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Metabolite Identification and Quantification in Biofluids Using Liquid Chromatography Mass Spectrometry Open Access

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Other title
Subject/Keyword
Metabolomics, Metabolite identification, Quantification, Acylglycines, Methylmalonic acid, MycompoundID, Ultra-High Performance Liquid Chromatography, Mass spectrometry, Tandem mass spectrometry
Type of item
Thesis
Degree grantor
University of Alberta
Author or creator
Stanislaus, Avalyn E
Supervisor and department
Li, Liang (Chemistry)
Examining committee member and department
Waldron, Karen (Chemistry)
Curtis, Jonathan (Agricultural, Food and Nutritional Science)
Harynuk, James (Chemistry)
Hall, Dennis (Chemistry)
Klassen, John (Chemistry)
Department
Department of Chemistry
Specialization

Date accepted
2012-05-17T15:43:44Z
Graduation date
2012-11
Degree
Doctor of Philosophy
Degree level
Doctoral
Abstract
Metabolomics, one of the branches of systems biology, is the comprehensive measurement of all the endogenous metabolites in a biological system. It can be applied in the areas of biomarker discovery and diagnosis of critical diseases and disorders, and can significantly increase our understanding of the pathophysiology involved in development of acute diseases. For metabolomics to reach its true potential, there are many key areas that need to be addressed. This thesis focuses on the development and application of new mass spectrometric techniques aimed at improving several areas in metabolomics research: 1) detection and identification of compound classes, 2) enhancing the electrospray response and improving the chromatographic behavior of metabolites, 3) increasing sample throughput, 4) accurate quantification of endogenous metabolites and 5) expanding current databases used in metabolite identification. The initial work, the comprehensive analysis of acylglycines in human urine, illustrates the analytical challenges of detection and identification of trace levels of metabolites in urine. Mass spectrometric methods were optimized using fragmentation patterns and breakdown graphs, and putative identifications were made using a combination of diagnostic neutral losses. This method was successful in detecting and putatively identifying 43 new acylglycines. Quantification of these metabolites in human urine and plasma was performed using a new labeling strategy. Chemical modification was advantageous as it enhanced the electrospray response of these polar metabolites and provided a way to incorporate a stable isotope onto the molecule. Quantification using a new “surrogate matrix” strategy was also developed. A new fast LC method was developed for the analysis of several analytes in the vitamin B12 pathway in less than 36 seconds. This method was very useful in improving high-throughput and in the discovery of the role of ABC protein transporters in B12 metabolism. Finally, a web-based software called MyCompoundID was developed to expand the metabolome coverage of an existing database (HMBD) by generating metabolic products derived from 76 common biotransformations. This served to increase the number of entries from 8000 to more than 10 million.
Language
English
DOI
doi:10.7939/R39W0970G
Rights
Permission is hereby granted to the University of Alberta Libraries to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only. Where the thesis is converted to, or otherwise made available in digital form, the University of Alberta will advise potential users of the thesis of these terms. The author reserves all other publication and other rights in association with the copyright in the thesis and, except as herein before provided, neither the thesis nor any substantial portion thereof may be printed or otherwise reproduced in any material form whatsoever without the author's prior written permission.
Citation for previous publication
Lewis-Stanislaus, A.E., Li, L., 2010, Journal of American Society of Mass Spectrometry, 21, 2105 – 2116.

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