Halogenated epoxides and related compounds as inhibitors of epoxide hydrase

Abstract

A variety of chlorinated and fluorinated epoxides and related compounds were synthesized and evaluated as inhibitors of epoxide hydrase. The compounds were tested using chicken liver microsomes and a radiometric assay based on [ 3H]styrene oxide, and using partially purified chicken liver microsomal epoxide hydrase and a continuous photometric assay based on p-nitrostyrene oxide, whose hydration could be monitored at 310 nm. For the 16 compounds studied both assays gave similar patterns of inhibitory activity. As expected from the relative K m values of the two substrates, all inhibitors were considerably more active against styrene oxide ( K m =1.0 mM) than against p-nitrostyrene oxide ( K m = 4.2 μM), and styrene oxide was a weak alternate-substrate inhibitor against p-nitrostyrene oxide. 1,1,1-Trichloropropene oxide, however, was a potent alternate-substrate inhibitor against p-nitrostyrene oxide. Addition of various substituents to the α-carbon of styrene oxide generated a series of compounds whose inhibitory potency toward p-nitrostyrene oxide increased in the order H ≈ CF 3 < CH 3 < CH 2Cl < CHCl 2 < CCl 3 ≈ 1,1,1-trichloropropene oxide. In contrast, addition of a CH 3 or CCl 3 group to the β-carbon of styrene oxide resulted in only a modest increase in inhibitory potency. 2-Phenyl- and 3-phenyloxetane showed no pronounced inhibitory activity toward either styrene oxide or p-nitrostyrene oxide, but pentafluorophenyl ethylene oxide and 1,1, 1-trichlorobutane-3,4-oxide were moderately active inhibitors, although significantly less potent than 1,1,1-trichloroproene oxide. These results show that electronegativity, steric effects, and hydrophobic effects are each important in governing the interaction of epoxide hydrase substrates with the enzyme, although it is not yet possible to analyze separately the effects of each of these parameters on K m , V, and the catalytic mechanism.