Grant: Discovery Grants Program - Individual

Metabolomics is the science of large-scale analysis of all small molecule metabolites present in a given biospecimen in order to study dynamic processes occurring in living systems. Metabolomics is increasingly used in biomedicine, food science, agriculture and environmental sciences. However, the analytical methods for comprehensive measurement of hundreds of metabolites require better coverage of low abundance and signalling metabolites including their in-depth characterization across various types of biospecimens; lack adequate validation and harmonization and cannot be easily adopted to microscale quantities of tissue. Thus, the long-term objective of the proposed research program is to establish novel analytical methods in global and targeted metabolomics with the primary focus on improving the detection and identification of low abundance and unstable metabolites in biological fluids and tissues using  liquid chromatography - high-resolution mass spectrometry (LC-HRMS). To achieve this objective, my 5-year research program will establish novel LC-HRMS assays for poorly characterized NO2-modified lipids. Rigorous validation of the developed methods will be carried out including systematic evaluation of metabolite stability, recovery and ionization suppression effects, all of which remain critical issues severely impacting the quality of many current studies. Next, we will develop a complementary series of novel untargeted LC-MS metabolomics workflows focusing on class-specific extraction, isomer separation and MS analysis of phase II metabolites, to introduce for the first time deep comprehensive profiling of these species and expand the coverage of relevant spectral libraries for metabolite identification. Finally, we will perform fundamental studies to examine electrospray ionization behaviour of various metabolite classes, including systematic studies of ionization suppression and adduct formation of selected metabolites within and across classes in various biospecimen types. These studies will also be combined with standard addition calibration to probe the influence of the adduct selection on the overall method accuracy for a given lipid. These investigations will support better selection of internal standards and mobile phase additives to increase sensitivity and accuracy of targeted and untargeted metabolomics. Collectively, this research program will establish new analytical approaches to measure currently "unmeasurable" low abundance metabolites and will advance the fields of metabolomics and exposomics by (i) increasing metabolite coverage of modified lipids and phase II metabolites including the expansion of public metabolite databases and comprehensive phase II metabolism mapping and (ii) controlling metabolite susceptibility to ionization suppression and adduct formation to guide the design of more accurate metabolomics methods and aid in harmonization efforts.