Development and Applications of Chemical Isotope Labeling Methods for Metabolome Sample Normalization and Liquid Chromatography-Mass Spectrometry-Based Metabolomics

• Author / Creator

  Wu, Yiman

• Metabolomics refers to the global study of the metabolome of a biological system where comprehensive and systematic profiling of all metabolites in a given biological sample (i.e., the metabolome) is performed. As the end product of biological processes, the status of metabolites could reflect the physiological state of an organism and provide valuable and complementary information to the genomics, transcriptomics and proteomics data. However, as a relatively new research field, there are some challenges associated with LC-MS-based metabolomics. For example, the total metabolite concentrations often vary from sample to sample, which may complicate relative quantification of the metabolome changes in comparative studies. Also, the high complexity of biological samples makes it difficult to quantitatively extract all metabolites and sensitively detect low abundance metabolites. Towards these challenges, the objective of my research was two-fold. Firstly, a sample normalization method based on UV absorbance measurement was developed to allow quantification of the total concentration of chemically labeled metabolites (Chapters 2 and 3). This method can be readily applied to any type of biological samples for effective correction of sample concentration variations. Application of this normalization strategy was demonstrated on human urine and bacterial cell extracts with significantly reduced inter-group variations and improved statistical results. Secondly, differential isotope dansylation labeling LC-MS metabolomics workflows were developed for various biological matrices including bacterial cells, plasma and cerebrospinal fluid (Chapter 4 to 6). This labeling chemistry targets at the amine and phenol sub-metabolome with improved chromatographic separation and detection sensitivity. The developed protocols enabled the detection of over 1000 putative metabolites in each biological sample, and have been applied to bacterial differentiation and biofluid disease biomarker discovery studies. Overall, these research activities have demonstrated enhanced analytical performance and capability of LC-MS-based metabolomics methods for real biological applications.

• Subjects / Keywords

  • Metabolomics
  • Isotope labeling
  • LC-MS

• Graduation date

  Fall 2015

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/R32R3PB4W

• License

  This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.