Abstract
Metabolomics is a growing research field where new protocols are rapidly developed and new applications discovered. Common applications include biomarker discovery and elucidation of drug metabolism. However, the development of such protocols rarely includes a systematic optimization followed by validation with real samples. Here a GC/MS-based protocol using methoximation followed by silylation with N-tert-butyldimethylsilyl-N-methyltrifluoroacetamide (MTBSTFA) for analysis of blood plasma metabolites is thoroughly developed and optimized from derivatization to detection with statistical design of experiments (DOE). Validation was performed with blood plasma samples and proved the methodology to be efficient, rapid and reliable with a total of 51 analyses performed in 24 h, with linear responses, low detection limits and good precision. The obtained chromatograms were much cleaner, due to the absence of glucose overloading, and the data was found to drift less with MTBSTFA derivatisation than with MTBSTFA derivatisation.
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Acknowledgments
This work was supported by grants from Swedish Research Council (14196-06-3), the Crafoord Foundation, Lars Hierta, Fredrik and Ingrid Thuring, Åke Wiberg, Albert Påhlsson, O.E. and Edla Johansson Foundations, Knut and Alice Wallenberg Foundation, and the Royal Physiographic Society. Support from Inga and John Hain Foundation to PS is acknowledged.
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Danielsson, A.P.H., Moritz, T., Mulder, H. et al. Development of a gas chromatography/mass spectrometry based metabolomics protocol by means of statistical experimental design. Metabolomics 8, 50–63 (2012). https://doi.org/10.1007/s11306-011-0283-6
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DOI: https://doi.org/10.1007/s11306-011-0283-6