Continuous spectrophotometric assays for cytosolic epoxide hydrolase

Roger N. Wixtrom, Bruce D. Hammock

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Two convenient and sensitive continuous spectrophotometric assays for cytosolic epoxide hydrolase are described. The assays are based on the differences in the ultraviolet spectra of the epoxide substrates and their diol products. The hydrolysis of 1,2-epoxy-1-(p-nitrophenyl)pentane (ENP5) is accompanied by a decrease in absorbance at 302 nm, while the hydration of 1,2-epoxy-1-(2-quinolyl)pentane (EQU5) produces an increase in absorbance at 315.5 nm. The Km, Vmax values for ENP5 and EQU5 with purified mouse liver cytosolic epoxide hydrolase were 1.7 μm, 11,700 nmol/min/mg and 25 μm, 8300 nmol/min/mg, respectively. Both substrates are hydrolyzed significantly faster than trans-stilbene oxide, which is currently the most commonly used substrate for measuring cytosolic epoxide hydrolase activity. No spontaneous hydrolysis of the substrates is detectable under normal assay conditions. The assays are applicable to whole tissue homogenates as well as purified enzyme preparations. p-Nitrostyrene oxide and p-nitrophenyl glycidyl ether were also examined and found to be very poor substrates for cytosolic epoxide hydrolase from mouse liver.

Original languageEnglish (US)
Pages (from-to)291-299
Number of pages9
JournalAnalytical Biochemistry
Volume174
Issue number1
DOIs
StatePublished - 1988

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Epoxide Hydrolases
Assays
Substrates
Hydrolysis
Liver
Epoxy Compounds
Oxides
Hydration
Tissue
Enzymes
1,2-epoxy-1-(2-quinolyl)pentane
1,2-epoxy-1-(4-nitrophenyl)pentane

Keywords

  • epoxide hydrolase
  • spectrophotometric assay

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Continuous spectrophotometric assays for cytosolic epoxide hydrolase. / Wixtrom, Roger N.; Hammock, Bruce D.

In: Analytical Biochemistry, Vol. 174, No. 1, 1988, p. 291-299.

Research output: Contribution to journalArticle

Wixtrom, Roger N. ; Hammock, Bruce D. / Continuous spectrophotometric assays for cytosolic epoxide hydrolase. In: Analytical Biochemistry. 1988 ; Vol. 174, No. 1. pp. 291-299.
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