Solid Phase Extraction for 1-Hydroxypyrene as a Biomarker of Occupational Exposure to PAHs Prior to High Performance Liquid Chromatography

Abstract

Urinary 1-hydroxypyrene is frequently used as a major metabolite and biological indicator of the overall exposure to polycyclic aromatic hydrocarbons. In this study, solid phase extraction was appropriately conditioned with regard to sample pH, sample concentration, loading flow rate, elution solvent, washing solvent, sample volume and sorbent mass. Octadecyl silica cartridge (C18) was used as solid phase adsorbent and showed to be an efficient phase in simplifying sample preparation for1-hydroxypyrene. Methanol extracted analyte from spiked urine gave a clean sample for reverse-phase HPLC-florescence. In the developed solid phase extraction conditions (sample pH: 5, sample concentration: 10 μg/l, washing solvent: distilled water, eluent: methanol, sample volume: 200 ml sample flow rate: 10 ml/min), the extraction recovery exceeded 99.96%, achieving detection limit of 0.02 μg/l. The extraction factors (sample pH, sample concentration, washing solvent , eluent , sample volume and sample flow rate), were evaluated statistically and also the procedure was validated with three different pools of spiked urine samples at low, medium , and high sample concentrations and showed a good reproducibility over six consecutive days as well as six within-day experiments. All coefficients of variations were less than 3.1%. Finally, urinary 1-hydroxypyrene of industrially exposed workers was also measured, using the appropriate conditions obtained in this study, in which, the amount of the compound of interest in the total exposed subjects was significantly higher than those of non-exposed.