Quantitative exposomics targeting over 200 toxicants and key biomarkers at picomolar concentrations

The exposome encompasses environmental exposures throughout life and significantly impacts health and disease. Ex-posure chemicals, present at trace levels, are frequently quantified using targeted LC-MS/MS. Many existing methods are limited to a narrow range of analyte classes or lack sufficient sensitivity for exposomic analyses and applicability to large sample cohorts for exposome-wide association studies (ExWAS) remain to be demonstrated. Here, we present a scalable, fit-for-purpose next-generation human biomonitoring (HBM) workflow for analyzing >230 biomarkers in urine, plasma, and serum using solid-phase extraction in 96-well plates and LC-MS/MS. Moreover, a new conceptual framework for vali-dation criteria of assays designed to analyze large panels of highly diverse compounds at trace levels is proposed. Meth-od robustness was evaluated, demonstrating suitable extraction recovery (60-130%), matrix effects (SSE, 60-130%), inter-/intra-day precision (RSD <30%), and high sensitivity (limit of detection, 0.015-50 pg/mL) for 60-80% of the ana-lytes across the investigated matrices. To showcase the method's applicability in epidemiological studies, 200 urine samples from pregnant women in a longitudinal pregnancy cohort were analyzed. More than 140 analytes were detected, several for the first time in an U.S. urinary biomonitoring study. With its broad analyte coverage, ultimate sensitivity, robustness, and high sample throughput, this method meets performance requirements for future large-scale ExWAS stud-ies in public and personalized prevention.