Determination of 1-hydroxypyrene in human urine by high-performance liquid chromatography with fluorescence detection using a deuterated internal standard

A high-performance liquid chromatographic method has been developed for the quantification of 1-hydroxypyrene (1-OHP) in human urine using deuterated 1-hydroxypyrene ([H-2(9)]1-OHP) as an internal standard with fluorescence detection. [H-2(9)]1-OHP was prepared enzymatically from deuterated pyrene ([H-2(10)]Pyr) with cytochrome P450 1A1. It eluted immediately prior to non-deuterated 1-OHP on alkylamide-type reversed-phase columns and had nearly the same fluorescence characteristics as non-deuterated 1-OHP. The detection limit was 0.1 mug/L and the calibration range was from 1 to 100 nmol/L. Urine sample treatment involved enzymatic hydrolysis followed by solid-phase extraction using Sep-Pak C-18 cartridges. The proposed method was used to determine urinary 1-OHP in smokers and non-smokers. (C) 2002 Elsevier Science BV All rights reserved.