Mass Spectrometric Strategies and Applications for High-Efficiency Global Metabolomics

• Main Author: Rubio, Vanessa Y
• Format: Dissertation
• Language: English
• Published: ProQuest Dissertations & Theses 01-01-2020
• Subjects: Analytical chemistry
• ISBN: 9798357544841

In clinical metabolomics, when the goal is predicting or evaluating systemic health and disease risk, having the ability to characterize individual phenotypic expression beyond the scope of the genome and proteome is significant. As a high-resolution platform, metabolomics performance using ultra high-performance liquid chromatography coupled to high-resolution mass spectrometry (UHPLC-HRMS) can be placed at the forefront of biomarker discovery for clinical research. However, current metabolomic workflows are considerably fragmented with analyses being performed as separate tasks. We evaluated a combined metabolomics acquisition workflow for the quantification of plasma or serum metabolites in a global metabolomics experiment. This was done by the introduction of a wide-isolation MS/MS acquisition method in a global metabolomics workflow for the analysis of intestinal microbiota composition and its impact on the metabolome as a result of dietary stress. Implementation of a wide-isolation MS/MS technique reduces the number of analytical scanning events performed by the mass spectrometer to compose a spectral depiction of a metabolite within a chromatographic peak. This combined workflow was expanded to include the utilization of a unique isotopic labeling technique. In addition to gaining the ability to perform quantitation, this isotopic labeling technique employs an enrichment of 5% and 95% 13C metabolites for experimental and control group applications, respectively. This generates a unique mass spectral pattern that allows for differentiation of peaks of biological significance rather than artifactual noise. Given this combined workflow, the purpose of our study herein was to expand the effort to understand the metabolic differences between the two short stature populations, Laron and Guevara-Rosenbloom syndromes (LS and GRS), with unique relationships to insulin resistance making them phenotypically similar, but metabolically different. The commonality between the two unassociated cohorts and diabetes provides an interesting opportunity to investigate the metabolome and its impact as a result of these unique diseases and their respective responses to glucose. Finally, we have shown a novel application of paper spray by using direct analysis for global metabolomics acquisition within a 30-sec analytical time and automated data processing that could rival traditional LC-MS global metabolomics workflows.