A simple analytical method of determining1-hydroxypyrene glucuronide in human urine by isotope dilution with ultra performance liquid chromatography-tandem mass spectrometry.

Abstract

The glucuronide conjugate of 1-hydroxypyrene (1-OHP-G) is a sensitive and reliable index biomarker for assessing low exposure to polycyclic aromatic hydrocarbons (PAHs). A simple method for determining 1-OHP-G in human urine with ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was established and applied to evaluate the exposure level of PAHs of pregnant women in a large sample size. After theurine sample was extracted with ethyl acetate, 0.2mL of the aqueous phase was diluted to 1.0mL with 5mmol/L ammonium acetate before injection. The chromatographic separation was performed on a C18 column with a gradient elution and identification was conducted on a tandem mass spectrometry with electrospray ionization in negative mode. 1-OHP-d9-G was used as an internal standard to improve precision. The method was validated and good linearity was obtained in the range of 0.1∼2.0ng/mL. The limit of detection (LOD) and the limit of quantification (LOQ) of 1-OHP-G were 0.015 and 0.051ng/mL. Intra-day and inter-day precision were 4.3 and 6.7%, respectively. The spiked recoveries were 79.4∼106% for urine samples. This method was rapid, sensitive, and very suitable for batch analysis of urine. Six hundred seventy-five urine samples of pregnant women from the cities of Fuzhou, Shenzhen, and Nanning of P.R. China were analyzed with the proposed method. The medians of 1-OHP-G concentration were 0.27μg/g.cr (n = 201), 0.30μg/g.cr (n = 212), and 0.51μg/g.cr (n = 262) for the cities of Fuzhou, Shenzhen, and Nanning, respectively. 1-OHP-G concentrations in urine samples of pregnant women from the cities of Fuzhou and Shenzhen in coastal areas were both significantly lower than that of Nanning City in inland region (p < 0.001). Graphical Abstract The internal standard 1-OHP-d9-G and 2.0 mL of ethyl acetate were added to 1.0 mL of urine sample, after vortex vibration and centrifugation the aqueousphasewas removed and diluted, and 5 μl of aliquot was analyzed by UPLC-MS/MS.