First identification of a diuretic, hydrochlorothiazide, in hair: Application to a doping case and interpretation of the results

Abstract

An athlete contested an adverse analytical finding involving hydrochlorothiazide, and requested our laboratory for testing his hair. As there is no reference in the literature about identification of hydrochlorothiazide in hair, several volunteers were first enrolled (4 after a single 25 mg administration and 10 with daily therapeutic treatment). A specific method was developed by ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS. Hair samples were decontaminated with dichloromethane and 30 mg were incubated in buffer at pH 7.0 for 15 hours at 50°C. Then, 5 mL ethyl acetate was added for extraction. Linearity was observed for hydrochlorothiazide concentrations ranging from 5 to 2000 pg/mg. The limit of quantification was 5 pg/mg. The coefficients of variation (CVs) of repeatability and matrix effect were lower than 20%. Analysis of the 0–2-cm segment of the 4 volunteers having received a single dose, collected 1 month after administration, was negative at the limit of quantification. The hair of the 10 patients (proximal 2 cm) on daily treatment was positive with concentrations ranging from 12 to 1845 pg/mg, with no correlation between daily dose and concentration. The athlete's hair tested positive for hydrochlorothiazide at 36 pg/mg in the segment corresponding to the period of the urinary control. Since a single exposure to hydrochlorothiazide is not detectable in hair and based on the results of the patients on daily treatment, the concentration found in the athlete has been interpreted as corresponding to repeated exposures, where it was not possible to establish the dosage and the frequency.