EROD and MROD as Markers of Cytochrome P450 1A Activities in Hepatic Microsomes from Entire and Castrated Male Pigs.docVIP

Sensors 2009, 9, 2134-2147; doi:10.3390/OPEN ACCESS sensors ISSN 1424-8220 /journal/sensors Article EROD and MROD as Markers of Cytochrome P450 1A Activities in Hepatic Microsomes from Entire and Castrated Male Pigs Galia Zamaratskaia 1,* and Vladimir Zlabek 1,2 1 Department of Food Science, Swedish University of Agricultural Sciences, SE-750 07, Uppsala, Sweden; E-mail: Vladimir.Zlabek@lmv.slu.se (V.Z.) 2 University of South Bohemia, Ceske Budejovice, Research Institute of Fish Culture and Hydrobiology, Zatisi 728, 389 25 Vodňany, Czech Republic * Author to whom correspondence should be addressed; E-Mails: Galia.Zamaratskaia@lmv.slu.se; Tel.: +46-18-672003; Fax: +46-18-672995 Received: 21 February 2009; in revised form: 18 March 2009 / Accepted:19 March 2009 / Published: 23 March 2009 Abstract: In the present study, we characterized the kinetic parameters of 7-ethoxy- resorufin O-deethylation (EROD) and 7-methoxyresorufin O-demethylation (MROD) in hepatic microsomes from entire and castrated male pigs. Validation parameters of an HPLC-based method to analyse EROD and MROD activities are also described. Eadie- Hofstee plot analysis demonstrated a biphasic kinetic of EROD, indicating that at least two forms of cytochrome P450 are involved in this reaction. MROD followed monophasic kinetic, suggesting that a single enzyme, or enzymes with similar affinities, is responsible for the reaction. Inhibitory effects of α-naphthoflavone (ANF), ellipticine and furafylline were studied using microsomes from entire and castrated male pigs. ANF is a known inhibitor of both cytochrome P450 1A1 and 1A2 (CYP1A1 and CYP1A2); the presence of ANF in the incubations resulted in the inhibition of both EROD and MROD activities in porcine liver microsomes. EROD activities in porcine liver microsomes were also inhibited by selective CYP1A1 inhibitor ellipticine, but not by