Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1976 Feb;73(2):607–611. doi: 10.1073/pnas.73.2.607

Identification of mutagenic metabolites of benzo(a)pyrene in mammalian cells.

E Huberman, L Sachs, S K Yang, V Gelboin
PMCID: PMC335960  PMID: 1061161

Abstract

The mutagenicity of benzo[a]pyrene and 15 of its derivatives, which included phenols, the benzo[a]yrene-4,5-epoxide (the K-region epoxide), dihydrodiols, two isomeric 7,8-diol-9,10-epoxides, a 6-methyl derivative, and a 6-hydroxymethyl derivative, were tested with Chinese hamster V79 cells in order to identify the mutagenic metabolites of benzo[a]pyrene. Mutations were characterized by resistance to ouabain or 8-azaguanine. Since V79 cells do not metabolize polycyclic hydrocarbons, mutagenesis was tested both in the presence and absence of benzo[a]pyrene-metabolizing normal golden hamster cells. All the tested phenols, 4,5-diols, trans-9,10-diol, 6-methyl, and 6-hydroxymethyl derivatives of benzo[a]pyrene showed little or no mutagenicity for both genetic markers. The (+/-)7alpha,8beta-dihydroxy-9alpha,10alpha-epoxy-7,8;9,10-tetrahydrobenzo[a]pyrene and K-region 4,5-epoxide exhibited similar and moderate mutagenicity in the absence of benzo[a]pyrene-metabolizing cells, but the (+/-)7alpha,8beta-dihydroxy-9beta,10beta-epoxy-7,8,9,10-tetrahydrobenzo[a]-pyrene showed a 2000- and 270-fold higher mutation frequency for ouabain and 8-azaguanine resistance, respectively, than did the K-region 4,5-epoxide. The trans-7,8-diol which was not mutagenic in the absence of benzo[a]pyrene-metabolizing cells was more mutagenic than benzo[a]pyrene after metabolism and mutagenesis by trans-7,8-diol in these cells was inhibited by 7,8-benzoflavone, an inhibitor of mixed-function oxidases. Metabolically formed trans-7,8-diol was isolated and incubated with rat liver microsomes in the presence of co-factors. High-pressure liquid chromatography analysis indicated that the major metabolite of trans-7,8-diol is 7alpha,8beta-dihydroxy-9beta,10beta-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene. The results indicate that the latter compound is metabolically formed and the major mutagenic intermediate of benzo[a]yrene metabolism.

