Biology subjects

Gouda, H.

Publications and source records attributed to Gouda, H..

9 recordsLinked to original sources

bioRxiv2026

Widespread detection of Polyethylene glycol reveals chronic human exposure through pharmaceutical drugs

Polyethylene glycol (PEG) is a synthetic polymer ubiquitous in pharmaceuticals, personal care products, food additives, and industrial manufacturing. Despite its widespread use and potential importance as an exposure chemical, the prevalence of PEG exposure and its excretion in human populations remain largely uncharacterized. Moreover, PEG detected in human biofluids is frequently assumed to arise from analytical contamination during sample preparation, potentially obscuring its contribution to the human exposome and confounding metabolic phenotyping studies. Here, we show that PEG is as component of the human xenobiotic exposome and is further metabolized into PEG hydoxy acid and diacid metabolites in humans. We further find that PEG exposure is associated with alterations in microbiome composition and short-chain fatty acid metabolism, suggesting its biological impact of its exposure. We identify PEG exposure in approximately 2.3% of publicly available metabolomics data files and provide a reusable 85,484 candidate PEG and PEGylated MS/MS spectral library for future use for the metabolomic community. Together, these findings establish PEG signal in human biofluids can reflect genuine exposure, and that PEG exposure is neither metabolically inert nor biologically silent.

biochemistry↗

bioRxiv2026

Alkamines reveal a hidden layer of steroid and drug metabolism

Biomedical research overlooks most genes in favor of a well-studied minority, yet whether analogous blind spots exist in metabolomics remains unknown. We show that reductive amination, forming secondary amines from aldehydes or ketones and amines, generates a previously hidden class of metabolites we term alkamines. Multiplexed synthesis of 8,475 alkamines combined with MS/MS searches across 1.7 billion spectra identified 1,626 candidates across multiple species and organs. Of these, 56 were confirmed in biological samples, including 27 steroid- and 12 drug-derived alkamines matching prescription patterns. Notably, 77% of synthesized alkamines are absent from PubChem. This combinatorial logic likely explains why alkamines have evaded detection and suggests drug metabolism frameworks substantially underestimate drug-derived metabolite diversity. Reductive amination is an overlooked route modifying steroids, bile acids, and xenobiotics.

biochemistry↗

bioRxiv2026

Pan-Metabolomics Repository Mapping of the Carnitine Landscape

Carnitines are a structurally diverse class of metabolites formed by conjugation of L-carnitine with fatty acids, amino acids, xenobiotics, and microbial metabolites. They play roles in transport, mitochondrial and peroxisomal metabolism, detoxification, and systemic signaling, yet their chemical diversity remains incompletely defined. We applied a pan-repository data mining strategy of LC-MS/MS data across GNPS/MassIVE, MetaboLights, and Metabolomics Workbench using MassQL diagnostic fragment ion filtering to systematically extract acylcarnitine spectra. This yielded a library of 34,222 unique MS/MS spectra representing 2,857 atomic compositions, corresponding to 3,872,050 detections. These datasets provide an MS/MS library for annotation, discovery, and contextualization of acylcarnitines, enabling identification of previously unknown carnitines, such as dihydroferulic acid conjugated carnitines and supporting future exploration of this metabolite class across host metabolism, diet, microbial activity, pharmacological exposures, and metabolic dysregulation.

bioinformatics↗

bioRxiv2026

Pan-repository analysis reveals a drug-activating function of microbial bile acid conjugation

Microbially modified bile acids shape host physiology by regulating nutrient absorption, glucose homeostasis, circadian rhythms and thermoregulation. Here we identify a previously unrecognized drug-activating function of microbial bile acid conjugation. By systematically mining human LC-MS/MS datasets across public repositories and linking uncharacterized bile acid spectra to health-associated metadata, we discovered conjugates of the >75-year-old anti-inflammatory drug 5-aminosalicylic acid (5-ASA) with primary and secondary bile acids, including cholic, deoxycholic and lithocholic acids. These bile acid-drug conjugates were detected specifically in individuals treated with 5-ASA or its prodrugs. Multiple gut bacteria, including members of the Bacteroidota and Bacillota, generated cholyl-5-ASA in vitro, and bile salt hydrolase-associated transaminase activity was required for conjugate formation. In a mouse model of colitis, cholyl-5-ASA was associated with reduced intestinal inflammatory pathology and showed markedly enhanced activation of PPAR-{gamma} in cell-based reporter assays compared with 5-ASA alone. Consistent with this activity, cholyl-5-ASA elicited selective immunophenotypic changes in CD4 T cells in vitro, including increased Foxp3+ regulatory T cells. Together with prior evidence that 5-ASA efficacy depends on the microbiome, these findings support a model in which microbial bile acid conjugation represents a key activation step for 5-ASA therapy. More broadly, this work demonstrates how pan-repository metabolomics can uncover previously unrecognized microbiome-dependent chemical functions with direct therapeutic relevance.

