The purpose of this study was to evaluate the in vitro inhibitory potency of various amide analogues and derivatives of valproic acid toward human microsomal epoxide hydrolase (mEH). mEH inhibition was evaluated in human liver microsomes with 25 microM (S)-(+)-styrene oxide as the substrate. Inhibitory potency expressed as the median inhibitory concentration (IC50) was calculated from the formation rate of the enzymatic product, (S)-(+)-1-phenyl-1,2-ethanediol. Inhibitory potency was directly correlated with lipophilicity and became significant for amides with a minimum of eight carbon atoms. Branched eight-carbon amides were more potent inhibitors than their straight chain isomer, octanamide. N-substituted valproylamide analogues had reduced or abolished inhibition potency with the exception of valproyl hydroxamic acid being a potent inhibitor. Inhibition potency was not stereoselective in two cases of chiral valpromide isomers. Valproyl glycinamide, a new antiepileptic drug currently undergoing phase II clinical trials and its major metabolite valproyl glycine were weak mEH inhibitors. Acid isomers of valproic acid were not potent mEH inhibitors. The structural requirements for valproylamide analogues for potent in vitro mEH inhibition are: an unsubstituted amide moiety; two saturated alkyl side chains; a minimum of eight carbons in the molecule.