Simultaneous Determination of 6 l-Arginine Metabolites in Human and Mouse Plasma by Using Hydrophilic-Interaction Chromatography and Electrospray Tandem Mass Spectrometry

Abstract

BACKGROUND Macrophages and related cells are important cellular mediators of the innate immune system and play important roles in wound healing and fibrosis. Flux through different l-arginine metabolic pathways partially defines the functional behavior of macrophages. Methods to measure metabolites within the nitric oxide synthase/arginase pathways could provide insights into local and systemic inflammatory processes. METHODS A targeted metabolomics approach was developed by using hydrophilic-interaction liquid chromatography and electrospray ionization–tandem mass spectrometry to simultaneously measure l-arginine, asymmetric dimethylarginine, symmetric dimethylarginine, l-citrulline, l-ornithine, and l-proline in plasma from humans and mice. RESULTS All analytes were quantifiable in human and mouse plasma with a small volume (25 μL), minimal sample preparation, and no derivatization. Patients with high plasma concentrations of C-reactive protein and mice with acute inflammation induced by lipopolysaccharide had significant reductions of arginine metabolites in plasma compared with controls. CONCLUSIONS This new assay uses plasma metabolomic measurements to help provide new insights into metabolic changes coupled to the innate immune response. We identified significant changes in arginine metabolism in both humans and mice following an inflammatory stimulus. These changes were associated with decreased plasma arginine metabolite concentrations and increased methylated arginine concentrations.