Abstract
Metabolome sample preparation is one of the key factors in metabolomics analyses. The quality of the metabolome data will depend on the suitability of the experimental procedures to the cellular system (e.g., yeast cells) and the analytical performance. Here, we summarize a protocol for metabolome analysis of yeast cells using gas chromatography–mass spectrometry (GC–MS). First, the main phases of a metabolomics analysis are identified: sample preparation, metabolite extraction, and analysis. We also provide an overview on different methods used to quench samples and extract intracellular metabolites from yeast cells. This protocol provides a detailed description of a GC-MS-based analysis of yeast metabolome, in particular for metabolites containing amino and/or carboxyl groups, which represent most of the compounds participating in the central carbon metabolism.
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References
Canelas AB, Ras C, Pierick A et al (2008) Leakage-free rapid quenching technique for yeast metabolomics. Metabolomics 4:226–239
De Koning W, Van Dam K (1992) A method for the determination of changes of glycolytic metabolites in yeast on a subsecond time scale using extraction at neutral pH. Anal Biochem 204:118–123
Gonzalez B, François J, Renaud M (1997) A rapid and reliable method for metabolite extraction in yeast using boiling buffered ethanol. Yeast (Chichester, England) 13: 1347–1355
Taymaz-Nikerel H, de Mey M, Ras C et al (2009) Development and application of a differential method for reliable metabolome analysis in Escherichia coli. Anal Biochem 386: 9–19
Wittmann C, Krömer JO, Kiefer P et al (2004) Impact of the cold shock phenomenon on quantification of intracellular metabolites in bacteria. Anal Biochem 327:135–9
Castrillo JI, Hayes A, Mohammed S et al (2003) An optimized protocol for metabolome analysis in yeast using direct infusion electrospray mass spectrometry. Phytochemistry 62: 929–937
Faijes M, Mars AE, Smid EJ (2007) Comparison of quenching and extraction methodologies for metabolome analysis of Lactobacillus plantarum. Microb Cell Fact 6:27
Boer VM, Crutchfield CA, Bradley PH et al (2010) Growth-limiting intracellular metabolites in yeast growing under diverse nutrient limitations. Mol Biol Cell 21:198–211
Ewald JC, Heux S, Zamboni N (2009) High-throughput quantitative metabolomics: workflow for cultivation, quenching, and analysis of yeast in a multiwell format. Anal Chem 81: 3623–9
Sasidharan K, Soga T, Tomita M et al (2012) A yeast metabolite extraction protocol optimised for time-series analyses. PLoS One 7:e44283
Smart KF, Aggio RBM, Van Houtte JR et al (2010) Analytical platform for metabolome analysis of microbial cells using methyl chloroformate derivatization followed by gas chromatography-mass spectrometry. Nat Protoc 5: 1709–29
Spura J, Reimer LC, Wieloch P et al (2009) A method for enzyme quenching in microbial metabolome analysis successfully applied to gram-positive and gram-negative bacteria and yeast. Anal Biochem 394:192–201
Weuster-Botz D (1997) Sampling tube device for monitoring intracellular metabolite dynamics. Anal Biochem 246:225–33
Buziol S, Bashir I, Baumeister A et al (2002) New bioreactor-coupled rapid stopped-flow sampling technique for measurements of metabolite dynamics on a subsecond time scale. Biotechnol Bioeng 80:632–6
Paczia N, Nilgen A, Lehmann T et al (2012) Extensive exometabolome analysis reveals extended overflow metabolism in various microorganisms. Microb Cell Fact 11:122
Mashego MR, Wu L, Van Dam JC et al (2004) MIRACLE: mass isotopomer ratio analysis of U-13C-labeled extracts. A new method for accurate quantification of changes in concentrations of intracellular metabolites. Biotechnol Bioeng 85:620–8
Canelas AB, ten Pierick A, Ras C et al (2009) Quantitative evaluation of intracellular metabolite extraction techniques for yeast metabolomics. Anal Chem 81:7379–89
Bolten CJ, Wittmann C (2008) Appropriate sampling for intracellular amino acid analysis in five phylogenetically different yeasts. Biotechnol Lett 30:1993–2000
Brown SD (1988) Chemometrics: A textbook. D. L. Massart. B. G. M. Vandeginste, S. N. Deming, Y. Michotte, and L. Kaufman, Elsevier, Amsterdam, 1988. ISBN 0-444-42660-4. Price Dfl 175.00. J Chemometr 2: 298–299
Putri SP, Yamamoto S, Tsugawa H et al (2013) Current metabolomics: technological advances. J Biosci Bioeng 116:9–16
Van Der Greef J, Smilde AK (2005) Symbiosis of chemometrics and metabolomics: past, present, and future. J Chemometr 19:376–386
Koek MM, Jellema RH, van der Greef J et al (2011) Quantitative metabolomics based on gas chromatography mass spectrometry: status and perspectives. Metabolomics 7:307–328
Villas-Bôas SG, Højer-Pedersen J, Akesson M et al (2005) Global metabolite analysis of yeast: evaluation of sample preparation methods. Yeast (Chichester, England) 22:1155–69
Winder CL, Dunn WB (2011) Fit-for-purpose quenching and extraction protocols for metabolic profiling of yeast using chromatography-mass spectrometry platforms. Methods Mol Biol (Clifton, NJ) 759:225–38
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Carneiro, S., Pereira, R., Rocha, I. (2014). Yeast Metabolomics: Sample Preparation for a GC/MS-Based Analysis. In: Mapelli, V. (eds) Yeast Metabolic Engineering. Methods in Molecular Biology, vol 1152. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0563-8_12
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DOI: https://doi.org/10.1007/978-1-4939-0563-8_12
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