出四种主要加合物。已知的细胞色素P450抑制剂可显著降低在微粒体活化系统存在下OPP或苯氢醌在DNA中形成的加合物。在微粒体活化系统存在下，OPP或苯氢醌在DNA中形成的主要加合物的色谱迁移率与在脱氧鸟苷3'-磷酸或与纯苯醌反应的DNA中观察到的主要加合物相匹配。这些数据表明，OPP或OPP的羟基化代谢产物苯氢醌在微粒体细胞色素P450活化系统存在下能够与DNA共价结合。苯醌是OPP的DNA结合代谢产物之一。得出的结论是，OPP在体外系统中具有遗传毒性，并且由OPP反应性代谢产物产生的遗传毒性可能在OPP诱导的细胞毒性或癌症中起作用。
