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Bulletin of the Korean Chemical Society (BKCS)

ISSN 0253-2964(Print)
ISSN 1229-5949(Online)
Volume 24, Number 5
BKCSDE 24(5)
May 20, 2003 

Studies on the Analysis of Benzo(a)pyrene and Its Metabolites in Biological Samples by Using High Performance Liquid Chromatograpy/Fluorescence Detection and Gas Chromatography/Mass Spectrometry
Won Lee, Hye Seung Shin, Jee Eun Hong, Heesoo Pyo, Yunje Kim
Benzo(a)pyrene, Hydroxylated metabolites, Rat urine, Blood plasma, HPLC/FLD
An analytical method the determination of benzo(a)pyrene (BaP) and its hydroxylated metabolites, 1-hydroxybenzo(a)pyrene (1-OHBaP), 3-hydroxybenzo(a)pyrene (3-OHBaP), benzo(a)pyrene-4,5-dihydrodiol (4,5-diolBaP) and benzo(a)pyrene-7,8-dihydrodiol (7,8-diolBaP), in rat urine and plasma has been developed by HPLC/FLD and GC/MS. The derivatization with alkyl iodide was employed to improve the resolution and the detection of two mono hydroxylated metabolites, 1-OHBaP and 3-OHBaP, in LC and GC. BaP and its four metabolites in spiked urine were successfully separated by gradient elution on reverse phase ODS C18 column (4.6 mm I.D., 100 mm length, particle size 5 mm) using a binary mixture of MeOH/H2O (85/15, v/v) as mobile phase after ethylation at 90 oC for 10 min. The extraction recoveries of BaP and its metabolites in spiked samples with liquid-liquid extraction, which was better than solid phase extraction, were in the range of 90.3- 101.6% in n-hexane for urine and 95.7-106.3% in acetone for plasma, respectively. The calibration curves has shown good linearity with the correlation coefficients (R2) varying from 0.992 to 1.000 for urine and from 0.996 to 1.000 for plasma, respectively. The detection limits of all analytes were obtained in the range of 0.01- 0.1 ng/mL for urine and 0.1-0.4 ng/mL for plasma, respectively. The metabolites of BaP were excreted as mono hydroxy and dihydrodiol forms after intraperitoneal injection of 20 mg/kg of BaP to rats. The total amounts of BaP and four metabolites excreted in dosed rat urine were 3.79 ng over the 0-96 hr period from adminstration and the excretional recovery was less than 0.065% of the injection amounts of BaP. The proposed method was successfully applied to the determination of BaP and its hydroxylated metabolites in rat urine and plasma for the pharmacokinetic studies.
559 - 565
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