Analysis of r -7, t -8,9, c -10-Tetrahydroxy-7,8,9,10-tetrahydrobenzo[ a ]pyrene in Human Urine: A Biomarker for Directly Assessing Carcinogenic Polycyclic Aromatic Hydrocarbon Exposure Plus Metabolic Activation (original) (raw)

2011, Chemical Research in Toxicology

Polycyclic aromatic hydrocarbons (PAH) are believed to be causative agents for various types of cancers in humans. Benzo[a]pyrene (BaP) is a prototypic carcinogenic PAH, which requires metabolic activation to elicit its detrimental effects. The major end product of its diol epoxide metabolic activation pathway is r-7, t-8,9,c-10-tetrahydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene (trans, anti-BaPT). Individual differences in exposure to, and metabolic activation of, carcinogenic PAH may influence cancer risk. Measurement of PAH metabolites in human urine could provide a direct way to assess individual differences in susceptibility to PAH-related cancer. In this paper, we describe a sensitive and reliable method for quantitation of trans, anti-BaPT in human urine using gas chromatography-negative ion chemical ionization-tandem mass spectrometry (GC-NICI-MS/MS). [ 13 C 6 ] trans, anti-BaPT was used as the internal standard. The urine was treated with β-glucuronidase and sulfatase, and then trans, anti-BaPT was enriched by solid-phase extraction with polymeric reversed phase and phenylboronic acid cartridges. The sample was silylated and analyzed by GC-NICI-MS/MS with selected reaction monitoring (SRM) for the trimethylsilyl (TMS) derivatives of trans, anti-BaPT (m/z 446→ m/z 255) and [ 13 C 6 ]trans, anti-BaPT (m/z 452→ m/z 261). The mean assay recovery was 44%. The instrumental on-column detection limit was about 20 amol of trans, anti-BaPT (as BaPT-TMS). trans, anti-BaPT was readily detected in all urine samples analyzed including 30 smokers (0.71 ± 0.64 fmol/mg creatinine) and 30 non-smokers (0.34 ± 0.2 fmol/mg creatinine) (P = 0.0018). The results of this study demonstrate a highly sensitive and selective method for quantitation of trans, anti-BaPT in human urine. This is to our knowledge the first study to show that smokers have significantly higher levels of trans, anti-BaPT in their urine than do non-smokers. This method may be useful as a direct phenotyping approach to assess individual differences in uptake and metabolic activation of carcinogenic PAH.