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. 1974 Nov;71(11):4356–4360. doi: 10.1073/pnas.71.11.4356

Effects of Inducers and Epoxide Hydrase on the Metabolism of Benzo[a]pyrene by Liver Microsomes and a Reconstituted System: Analysis by High Pressure Liquid Chromatography

Gerald Holder *, Haruhiko Yagi *, Patrick Dansette *, Donald M Jerina *, W Levin †, Anthony Y H Lu †, A H Conney †
PMCID: PMC433882  PMID: 4530987

Abstract

The mobilities of 24 potential metabolites of benzo[a]pyrene were examined with high pressure liquid chromatography. Twelve phenols, five quinones, four dihydrodiols, and three oxides were studied. The chromatographic procedure employed allowed the separation and quantitation of benzopyrene metabolites into three major groups consisting of phenols, quinones, and dihydrodiols. Two of the benzopyrene oxides were unstable during chromatography, whereas the third oxide was more stable and chromatographed in the quinone fraction.

Treatment of rats with phenobarbital or 3-methylcholanthrene enhanced the metabolism of benzopyrene by liver microsomes and altered the relative amounts of the various metabolites formed. In the absence of epoxide hydrase (EC 4.2.1.63), benzopyrene was metabolized primarily to phenols and quinones but was not appreciably metabolized to dihydrodiols by a solubilized, reconstituted cytochrome P-448 monooxygenase system. Addition of partially purified epoxide hydrase resulted in the formation of benzopyrene dihydrodiols with a concomitant decrease in the formation of phenolic metabolites, indicating that benzopyrene undergoes metabolism via arene oxides that are precursors for dihydrodiols and phenols.

