A Tutorial on Using Untargeted Metabolomics Data of Human Excreta to Investigate Drug Excretion and Wastewater Entry

Environmental scientists are increasingly monitoring therapeutic drugs and their metabolites in water systems, requiring knowledge of human drug metabolism and excretion. Many published studies, however, rely on data from small-scale human metabolism trials, typically involving around six (healthy, young, male) volunteers. Their generalizability to real-world drug users may be limited, potentially biasing environmental monitoring efforts. Here, we leveraged untargeted LC-SWATH/MS pharmacometabolomics data from 283 potential living kidney donors and 688 kidney transplant recipients to characterize the 24 h urinary excretion profiles of two widely used diuretics frequently monitored in wastewater, hydrochlorothiazide and furosemide. Both are expected to be excreted largely unchanged, which our analyses confirmed. For hydrochlorothiazide, however, we also identified (using reference standards) the previously underreported transformation products chlorothiazide and salamide. These findings highlight the relevance and capability of using untargeted metabolomics data from human excreta to provide insights from large, real-world cohorts into which chemicals enter wastewater systems, with both drugs serving as exemplary case studies for analogous analyses of other drugs. In particular, the qualitative information obtained (e.g., accurate mass, retention time, fragment spectra) may inform targeted biomonitoring and highlight cases where consensus-based estimates of excreted drug or metabolite fractions are overestimated.