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UPLC-TOF-MS for high resolution plant metabolite profiling and metabolomics
Among the different techniques enlisted for metabolome analysis, ultra-performance liquid chromatography coupled to time-of-flight mass spectrometry (UPLC-TOF-MS) represents a powerful platform for rapid and sensitive metabolite fingerprinting which provides very reproducible datasets in both chromatographic and mass dimensions. This technique is well established for bio fluid analysis  but has only scarcely be used for plant metabolomics. In our search for new bioactive stress-induced plant constituents, a generic UPLC-TOF-MS approach has been devised  and consists of (i) high throughput metabolite fingerprinting involving rapid gradients on numerous control and stressed plants (ii) high resolution metabolite profiling of selected pool samples on high peak capacity UPLC columns after efficient gradient transfer. Multivariate analysis applied to the fingerprinting data enables group discrimination and evidences ions (m/z) responsible for the main differences. High resolution profiling with long gradients provides a precise localisation and deconvolution of the putative biomarkers. Applied to the study of the wound response in Arabidopsis thaliana (Brassicaceae), this sequential strategy has been used to detect new key minor biomarkers and enabled the discrimination of different stress states in relation with global metabolome variations. Microgram amount of the biomarkers of interest was obtained by LC-MS triggered microfractionation. Their full structure determination was ensured by 1D and 2D CapNMR experiments . Original derivatives of the plant hormone jasmonic acid  were thus identified and a global picture of their temporal and spatial dynamics was obtained. The defence gene expression potential of these oxylipins was evaluated based on DNA microarray experiments.
Acknowledgements: Swiss National Science Foundation (Grants no. 205320–107735/1 to J-L.W. and S.R.).
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