EPOXIDE HYDROLASE, ITS FUNCTION AND DETERMINATION OF ITS ACTIVITY IN RAT-LIVER

Epoxides are a group of reactive intermediates formed by the cytochrome P-450-mediated monooxygenation of unsaturated xenobiotics. Epoxide hydrolase inactivates these epoxides by addition of water to form diols. Commonly the function of epoxide hydrolase is finally followed by excretion of the diols. However, reactivation of certain diols by a second epoxidation may happen. Epoxide hydrolase inactivates also the epoxides existing in the metabolism of endogenous compounds. The determination of the activity of epoxide hydrolase by the addition of water to styrene oxide (1,2-epoxyethylbenzene) and measurement of the concentration of the produced phenylglycol (1-phenyl-1,2-ethandiol) with subsequent separation of the 2 substances by HPLC is described. Lipophilic xenobiotics tend to accumulate into tissues, and they must be transformed to water soluble compounds to enable the excretion. In this transformation process reactive intermediates are produced. If biotransformation fails to detoxify these reactive intermediates, they may react covalently with critical targets like the genetic material, or start harmful reaction chains like lipid peroxidation. As a result of this carcinogenicity, mutagenicity etc. may ensue Miller and Miller (1976). Depending on the chemical structure of the molecule, different kinds of reactive substances are generated. Epoxides originate from oxidation of an aliphatic or aromatic double bond by the action of cytochrome P-450-mediated monooxygenases (Leibmann et al. 1979). One detoxifying pathway is the addition of water to form diols, which are of low reactivity; this reaction is catalized by epoxide hydrolase. Other possible pathways are the formation of glutathion conjugates or the rearrangement to aldehydes or ketones (Habig et al. 1974; Oesch 1979). Epoxide hydrating enzymes have been detected in many species (Walker et al. 1978). In rat liver it becomes detectable on the 15 th day of gestation and increases slowly to the day of delivery. After birth there is a further growth of activity, and adult levels are reached in the second month of life (James et al. 1977). The activity of epoxide hydrolase increases in response to exposure of many foreign compounds. Phenobarbital was shown as a typical inducer of epoxide hydrolase in rat liver, also a large number of chlorinated hydrocarbons elevates the activity of the enzyme. An inhibition is reached by many epoxides, alkylating chemicals, heavy metals and organophosphates (Oesch et al. 1973; Parkki and Aitio 1978; Parkki 1980). The substrate specificity of epoxide hydrolase is very low, many substrates have been investigated. Numerous aliphatic, alicyclic and aromatic epoxides are biotransformed by this enzyme. A very often used substrate is styrene oxide which is also used in our institute for the determination of enzyme activity in rat liver. The method was published by Schoket and Vincze (1985) and modified for our conditions and purposes. © 1990, VEB Gustav Fischer Verlag Jena. All rights reserved.