Full text

PDF
607

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ames B. N., Durston W. E., Yamasaki E., Lee F. D. Carcinogens are mutagens: a simple test system combining liver homogenates for activation and bacteria for detection. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2281–2285. doi: 10.1073/pnas.70.8.2281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Baird W. M., Harvey R. G., Brookes P. Comparison of the cellular DNA-bound products of benzo(alpha)pyrene with the products formed by the reaction of benzo(alpha)pyrene-4,5-oxide with DNA. Cancer Res. 1975 Jan;35(1):54–57. [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. CHANOCK R. M., HAYFLICK L., BARILE M. F. Growth on artificial medium of an agent associated with atypical pneumonia and its identification as a PPLO. Proc Natl Acad Sci U S A. 1962 Jan 15;48:41–49. doi: 10.1073/pnas.48.1.41. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chu E. H., Malling H. V. Mammalian cell genetics. II. Chemical induction of specific locus mutations in Chinese hamster cells in vitro. Proc Natl Acad Sci U S A. 1968 Dec;61(4):1306–1312. doi: 10.1073/pnas.61.4.1306. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Diamond L., Gelboin H. V. Alpha-naphthoflavone: an inhibitor of hydrocarbon cytotoxicity and microsomal hydroxylase. Science. 1969 Nov 21;166(3908):1023–1025. doi: 10.1126/science.166.3908.1023. [DOI] [PubMed] [Google Scholar]
  7. 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]
  8. Gelboin H. V., Huberman E., Sachs L. Enzymatic hydroxylation of benzopyrene and its relationship to cytotoxicity. Proc Natl Acad Sci U S A. 1969 Dec;64(4):1188–1194. doi: 10.1073/pnas.64.4.1188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Huberman E., Aspiras L., Heidelberger C., Grover P. L., Sims P. Mutagenicity to mammalian cells of epoxides and other derivatives of polycyclic hydrocarbons. Proc Natl Acad Sci U S A. 1971 Dec;68(12):3195–3199. doi: 10.1073/pnas.68.12.3195. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Huberman E. Mammalian cell transformation and cell-mediated mutagenesis by carcinogenic polycyclic hydrocarbons. Mutat Res. 1975 Aug;29(2):285–291. doi: 10.1016/0027-5107(75)90181-5. [DOI] [PubMed] [Google Scholar]
  11. Huberman E., Sachs L. Cell-mediated mutagenesis of mammalian cells with chemical carcinogens. Int J Cancer. 1974 Mar 15;13(3):326–333. doi: 10.1002/ijc.2910130308. [DOI] [PubMed] [Google Scholar]
  12. Huberman E., Sachs L. Mutability of different genetic loci in mammalian cells by metabolically activated carcinogenic polycyclic hydrocarbons. Proc Natl Acad Sci U S A. 1976 Jan;73(1):188–192. doi: 10.1073/pnas.73.1.188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hulbert P. B. Carbonium ion as ultimate carcinogen of polycyclic aromatic hydrocarbons. Nature. 1975 Jul 10;256(5513):146–148. doi: 10.1038/256146a0. [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. Malaveille C., Bartsch H., Grover P. L., Sims P. Mutagenicity of non-K-region diols and diol-epoxides of benz(a)anthracene and benzo(a)pyrene in S. typhimurium TA 100. Biochem Biophys Res Commun. 1975 Sep 16;66(2):693–700. doi: 10.1016/0006-291x(75)90565-3. [DOI] [PubMed] [Google Scholar]
  16. Miller J. A. Carcinogenesis by chemicals: an overview--G. H. A. Clowes memorial lecture. Cancer Res. 1970 Mar;30(3):559–576. [PubMed] [Google Scholar]
  17. Selkirk J. K., Croy R. G., Roller P. P., Gelboin H. V. High-pressure liquid chromatographic analysis of benzo(alpha)pyrene metabolism and covalent binding and the mechanism of action of 7,8-benzoflavone and 1,2-epoxy-3,3,3-trichloropropane. Cancer Res. 1974 Dec;34(12):3474–3480. [PubMed] [Google Scholar]
  18. Sims P., Grover P. L. Epoxides in polycyclic aromatic hydrocarbon metabolism and carcinogenesis. Adv Cancer Res. 1974;20:165–274. doi: 10.1016/s0065-230x(08)60111-6. [DOI] [PubMed] [Google Scholar]
  19. Sims P., Grover P. L., Swaisland A., Pal K., Hewer A. Metabolic activation of benzo(a)pyrene proceeds by a diol-epoxide. Nature. 1974 Nov 22;252(5481):326–328. doi: 10.1038/252326a0. [DOI] [PubMed] [Google Scholar]
  20. 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]
  21. Wood A. W., Goode R. L., Chang R. L., Levin W., Conney A. H., Yagi H., Dansette P. M., Jerina D. M. Mutagenic and cytotoxic activity of benzol[a]pyrene 4,5-, 7,8-, and 9,10-oxides and the six corresponding phenols. Proc Natl Acad Sci U S A. 1975 Aug;72(8):3176–3180. doi: 10.1073/pnas.72.8.3176. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Yagi H., Hernandez O., Jerina D. M. Letter: Synthesis of (+/-)-7 beta,8alpha-dihydroxy-9 beta,10beta-epoxy-7,8,-9,10-tetrahydrobenzo(a)pyrene, a potential metabolite of the carcinogen benzo(a)pyrene with stereochemistry related to the antileukemic triptolides. J Am Chem Soc. 1975 Nov 12;97(23):6881–6883. doi: 10.1021/ja00856a057. [DOI] [PubMed] [Google Scholar]
  23. Yang S. K., Selkirk J. K., Plotkin E. V., Gelboin H. V. Kinetic analysis of the metabolism of benzo(a)pyrene to phenols, dihydrodiols, and quinones by high-pressure chromatography compared to analysis by aryl hydrocarbon hydroxylase assay, and the effect of enzyme induction. Cancer Res. 1975 Dec;35(12):3642–3650. [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES