<abstract xml:lang="en" xmlns="http://www.w3.org/1999/xhtml"><p>A simple and sensitive method was developed to separate the carcinogenic polycyclic aromatic hydrocarbon (PAH), benzo[<italic>a</italic>]pyrene (BaP), and six of its oxidation metabolites generated by rat hepatic microsomes enriched with cytochrome P450 (CYP) 1A1, by high pressure liquid chromatography (HPLC). The HPLC method, using an acetonitrile/water gradient as mobile phase and UV detection, provided appropriate separation and detection of both mono- and di-hydroxylated metabolites of BaP as well as BaP diones formed by rat hepatic microsomes and the parental BaP. In this enzymatic system, 3-hydroxy BaP, 9-hydroxy BaP, BaP-4,5-dihydrodiol, BaP-7,8-dihydrodiol, BaP-9,10-dihydrodiol and BaP-dione were generated. Among them the mono-hydroxylated BaP metabolite, 3-hydroxy BaP followed by di-hydroxylated BaP products, BaP-7,8-dihydrodiol and BaP-9,10-dihydrodiol, predominated, while BaP-dione was a minor metabolite. This HPLC method will be useful for further defining the roles of the CYP1A1 enzyme with both <italic>in vitro</italic> and <italic>in vivo</italic> models in understanding its real role in activation and detoxification of BaP.</p></abstract>

A simple and sensitive method was developed to separate the carcinogenic polycyclic aromatic hydrocarbon (PAH), benzo[a]pyrene (BaP), and six of its oxidation metabolites generated by rat hepatic microsomes enriched with cytochrome P450 (CYP) 1A1, by high pressure liquid chromatography (HPLC). The HPLC method, using an acetonitrile/water gradient as mobile phase and UV detection, provided appropriate separation and detection of both mono- and di-hydroxylated metabolites of BaP as well as BaP diones formed by rat hepatic microsomes and the parental BaP. In this enzymatic system, 3-hydroxy BaP, 9-hydroxy BaP, BaP-4,5-dihydrodiol, BaP-7,8-dihydrodiol, BaP-9,10-dihydrodiol and BaP-dione were generated. Among them the mono-hydroxylated BaP metabolite, 3-hydroxy BaP followed by di-hydroxylated BaP products, BaP-7,8-dihydrodiol and BaP-9,10-dihydrodiol, predominated, while BaP-dione was a minor metabolite. This HPLC method will be useful for further defining the roles of the CYP1A1 enzyme with both in vitro and in vivo models in understanding its real role in activation and detoxification of BaP.

Analysis of benzo[<italic>a</italic>]pyrene metabolites formed by rat hepatic microsomes using high pressure liquid chromatography: optimization of the method

Department of Biochemistry, Faculty of Science, Charles University, Albertov 2030, 128 40 Prague 2, Czech Republic

Department of Molecular Toxicology, German Cancer Research Center, 69 120 Heidelberg, Germany

Interdisciplinary Toxicology

Analysis of benzo[]pyrene metabolites formed by rat hepatic microsomes using high pressure liquid chromatography: optimization of the method

A simple and sensitive method was developed to separate the carcinogenic polycyclic aromatic hydrocarbon (PAH), benzo[a]pyrene (BaP), and six of its oxidation...

Analysis of benzo[a]pyrene metabolites formed by rat hepatic microsomes using high pressure liquid chromatography: optimization of the method

Published Online: Jan 06, 2010

Page range: 239 - 244

DOI: https://doi.org/10.2478/v10102-009-0024-0

Keywords
benzo[]pyrene, metabolism, HPLC

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