Simultaneous determination of etomidate and its derivatives in wastewater with online SPE-HPLC-MS/MS

Etomidate (ETM), a short-acting intravenous sedative, has been increasingly abused, particularly among young people in China and other parts of East Asia, leading to severe health issues and prompting regulatory control. Along with the control of ETM, several new ETM derivatives have emerged as substitutes. High-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) is a widely used analytical technique for identifying and quantifying abused substances in wastewater, which faces challenges due to low analyte concentrations and matrix interference. This study introduces an online solid-phase extraction (SPE)-HPLC-MS/MS method for the simultaneous determination of ETM, its major metabolite etomidate acid, and eight ETM derivatives in wastewater. The HPLC-MS/MS conditions, including column selection, mobile phase composition, and sample pretreatment conditions, were optimized, respectively. Method validation demonstrated excellent linearity across calibration ranges of 1 to 1000 ng L-1, with limits of detection (LOD) and limits of quantification (LOQ) ranging from 0.2 to 3 ng L-1 and 1 to 10 ng L-1, respectively. The method exhibited satisfactory accuracy with recovery values ranging from 86.6% to 108.6%, and high precision with relative standard deviations between 4.8% and 13.9%. The validated method was successfully applied to analyze 40 authentic wastewater samples, of which four tested positive. The detected compounds included ETM (8.2 and 13.4 ng L-1 in two samples), metomidate (MEM) (6.8 ng L-1), and isopropoxate (IPX) (14.5 ng L-1). These findings make the method suitable for accurate and rapid monitoring of ETM and its derivatives in wastewater, providing a robust analytical tool for future wastewater-based epidemiology investigations.