ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS Vol. 269, No. 1, February 15, pp. 30%312,1989 Metabolism of Retinol and Retinoic Acid by Human Liver Cytochrome P45OllC8’ MARIA ANNA LEO, JEROME M. LASKER, JUDY L. RAUCY,* CHO-IL KIM, MARTIN BLACK,* AND CHARLES S. LTEBER2 Section of Liver Disease and Nutrition, Alcohol Research and Treatment Center, Brow Veterans Medical Center, Mt. Sinai School of Medicine, New York, New York 10468, and *Department of Pharmacology, Temple University School of Medicine, Ph,iladelphia. Pennsylvania 19140 Received August 4,1988, and in revised form October 24,1988 Liver microsomes obtained from nine subjects were found to metabolize retinol to polar metabolites, including 4-hydroxyretinol. In a reconstituted monooxygenase system containing human liver P45OIIC8, retinol was converted to 4-hydroxyretinol and other polar metabolites, with a & of 0.071 MM and a V,,, of 1.73 nmol/min/nmol P450. Nei- ther P45OIIC9 nor P450IIE1, two other purified human P45Os, displayed significant reti- no1 hydroxylase activity. Immunoblots performed with a monospecific antibody directed against human P45OIIC8 revealed that appreciable amounts of this enzyme were present in human liver microsomes. The same antibody significantly inhibited retinol metabo- lism in liver microsomes and in the system reconstituted with P45OIIC8. The system reconstituted with P45OIIC8 also converted retinoic acid to polar metabolites. Thus, this study shows, for the first time, metabolism of two physiologic substrates by a human liver cytochrome P450 related to a group of “constitutive” rodent P45Os believed to par- ticipate in the metabolism of endogenous compounds. Through its involvement in vita- min A metabolism, P45OIIC8 may participate in maintaining the balance between those vitamin A concentrations that promote cellular integrity (and oppose the development of cancer) and those concentrations that cause cellular toxicity. 0 1989 Academic Press, Inc. It is generally accepted that the first step in the hepatic metabolism of vitamin A is the oxidation of retinol to retinal by the cytosolic enzyme retinol dehydroge- nase (1). Recently, however, a new path- way of retinol metabolism was described: rat liver microsomes, when fortified with NADPH, converted retinol to polar metab- olites, including 4-hydroxyretinol (2). This activity was also demonstrated in a recon- stituted monooxygenase system contain- ing purified forms of rat cytochromes P450”, including P4501IBl (a phenobarbi- ’ This study was supported by DHSS Grants DK32810, AA03508, and AA05934 and the Veterans Administration. a To whom correspondence should be addressed. ’ Abbreviations used: P450, liver microsomal cyto- chrome P450; Fp, liver microsomal NADPH-cyto- chrome P450 reductase; IgG, immunoglobulin G; tal-inducible isozyme) and P4501IC7 (a “constitutive” isozyme, also called P450f) (2). The present study was undertaken to determine whether human liver micro- somes can metabolize retinol. Having veri- fied this fact, we then attempted to estab- lish the possible forms of cytochrome P450 involved. We focused our investigation on P45OIIC8 because recent studies have shown that this human liver cytochrome is immunochemically related to a family of constitutive rat P45Os, including P450f (3). Furthermore, we had previously observed that benzphetamine competitively inhibits the metabolism of retinol in rat liver mi- crosomes (4). Of the various substrates ex- amined, only benzphetamine was reported PAGE, polyacrylamide gel electrophoresis; SDS, so- dium dodecyl sulfate. 305 0003-9861189 $3.00 Copyright 0 1989 hy Academic Press. Inc All rights of reproduction in any form reserved.