title: Global metabolic profiling procedures for urine using UPLC-MS.
abstract: The production of 'global' metabolite profiles involves measuring low molecular-weight metabolites (<1 kDa) in complex biofluids/tissues to study perturbations in response to physiological challenges, toxic insults or disease processes. Information-rich analytical platforms, such as mass spectrometry (MS), are needed. Here we describe the application of ultra-performance liquid chromatography-MS (UPLC-MS) to urinary metabolite profiling, including sample preparation, stability/storage and the selection of chromatographic conditions that balance metabolome coverage, chromatographic resolution and throughput. We discuss quality control and metabolite identification, as well as provide details of multivariate data analysis approaches for analyzing such MS data. Using this protocol, the analysis of a sample set in 96-well plate format, would take ca. 30 h, including 1 h for system setup, 1-2 h for sample preparation, 24 h for UPLC-MS analysis and 1-2 h for initial data processing. The use of UPLC-MS for metabolic profiling in this way is not faster than the conventional HPLC-based methods but, because of improved chromatographic performance, provides superior metabolome coverage.
authors.affiliation: Biomolecular Medicine, Department of Surgery and Cancer, Faculty of Medicine, Imperial College London, London, UK. e.want@imperial.ac.uk
authors.forename: Elizabeth J
authors.initials: EJ
authors.lastname: Want
authors.forename: Ian D
authors.initials: ID
authors.lastname: Wilson
authors.forename: Helen
authors.initials: H
authors.lastname: Gika
authors.forename: Georgios
authors.initials: G
authors.lastname: Theodoridis
authors.forename: Robert S
authors.initials: RS
authors.lastname: Plumb
authors.forename: John
authors.initials: J
authors.lastname: Shockcor
authors.forename: Elaine
authors.initials: E
authors.lastname: Holmes
authors.forename: Jeremy K
authors.initials: JK
authors.lastname: Nicholson
mesh_terms: D015415:Biomarkers / Q000652:urine; D002853:Chromatography, Liquid / Q000379:methods*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D055442:Metabolome; D055432:Metabolomics / Q000379:methods*; D015394:Molecular Structure; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D014556:Urine / Q000737:chemistry*
chemical_list: D015415:Biomarkers
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