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PubMed · 12479830

Method for determining monohydroxybenzo[a]pyrene isomers using column-switching high-performance liquid chromatography.

Akira Toriba·Hiroaki Nakamura·Thaneeya Chetiyanukornkul·Ryoichi Kizu·Tsunehisa Makino·Hiroyuki Nakazawa·Tsuyoshi Yokoi·Kazuichi Hayakawa
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Abstract

A method for determining monohydroxybenzo[a]pyrene (OHBaP) isomers using column-switching high-performance liquid chromatography with fluorescence detection was developed. Eleven of 12 isomers of OHBaP (all except 6-OHBaP) were separated on an alkylamide-type reversed-phase column and, via column-switching, on a beta-cyclodextrin-bonded silica gel column. The detection limits for the OHBaPs were in the range 0.3-8 pg/injection (S/N=3). By using this method, 1-, 3-, and 9-OHBaPs were identified as major metabolites of benzo[a]pyrene in vitro by human recombinant p450 1A1. The method was used to determine OHBaPs in the urine of a nonsmoker subject. After enzymatic hydrolysis of the conjugated metabolites by beta-glucuronidase/aryl sulfatase, the analytes were selectively adsorbed on blue rayon (a cellulose-supported copper phthalocyanine) from the urine matrix. Methanol as the eluting solvent from the rayon gave the best recoveries of OHBaPs and 1-hydroxypyrene (1-OHP) in the range of 91-103%, which was superior to that of the solid-phase extraction method. 1-OHP, a well-known biomarker of the exposure to polycyclic aromatic hydrocarbons, was simultaneously analyzed. Intra- and interday accuracy values for the determination of 3-OHBaP in 200 ml of urine were 95.5 and 100.9%, and those for 1-OHP were 96.4 and 103.6%, respectively. The intra- and interday precision values were 3.9 and 2.4% for 3-OHBaP and 2.4 and 3.2% for 1-OHP, respectively. In 11 kinds of isomers, only 3-OHBaP was detected in the human urine. Urinary concentration of 3-OHBaP was quantified at 0.5 ng/g creatinine concentration and the 3-OHBaP/1-OHP ratio was approximately 1/130.

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Benzo(a)pyreneChromatography, High Pressure LiquidCytochrome P-450 CYP1A1IsomerismRecombinant ProteinsReproducibility of ResultsJournal ArticleResearch Support, Non-U.S. Gov't

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Akira Toriba, Hiroaki Nakamura, Thaneeya Chetiyanukornkul, Ryoichi Kizu, Tsunehisa Makino, Hiroyuki Nakazawa, Tsuyoshi Yokoi, Kazuichi Hayakawa. 2003-01-01. Method for determining monohydroxybenzo[a]pyrene isomers using column-switching high-performance liquid chromatography.. https://doi.org/10.1016/s0003-2697(02)00429-3

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