Keywords: carcinogen metabolism, microsomal cytochrome P-450

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Selected References

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  1. Alvares A. P., Schilling G., Levin W., Kuntzman R. Alteration of the microsomal hemoprotein by 3-methylcholanthrene: effects of ethionine and actinomycin D. J Pharmacol Exp Ther. 1968 Oct;163(2):417–424. [PubMed] [Google Scholar]
  2. BERENBLUM I., SCHOENTAL R. Metabolism of 3,4-benzopyrene. Science. 1955 Sep 9;122(3167):470–470. doi: 10.1126/science.122.3167.470. [DOI] [PubMed] [Google Scholar]
  3. Borgen A., Darvey H., Castagnoli N., Crocker T. T., Rasmussen R. E., Wang I. Y. Metabolic conversion of benzo(a)pyrene by Syrian hamster liver microsomes and binding of metabolites to deoxyribonucleic acid. J Med Chem. 1973 May;16(5):502–506. doi: 10.1021/jm00263a020. [DOI] [PubMed] [Google Scholar]
  4. CONNEY A. H., MILLER E. C., MILLER J. A. Substrate-induced synthesis and other properties of benzpyrene hydroxylase in rat liver. J Biol Chem. 1957 Oct;228(2):753–766. [PubMed] [Google Scholar]
  5. Flesher J. W., Sydnor K. L. Possible role of 6-hydroxymethylbenzo(a)pyrene as a proximate carcinogen of benzo(a(pyrene and 6-methylbenzo(a)pyrene. Int J Cancer. 1973 Mar 15;11(2):433–437. doi: 10.1002/ijc.2910110221. [DOI] [PubMed] [Google Scholar]
  6. Gelboin H. V. A microsome-dependent binding of benzo[a]pyrene to DNA. Cancer Res. 1969 Jun;29(6):1272–1276. [PubMed] [Google Scholar]
  7. Grover P. L., Hewer A., Sims P. Epoxides as microsomal metabolites of polycyclic hydrocarbons. FEBS Lett. 1971 Oct 15;18(1):76–80. doi: 10.1016/0014-5793(71)80411-8. [DOI] [PubMed] [Google Scholar]
  8. Grover P. L., Hewer A., Sims P. Formation of K-region epoxides as microsomal metabolites of pyrene and benzo(a)pyrene. Biochem Pharmacol. 1972 Oct 15;21(20):2713–2726. doi: 10.1016/0006-2952(72)90020-2. [DOI] [PubMed] [Google Scholar]
  9. Grover P. L., Sims P., Huberman E., Marquardt H., Kuroki T., Heidelberger C. In vitro transformation of rodent cells by K-region derivatives of polycyclic hydrocarbons. Proc Natl Acad Sci U S A. 1971 Jun;68(6):1098–1101. doi: 10.1073/pnas.68.6.1098. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Jerina D. M., Daly J. W. Arene oxides: a new aspect of drug metabolism. Science. 1974 Aug 16;185(4151):573–582. doi: 10.1126/science.185.4151.573. [DOI] [PubMed] [Google Scholar]
  11. Jerina D. M., Daly J. W., Witkop B., Zaltzman-Nirenberg P., Udenfriend S. 1,2-naphthalene oxide as an intermediate in the microsomal hydroxylation of naphthalene. Biochemistry. 1970 Jan 6;9(1):147–156. doi: 10.1021/bi00803a019. [DOI] [PubMed] [Google Scholar]
  12. Jerina D. M., Kaubisch N., Daly J. W. Arene oxides as intermediates in the metabolism of aromatic substrates: alkyl and oxygen migrations during isomerization of alkylated arene oxides. Proc Natl Acad Sci U S A. 1971 Oct;68(10):2545–2548. doi: 10.1073/pnas.68.10.2545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kaubisch N., Daly J. W., Jerina D. M. Arene oxides as intermediates in the oxidative metabolism of aromatic compounds. Isomerization of methyl-substituted arene oxides. Biochemistry. 1972 Aug 1;11(16):3080–3088. doi: 10.1021/bi00766a022. [DOI] [PubMed] [Google Scholar]
  14. Kinoshita N., Shears B., Gelboin H. V. K-region and non-K-region metabolism of benzo(a)pyrene by rat liver microsomes. Cancer Res. 1973 Aug;33(8):1937–1944. [PubMed] [Google Scholar]
  15. Lesko S. A., Jr, Ts'o P. O., Umans R. S. Interaction of nucleic acids. V. Chemical linkage of 3,4-benzpyrene to deoxyribonucleic acid in aqueous solution. Biochemistry. 1969 Jun;8(6):2291–2298. doi: 10.1021/bi00834a009. [DOI] [PubMed] [Google Scholar]
  16. Levin W., Ryan D., West S., Lu A. Y. Preparation of partially purified, lipid-depleted cytochrome P-450 and reduced nicotinamide adenine dinucleotide phosphate-cytochrome c reductase from rat liver microsomes. J Biol Chem. 1974 Mar 25;249(6):1747–1754. [PubMed] [Google Scholar]
  17. Lubet R. A., Brown D. Q., Kouri R. E. The role of 3-OH benzo(a)pyrene in mediating benzo(a)pyrene induced toxicity and transformation in cell culture. Res Commun Chem Pathol Pharmacol. 1973 Nov;6(3):929–942. [PubMed] [Google Scholar]
  18. Marquardt H., Kuroki T., Huberman E., Selkirk J. K., Heidelberger C., Grover P. L., Sims P. Malignant transformation of cells derived from mouse prostate by epoxides and other derivatives of polycyclic hydrocarbons. Cancer Res. 1972 Apr;32(4):716–720. [PubMed] [Google Scholar]
  19. Meunier M., Chauveau J. Binding of benzo(a)pyrene metabolite(s) to main and satellite calf thymus DNA's in vitro. FEBS Lett. 1973 May 1;31(3):327–331. doi: 10.1016/0014-5793(73)80133-4. [DOI] [PubMed] [Google Scholar]
  20. Nagata C., Tagashira Y., Kodama M., Ioki Y., Oboshi S. Effect of hydrogen peroxide, Fenton's reagent, and iron ions on the carcinogenicity of 3, 4-benzopyrene. Gan. 1973 Jun;64(3):277–285. [PubMed] [Google Scholar]
  21. Oesch E., Jerina D. M., Daly J. A radiometric assay for hepatic epoxide hydrase activity with [7-3H] styrene oxide. Biochim Biophys Acta. 1971 Mar 10;227(3):685–691. doi: 10.1016/0005-2744(71)90017-9. [DOI] [PubMed] [Google Scholar]
  22. Raha C. R. Metabolism of benzo(a)pyrene at the 4,5-position. Indian J Biochem Biophys. 1972 Mar;9(1):105–110. [PubMed] [Google Scholar]
  23. Selkirk J. K., Croy R. G., Gelboin H. V. Benzo(a)pyrene metabolites: efficient and rapid separation by high-pressure liquid chromatography. Science. 1974 Apr 12;184(4133):169–171. doi: 10.1126/science.184.4133.169. [DOI] [PubMed] [Google Scholar]
  24. Selkirk J. K., Huberman E., Heidelberger C. An epoxide is an intermediate in the microsomal metabolism of the chemical carcinogen, dibenz(a,h)anthracene. Biochem Biophys Res Commun. 1971 Jun 4;43(5):1010–1016. doi: 10.1016/0006-291x(71)90562-6. [DOI] [PubMed] [Google Scholar]
  25. Sims P. The metabolism of benzo[a]pyrene by rat-liver homogenates. Biochem Pharmacol. 1967 Apr;16(4):613–618. doi: 10.1016/0006-2952(67)90071-8. [DOI] [PubMed] [Google Scholar]
  26. Sims P. The synthesis of 8- and 9-hydroxybenzo[a]pyrene and the role of the products in benzo[a]pyrene metabolism. J Chem Soc Perkin 1. 1968;1:32–34. doi: 10.1039/j39680000032. [DOI] [PubMed] [Google Scholar]
  27. TARBELL D. S., BROOKER E. G., SEIFERT P., VANTERPOOL A., CLAUS C. J., CONWAY W. Studies on the metabolic products obtained from mouse skin after painting with 3, 4-benzpyrene. Cancer Res. 1956 Jan;16(1):37–47. [PubMed] [Google Scholar]
  28. Wang I. Y., Rasmussen R. E., Crocker T. T. Isolation and characterization of an active DNA-binding metabolite of benzo(a)pyrene from hamster liver microsomal incubation systems. Biochem Biophys Res Commun. 1972 Nov 15;49(4):1142–1149. doi: 10.1016/0006-291x(72)90332-4. [DOI] [PubMed] [Google Scholar]
  29. Waterfall J. F., Sims P. Epoxy derivatives of aromatic polycyclic hydrocarbons. The preparation and metabolism of epoxides related to benzo(a)pyrene and to 7,8- and 9,10-dihydrobenzo(a)pyrene. Biochem J. 1972 Jun;128(2):265–277. doi: 10.1042/bj1280265. [DOI] [PMC free article] [PubMed] [Google Scholar]

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