| Autor: |
Papadimitropoulos MP; Metabolic Engineering and Systems Biology Laboratory, Institute of Chemical Engineering Sciences, Foundation for Research & Technology - Hellas (FORTH/ICE-HT), Patras, 26504, Greece.; Division of Genetics, Cell & Developmental Biology, Department of Biology, University of Patras, Patras, 26500, Greece., Vasilopoulou CG; Metabolic Engineering and Systems Biology Laboratory, Institute of Chemical Engineering Sciences, Foundation for Research & Technology - Hellas (FORTH/ICE-HT), Patras, 26504, Greece.; Human and Animal Physiology Laboratory, Department of Biology, University of Patras, Patras, 26500, Greece., Maga-Nteve C; Metabolic Engineering and Systems Biology Laboratory, Institute of Chemical Engineering Sciences, Foundation for Research & Technology - Hellas (FORTH/ICE-HT), Patras, 26504, Greece.; School of Medicine, University of Patras, Patras, 26500, Greece., Klapa MI; Metabolic Engineering and Systems Biology Laboratory, Institute of Chemical Engineering Sciences, Foundation for Research & Technology - Hellas (FORTH/ICE-HT), Patras, 26504, Greece. mklapa@iceht.forth.gr.; Department of Chemical and Biomolecular Engineering, University of Maryland, College Park, MD 20742, USA. mklapa@iceht.forth.gr.; Department of Bioengineering, University of Maryland, College Park, MD 20742, USA. mklapa@iceht.forth.gr. |
| Abstrakt: |
Untargeted metabolomics refers to the high-throughput analysis of the metabolic state of a biological system (e.g., tissue, biological fluid, cell culture) based on the concentration profile of all measurable free low molecular weight metabolites. Gas chromatography-mass spectrometry (GC-MS), being a highly sensitive and high-throughput analytical platform, has been proven a useful tool for untargeted studies of primary metabolism in a variety of applications. As an omic analysis, GC-MS metabolomics is a multistep procedure; thus, standardization of an untargeted GC-MS metabolomics protocol requires the integrated optimization of pre-analytical, analytical, and computational steps. The main difference of GC-MS metabolomics compared to other metabolomics analytical platforms, including liquid chromatography-MS, is the need for the derivatization of the metabolite extracts into volatile and thermally stable derivatives, the latter being quantified in the metabolic profiles. This analytical step requires special care in the optimization of the untargeted GC-MS metabolomics experimental protocol. Moreover, both the derivatization of the original sample and the compound fragmentation that takes place in GC-MS impose specialized GC-MS metabolomic data identification, quantification, normalization and filtering methods. In this chapter, we describe the integrated protocol of untargeted GC-MS metabolomics with both the analytical and computational steps, focusing on the GC-MS specific parts, and provide details on any sample depending differences. |
