Simultaneous determination of urinary hydroxylated metabolites of naphthalene, fluorene, phenanthrene, fluoranthene and pyrene as multiple biomarkers of exposure to polycyclic aromatic hydrocarbons

Abstract

A method is presented for determining monohydroxy polycyclic aromatic hydrocarbons (OHPAHs) having 2-, 3- and 4-rings in human urine by using high-performance liquid chromatography with fluorescence detection. A urine sample containing conjugates of OHPAHs was hydrolysed in the presence of beta-glucuronidase/aryl sulfatase and the solution was cleaned up with a solid-phase extraction (C(18) and silica). Eight OHPAHs, namely 1- and 2-hydroxynaphthalenes, 2-hydroxyfluorene, 2-, 3- and 4-hydroxyphenanthrenes, 3-hydroxyfluoranthene and 1-hydroxypyrene, were separated and 1- and 9-hydroxyphenanthrenes co-eluted on an alkylamide-type reversed-phase column with fluorimetric detection. The urinary concentrations of OHPAHs were quantified by using deuterated 1-hydoxypyrene as an internal standard. The method showed good repeatability for inter- and intra-day precisions as well as good linearity of calibration curves (r (2) ranged from 0.996 to 0.999). The limits of detection (S/N=3) were in the range from 2.3 fmol to 2.2 pmol per injection. This method was successfully applied to urine samples from non-smoking taxi drivers, traffic policemen and rural villagers of Chiang Mai, Thailand. The results showed higher urinary concentrations of OHPAHs in rural villagers, consistent with higher respiratory exposure to PAHs.

Direct measurement of the glucuronide conjugate of 1-hydroxypyrene in human urine by using liquid chromatography with tandem mass spectrometry

“…Urinary 1-OHP was determined as described previously with a slight modification [12]. A 3 mL aliquot of the sample was adjusted to pH 5.…”

“…The calibration range was based on the urinary concentrations of actual subjects [11,12,27]. The matrix effect on the mass spectrometric response was evaluated for the analytes by comparing the slope of the above calibration curve with that of the working curve obtained with the urine matrix.…”

“…1-OHP is a urinary PAH metabolite that has often been used as a biomarker for recent exposure to multiple routes of PAHs. 1-OHP in human urine has been measured with analytical equipments like HPLC with fluorescence detection (HPLC-FL), GC-MS, LC-MS/MS [11][12][13][14][15][16][17][18][19][20][21].…”

“…However, these methods need enzymatic hydrolysis of its conjugates to 1-OHP for generally 2 to 16 hours (overnight) [12,22]. The hydrolysis step is time-consuming, and its possible incomplete hydrolysis should be taken into consideration [8].…”

Quasi-targeted analysis of hydroxylation-related metabolites of polycyclic aromatic hydrocarbons in human urine by liquid chromatography–mass spectrometry

“…Some of the phase I metabolites, such as monohydroxylated PAHs (OH-PAHs) have been frequently detected in human urine and hair [28][29][30]. Urinary OH-PAHs, particularly 1-hydroxypyrene (1-OHP), have been used as biomarkers to indicate human exposure to PAHs [31][32][33][34]. Therefore, investigation of urinary OH-PAHs has attracted increasing attentions in environmental studies [35,36].…”