Analytical Method Development Projects

Summary of Projects

 

From 'omics discovery to targeted metabolite analysis: an optimal workflow

The overall aim of this new project, jointly funded by NERC and Thermo Fisher Scientific, is to develop novel strategies for efficiently developing targeted, high-throughput and quantitative LC-MS assays for measuring molecular biomarkers of environmental pollution. We will emphasise the discovery of robust biomarkers in the sentinel organism Daphnia magna.

Direct infusion FT-ICR mass spectrometry based metabolomics

We are developing analytical and bioinformatic methods for our Thermo FT-ICR mass spectrometer and Triversa chip-based nanoelectrospray assembly to maximise the number of metabolites detected, the sample throughput and the analytical reproducibility, while maintaining high mass accuracy and resolution.

  • R. J. M. Weber, A. D. Southam, U. Sommer, M. R. Viant, Characterization of isotopic abundance measurements in high resolution FT-ICR and Orbitrap mass spectra for improved confidence of metabolite identification. Anal. Chem. 83, 3737-3743 (2011).
  • A. D. Southam, T. G. Payne, H. J. Cooper, T. N. Arvanitis, M. R. Viant, Spectral Stitching Method Increases the Dynamic Range and Mass Accuracy of Wide-scan Direct Infusion nano-Electrospray Fourier Transform Ion Cyclotron Resonance Mass Spectrometry-based Metabolomics. Anal. Chem. 79, 4595-4602 (2007).
Sample preparation for metabolomics

The measurement of tissue-specific metabolic fingerprints can be of particular interest when investigating disease processes, mechanisms of toxicity, or when knowledge of the metabolic interactions between different organs is required. The preparation of tissue extracts for metabolomics analysis involves three critical steps. The first is to rapidly collect and freeze the tissue to ‘quench’ metabolism and preserve the metabolite concentrations. The second step involves mechanically disrupting the tissue to allow the extraction of the low molecular weight metabolites. The goals are to de-proteinise the sample to permanently halt metabolism, and to extract only those metabolites of interest (e.g., lipids, carbohydrates, amino acids and other small metabolites). The final step is to optimise the solution for high resolution NMR spectroscopy or mass spectroscopy. For NMR analyses, this entails re-suspending the metabolite extract in an appropriate deuterated solvent buffer and adding an NMR internal standard. We are engaged in optimising these methods for a variety of tissues from a variety of organisms.

  • H. Wu, A. D. Southam, A. Hines and M. R. Viant, High throughput tissue extraction protocol for NMR- and MS-based metabolomics. Anal. Biochem. 372, 204-212 (2008).

  • C.-Y. Lin, H. Wu, R. S. Tjeerdema and M. R. Viant, Evaluation of metabolite extraction strategies from tissue samples using NMR metabolomics. Metabolomics 3, 55-67 (2007).