A rapid radiometric assay for mammalian cytosolic epoxide hydrolase

C. A. Mullin, B. D. Hammock

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

A rapid and sensitive radiometric assay for cytosolic epoxide hydrolase is described. The assay is based on the highly efficient partitioning of unreacted 1-phenyl-1,2-epoxybutane (β-ethylstyrene oxide) from an aqueous mix into isooctane, with the product diol being retained in the aqueous phase. The trans-epoxide is an excellent substrate for both the glutathione transferase and epoxide hydrolase present in mouse liver cytosol, and both enzymes may be monitored simultaneously after appropriate modifications of the assay are made. Also, trans-β-ethylstyrene oxide is hydrated much faster than the cis-isomer in the hepatic cytosol of three different mammalian species, and both isomers are poor substrates for the microsomal epoxide hydrolase. Mouse liver hydrates the trans-epoxide at 69 nmol/min-mg protein and a K(m) of 7.2x10-5 M is observed. Reduction of 1-phenyl-1-bromo-2-butanone with sodium borotritide and subsequent ring closure of the bromohydrin mix with base was a highly efficient route for radiolabeling β-ethylstyrene oxide. The resulting cis- and trans-epoxides were separable by semipreparative high-performance liquid chromatography.

Original languageEnglish (US)
Pages (from-to)476-485
Number of pages10
JournalAnalytical Biochemistry
Volume106
Issue number2
StatePublished - 1980

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Epoxide Hydrolases
Epoxy Compounds
Oxides
Assays
Isomers
Liver
Cytosol
High performance liquid chromatography
Substrates
Glutathione Transferase
Hydrates
High Pressure Liquid Chromatography
Enzymes
Proteins

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Mullin, C. A., & Hammock, B. D. (1980). A rapid radiometric assay for mammalian cytosolic epoxide hydrolase. Analytical Biochemistry, 106(2), 476-485.

A rapid radiometric assay for mammalian cytosolic epoxide hydrolase. / Mullin, C. A.; Hammock, B. D.

In: Analytical Biochemistry, Vol. 106, No. 2, 1980, p. 476-485.

Research output: Contribution to journalArticle

Mullin, CA & Hammock, BD 1980, 'A rapid radiometric assay for mammalian cytosolic epoxide hydrolase', Analytical Biochemistry, vol. 106, no. 2, pp. 476-485.
Mullin, C. A. ; Hammock, B. D. / A rapid radiometric assay for mammalian cytosolic epoxide hydrolase. In: Analytical Biochemistry. 1980 ; Vol. 106, No. 2. pp. 476-485.
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