Glucuronidation metabolomic fingerprinting to map host-microbe metabolism.

Glucuronidation is an important detoxification pathway that operates in balance with gastrointestinal microbial β-glucuronidase (GUS) activity, which can regenerate bioactive metabolites from their glucuronidated forms. How this host-microbe interaction shapes the distribution and pool of glucuronidated metabolites (i.e., the glucuronidome) remains poorly understood. In this study, we employed pattern-filtering data science approaches in conjunction with untargeted LC-MS/MS metabolomics to map the glucuronidome in urine, serum, and colon/fecal samples from gnotobiotic and conventional mice, and in humans. We find that microbial colonization and GUS activity compress the colonic glucuronidome and expand urinary glucuronidome diversity, revealing a compartmental redistribution of glucuronidated metabolites. Reverse metabolomics of known glucuronidated chemicals and glucuronidation pattern filtering searches in public metabolomics datasets exposed the diversity of glucuronidated metabolites in human and mouse ecosystems. In summary, we present a glucuronidation fingerprint resource that provides broader access to and analysis of the glucuronidome. Together, this work establishes a scalable analytical framework and provides mechanistic insight into how microbial activity reshapes systemic glucuronidation, with implications for drug metabolism, diet-microbe interactions, and biomarker discovery.