biochemistry↗

bioRxiv2026

Navigating the conjugated metabolome

Lifes chemical diversity far exceeds current biochemical maps. While metabolomics has catalogued tens of thousands of small molecules, conjugated metabolites, formed when two or more molecular entities are covalently fused through amidation, esterification, or related chemistries, remain underexplored. These molecules can act as microbial signals, detoxification intermediates, or endogenous regulators. Here, we mined 1.32 billion MS/MS spectra across public metabolomics repositories using reverse spectral searching coupled with delta-mass inference to map conjugation events. We generated structural hypotheses for 24,227,439 MS/MS clusters. From these, we inferred 217,291 substructure pairs with dual spectral support and 3,412,720 candidate conjugates with single-match support. Predictions span host-microbe co-metabolites, diet-derived conjugates, and drug-derived species, including drug-ethanolamine and creatinine conjugates with altered bioactivities. We also uncover a family of steroid-phosphoethanolamine conjugates. Fifty-five conjugates were matched by MS/MS of synthetic standards for this work, with 27 additionally supported by retention time matching in biological samples. Guidance on how to leverage this resource is also provided. Together, these results deliver a pan-repository map of potential conjugation chemistry, establish a resource for structural discovery and MS/MS annotation, and offer a scalable framework to explore the scope and diversity of the conjugated metabolome.

bioinformatics↗

bioRxiv2026

A searchable metadata network graph for microbiome metabolomics

Establishing the biological context of microbial metabolites remains a major challenge. We present microbiomeMASST, a metadata-driven network graph that maps metabolites across 467 available datasets with 144,424 mass spectrometry files from humans, animals, and microbial culture systems. MicrobiomeMASST integrates monocultures, synthetic communities, and host-associated samples across multiple body sites and plants. MS/MS spectra can be queried to trace occurrence across hosts, experimental conditions, and interventions, enabling cross-study integration. We demonstrate this framework by contextualizing microbial-conjugated bile acids and interrogating microbiome-mediated drug metabolism. Screening gut bacteria revealed deprolylation of the angiotensin-converting enzyme (ACE) inhibitor prodrug enalapril. Using microbiomeMASST, we traced this metabolite across human cohorts, microbial isolates, environmental samples, and in Gorilla gorilla. Structural modeling and enzymatic assays showed that microbial deprolylation abolishes ACE inhibition, thereby inactivating its therapeutic effect. Together, microbiomeMASST links MS/MS spectra to biological context, converting isolated observations into an interpretable microbiome map for cross-study analysis.

biochemistry↗

bioRxiv2026

Community Curation of Microbial Metabolites Enables Biological Insights of Metabolomics Data

Microbial metabolites play a critical role in regulating ecosystems, including the human body and its microbiota. However, understanding the physiologically relevant role of these molecules, especially through liquid chromatography tandem mass spectrometry (LC-MS/MS)-based untargeted metabolomics, poses significant challenges and often requires manual parsing of a large amount of literature, databases, and webpages. To address this gap, we established the Collaborative Microbial Metabolite Center knowledgebase (CMMC-KB), a platform that fosters collaborative efforts within the scientific community to curate knowledge about microbial metabolites. The CMMC-KB aims to collect comprehensive information about microbial molecules originating from microbial biosynthesis, drug metabolism, exposure-related molecules, food, host-derived molecules, and, whenever available, their known activities. Molecules from other sources, including host-produced, dietary, and pharmaceutical compounds, are also included. By enabling direct integration of this knowledgebase with downstream analytical tools, including molecular networking, we can deepen insights into microbiota and their metabolites, ultimately advancing our understanding of microbial ecosystems.

bioinformatics↗

bioRxiv2025

Charting the Undiscovered Metabolome with Synthetic Multiplexing

Most molecular features detected in untargeted metabolomics remain uncharacterized due to the limited scope of existing spectral reference libraries. We synthesized >100,000 biologically inspired compounds using multiplexed reactions, of which 91% were absent from existing structural databases, and searched the resulting MS/MS library across >1.7 billion public spectra, increasing annotation rates by 17.4%. This approach revealed previously undescribed exposure-derived metabolites, including ibuprofen-carnitine. Because ibuprofen has been linked to rhabdomyolysis, reduced mitochondrial function, and impaired muscle recovery in carnitine-limited contexts, we investigated the functional relevance of this conjugate. Ibuprofen-carnitine reduced carnitine transport via the OCTN2 transporter, and in a postpartum mouse muscle injury model, ibuprofen delayed muscle repair that could be rescued by carnitine supplementation, with urinary ibuprofen-carnitine:carnitine ratios tracking this effect. These findings support a hypothesis whereby NSAID-carnitine conjugates compete for carnitine transport, impairing energy metabolism and muscle recovery in susceptible individuals. Synthetic multiplexing thus provides a scalable route to annotate the dark metabolome and generate experimentally testable biological hypotheses.

biochemistry↗

bioRxiv2023

Architecture of the human G-protein-methylmalonyl-CoA mutase nanoassembly for B12 delivery and repair

G-proteins function as molecular switches to power cofactor translocation and confer fidelity in metal trafficking. MMAA, a G-protein motor, together with MMAB, an adenosyltransferase, orchestrate cofactor delivery and repair of B12-dependent human methylmalonyl-CoA mutase (MMUT). The mechanism by which the motor assembles and moves a >1300 Da cargo, or fails in disease, are poorly understood. Herein, we report the crystal structure of the human MMUT-MMAA nanomotor assembly, which reveals a dramatic 180{degrees} rotation of the B12 domain, exposing it to solvent. The nanomotor complex, stabilized by MMAA wedging between two MMUT domains, leads to ordering of the switch I and III loops, revealing the molecular basis of mutase-dependent GTPase activation. The structure explains the biochemical penalties incurred by methylmalonic aciduria-causing mutations that reside at the newly identified MMAA-MMUT interfaces.

biochemistry↗