Simultaneous profiling of mercapturic acids, glucuronic acids, and sulfates in human urine

Humans are constantly exposed to both naturally-occurring and anthropogenic chemicals. Targeted mass spectrometry approaches are frequently used to measure a small panel of chemicals and their metabolites in environmental or biological matrices, but methods for comprehensive individual-level exposure assessment are limited. In this study, we applied an integrated library-guided analysis (ILGA) with ultraperformance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF/MS) to profile phase II metabolites, specifically mercapturic acids (MAs), glucuronic acids (GAs), and sulfates (SAs) in human urine samples (n = 844). We annotated 424 metabolites (146 MAs, 171 GAs, 107 SAs) by querying chromatographic features with in-house structural libraries, filtering against fragmentation patterns (common neutral loss and ion fragment), and comparing mass spectra with in-silico fragmentations and external spectral libraries. These metabolites were derived from over 200 putative parent compounds of exogenous and endogenous sources, such as dietary compounds, benzene/monocyclic substituted aromatics, pharmaceuticals, polycyclic aromatic hydrocarbons, bile acids/bile salts, and 4-hydroxyalkenals associated with lipid peroxidation process. Further, we performed statistical analyses on 214 metabolites found in more than 75% of samples to examine the association between metabolites and demographic characteristics using integrated network analysis, principal component analysis (PCA), and multivariable linear regression models. The network analysis revealed four distinct communities of 37 positively correlated metabolites, and the PCA (using the 37 metabolites) presented 4 principal components that meaningfully explained at least 80% of the variance in the data. The multivariable linear regression models showed some positive and negative associations between metabolite profiles and certain demographic variables (e.g., age, sex, race, education, income, and tobacco use).