Development of a novel HPLC-HRMS method for quantitative analysis of resorcinol in urine: Application to...

Resorcinol is a widespread substance used in a large variety of manufacturing industries, including cosmetics, with endocrine-disrupting activity on the thyroid function. The aim of the present study was to develop and validate a sensitive, selective and robust method to quantify resorcinol in urine and thereby assess hairdressers' occupational exposure. As resorcinol is mainly excreted in urine as glucuronide or sulfate forms, the first step consisted in hydrolyzing urine samples with a β-glucuronidase-arylsulfatase enzyme for 16 h. Then, after cleaning with a supported-liquid extraction cartridge, the samples were derivatized with dansyl chloride to improve signal and signal-to-noise ratio. Analysis was carried out using an accurate high-resolution liquid chromatography-mass spectrometry instrument on a Kinetex Biphenyl analytical column. Particular attention was paid to the chromatographic separation of resorcinol from its two isomers, catechol and hydroquinone, also present in urine. Acquisition was performed in positive ESI mode, at m/z 577.14615 for dansylated resorcinol and m/z 581.17126 for dansylated resorcinol-d₄, with respective retention times of 8.63 and 8.60 min. The method passed all the performance tests included in the validation process. The lowest limit of quantification (LLOQ) was 0.3 µg/L resorcinol, which was sufficient to quantify resorcinol in all samples tested. The calibration curves were linear from LLOQ to 2000 µg/L, with coefficients of determination R² ranging from 99.82 % to 100 %. The method was accurate, reaching 95.6 to 101.7 % of target intraday and 99.8 to 105.0 % interday, and precise with RSDs between 0.88 and 1.99 % intraday and with RSDs between 1.75 and 8.65 % in interday assessments. It also proved robust, with a matrix effect of 8.25 %. Resorcinol stability was determined by studying long-term stability at -20 °C for sample storage up to 6 months, short-term stability (at + 20 °C and + 4°C for possible short-term storage), freeze-thaw cycles, and post derivatization stability. This method was successfully applied on samples from 17 women working as hairdressers. Urinary resorcinol concentrations ranged from 2 µg/L to 1824 µg/L (6 to 4475 µg/g creatinine) and were in line with those reported in the literature.