Improvement on recovery and reproducibility for quantifying urinary mono-hydroxylated polycyclic aromatic hydrocarbons (OH-PAHs)

Abstract

Efficient and reproducible measurements of multiple polycyclic aromatic hydrocarbon (PAH) metabolites in urinary samples are required to evaluate the complex health effects of PAH exposure. Here, we demonstrate a highly practical, automated off-line solid-phase extraction (SPE) of deconjugated hydroxylated PAHs followed by LC-MS/MS to simultaneously measure eight mono-hydroxylated PAH compounds: 1-hydroxynaphthalene, 2-hydroxynaphthalene, 2-hydroxyfluorene, 1-hydroxyphenanthrene, 2&3-hydroxyphenanthrene, 4-hydroxyphenanthrene and 1-hydroxypyrene. Initially, we observed low recovery rates (e.g., 16% for 1-hydroxypyrene) when using previously published methods. We optimized the procedure by choosing polymeric absorbent-based cartridges, automating the sample loading step by diluting samples with 15% methanol/sodium acetate, and most importantly, replacing acetonitrile with methanol as the eluting solvent. Optimized sample preparation has improved the recovery rates to more than 69% for analytes of interest. This improvement led to higher method sensitivity and detection frequency, especially for 1-hydroxypyrene, in all of 100 urine samples collected in the New York City site of the Legacy Girls Study. The limits of detection ranged from 7.6 pg/mL to 20.3 pg/mL using 1 mL of urine, compared to the 2 mL required in CDC, method 09-OD. The average coefficients of variance of quality control samples (n = 60) ranged between 7 and 21%; variance of repeated measurements (n = 45) was less than 10%. This efficient and reliable method for measuring PAH metabolites will greatly benefit epidemiology studies and biomonitoring programs.