The utility of the microsomal 4-chlorobiphenyl hydroxylase enzyme assay in distinguishing between phenobarbitone- and 3-methylcholanthrene-induced microsomal monooxygenases

Andrew Parkinson, Leslie Copp, Stephen Safe

Research output: Contribution to journalArticle

14 Scopus citations

Abstract

4-Chlorobiphenyl was used as a substrate for the in vitro determination of rat hepatic microsomal, cytochrome P-450-dependent monooxygenase activity. The 4-chlorobiphenyl hydroxylase assay was tested for its ability to distinguish between a variety of phenobarbitone- and 3-methylcholanthrene-type inducers. Two radiometric procedures were employed to investigate the metabolism of 4-chlorobiphenyl. First, the metabolite profile of 4-chlorobiphenyl was analyzed by radio-thin-layer chromatography. This procedure permitted an assessment of the effects of microsomal enzyme inducers on both the qualitative and quantitative aspects of 4-chlorobiphenyl metabolism. Second, the rate of 4-chlorobiphenyl metabolism was determined by a differential extraction procedure which separated unreacted starting material (hexane phase) from metabolites (base phase). This procedure provided a rapid measurement of the overall activity of 4-chlorobiphenyl hydroxylase. Irrespective of the animal pretreatment, the metabolite profile of 4-chlorobiphenyl was dominated by 4′-chloro-4-biphenyl. Unlike the qualitative aspects, the quantitative aspects of 4-chlorobiphenyl metabolism were markedly influenced by animal pretreatment. Specifically, 3-methylcholanthrene-type inducers (3-methylcholanthrene and 3,3′,4,4′-tetrachlorobiphenyl) enhanced the activity of 4-chlorobiphenyl hydroxylase at least 10 times more than phenobarbitone-type inducers (phenobarbitone, 2,2′,4,4′-tetrachlorobiphenyl, and 2,2′,4,4′,5,5′-hexachlorobiphenyl) enabling these two classes of inducers to be clearly distinguished. It is concluded that 4-chlorobiphenyl is preferentially metabolized by the 3-methylcholanthrene-inducible form(s) of cytochrome P-450 and that this class of microsomal enzyme inducers can be readily distinguished from phenobarbitone-type inducers by means of the 4-chlorobiphenyl hydroxylase assay.

Original languageEnglish (US)
Pages (from-to)65-73
Number of pages9
JournalAnalytical Biochemistry
Volume105
Issue number1
DOIs
StatePublished - Jun 1980

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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