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      Global metabolic profiling of animal and human tissues via UPLC-MS.

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          Abstract

          Obtaining comprehensive, untargeted metabolic profiles for complex solid samples, e.g., animal tissues, requires sample preparation and access to information-rich analytical methodologies such as mass spectrometry (MS). Here we describe a practical two-step process for tissue samples that is based on extraction into 'aqueous' and 'organic' phases for polar and nonpolar metabolites. Separation methods such as ultraperformance liquid chromatography (UPLC) in combination with MS are needed to obtain sufficient resolution to create diagnostic metabolic profiles and identify candidate biomarkers. We provide detailed protocols for sample preparation, chromatographic procedures, multivariate analysis and metabolite identification via tandem MS (MS/MS) techniques and high-resolution MS. By using these optimized approaches, analysis of a set of samples using a 96-well plate format would take ~48 h: 1 h for system setup, 8-10 h for sample preparation, 34 h for UPLC-MS analysis and 2-3 h for preliminary/exploratory data processing, representing a robust method for untargeted metabolic screening of tissue samples.

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          Author and article information

          Journal
          Nat Protoc
          Nature protocols
          Springer Science and Business Media LLC
          1750-2799
          1750-2799
          Jan 2013
          : 8
          : 1
          Affiliations
          [1 ] Biomolecular Medicine, Department of Surgery and Cancer, Faculty of Medicine, Imperial College, London, UK.
          Article
          nprot.2012.135
          10.1038/nprot.2012.135
          23222455
          402b5599-533c-4853-be31-ae69ba921bf5
          History

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