Simultaneous quantification of 18 serum steroids and catecholamines via LC-MS/MS: A derivatization-free method using...

The quantification of serum steroids and catecholamines is essential for the clinical diagnosis of adrenal diseases and tumors. Due to their distinct physicochemical properties and low serum concentrations, existing assays typically quantify these analytes separately using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Simultaneous quantification offers significant advantages, including enhanced diagnostic accuracy and improved sample comparability. Here, we developed a sensitive LC-MS/MS method to simultaneously measure 6 catecholamines, 10 steroids, and 2 other neuroendocrine hormones in a single run. Sample preparation involved a simple one-step solid-phase extraction without derivatization. All analytes were chromatographically separated on an HSS PFP column within 14 minutes. The method exhibited excellent linearity (r²>0.9930) with limits of quantification ranging from 0.0006 to 2 ng/mL. Extraction recoveries and matrix effects ranged from 82.6% to 117.4% and 82.0% to 107.6%, respectively. Intra- and inter-day reproducibility was validated, with coefficients of variation (CVs) between 0.6% and 11.4%. The method was successfully applied to serum samples from healthy volunteers in an exposure-intervention study. Detection rates exceeded 95% for most analytes, with the exception of dihydrotestosterone and dehydroepiandrosterone. The developed method demonstrates improved sensitivity and throughput compared to existing assays, serving as a valuable quantitative tool for large-scale epidemiological studies and clinical practice.