Improving the intestinal lipidome coverage in a gnotobiotic mouse model using UHPLC-MS-based approach through optimization of mobile phase modifiers and column selection.

Lipidomics is focusing on the screening of lipid species in complex mixtures using mass spectrometry-based approaches. In this work, we aim to enhance the intestinal lipidome coverage within the Oligo-Mouse-Microbiota (OMM12) colonized mouse model by testing eight mobile phase conditions on five reversed-phase columns. Our selected mobile phase modifiers included two ammonium salts, two concentrations, and the addition of respective acids at 0.1 %. We compared two columns with hybrid surface technology, two with ethylene bridged hybrid technology and one with core-shell particles. Best performance was attained for standards and intestinal lipidome, using either ammonium formate or acetate in ESI(+) or ammonium acetate in ESI(-) for all column technologies. Notably, a concentration of 5 mM ammonium salt showed optimal results for both modes, while the addition of acids had a negligible effect on lipid ionization efficiency. The HST BEH C18 column improved peak width and tailing factor parameters compared to other technologies. We achieved the highest lipid count in colon and ileum content, including ceramides, phosphatidylethanolamines and phosphatidylcholines, when using 5 mM ammonium acetate in ESI(-). Conversely, in ESI(+) 5 mM ammonium formate demonstrated superior coverage for diacylglycerols and triacylglycerols.

High-resolution mass spectrometry identifies delayed biomarkers for improved precision in acetaminophen/paracetamol human biomonitoring.

Paracetamol/acetaminophen (N-acetyl-p-aminophenol, APAP) is a top selling analgesic used in more than 600 prescription and non-prescription pharmaceuticals. To study efficiently some of the potential undesirable effects associated with increasing APAP consumption (e.g., developmental disorders, drug-induced liver injury), there is a need to improve current APAP biomonitoring methods that are limited by APAP short half-life. Here, we demonstrate using high-resolution mass spectrometry (HRMS) in several human studies that APAP thiomethyl metabolite conjugates (S-methyl-3-thioacetaminophen sulfate and S-methyl-3-thioacetaminophen sulphoxide sulfate) are stable biomarkers with delayed excretion rates compared to conventional APAP metabolites, that could provide a more reliable history of APAP ingestion in epidemiological studies. We also show that these biomarkers could serve as relevant clinical markers to diagnose APAP acute intoxication in overdosed patients, when free APAP have nearly disappeared from blood. Using in vitro liver models (HepaRG cells and primary human hepatocytes), we then confirm that these thiomethyl metabolites are directly linked to the toxic N-acetyl-p-benzoquinone imine (NAPQI) elimination, and produced via an overlooked pathway called the thiomethyl shunt pathway. Further studies will be needed to determine whether the production of the reactive hepatotoxic NAPQI metabolites is currently underestimated in human. Nevertheless, these biomarkers could already serve to improve APAP human biomonitoring, and investigate, for instance, inter-individual variability in NAPQI production to study underlying causes involved in APAP-induced hepatotoxicity. Overall, our findings demonstrate the potential of exposomics-based HRMS approach to advance towards a better precision for human biomonitoring.