Quantitative Exposomics Targeting over 200 Toxicants and Key Biomarkers at the Picomolar Level.

The exposome encompasses environmental exposures throughout life and significantly impacts health and disease. Exposure 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) remains 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 complementary conceptual framework for validation criteria of assays designed to analyze large panels of highly diverse compounds at trace levels is proposed. Method robustness was evaluated, demonstrating extraction recovery (60-130%), matrix effects (SSE, 60-130%), inter-/intraday precision (RSD <30%), and high sensitivity (limit of detection <0.1 ng/mL) for 59-80% of the analytes across the investigated biological 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 100 analytes including PFAS, drugs, air pollutants, pesticides, flame retardants, mycotoxins, industrial products, food processing contaminants, plastics-related chemicals, and phytotoxins, were detected, several for the first time in a U.S. urinary biomonitoring study. With its broad analyte coverage, ultimate sensitivity, robustness, and high sample throughput, this method meets the performance requirements for future large-scale ExWAS applications in public and personalized prevention research.