Abstract
Analysis of the entire metabolome as the sum of all detectable components in the sample rather than analysis of each individual metabolite is performed by the metabolomics approaches. To monitor in parallel hundreds or even thousands of metabolites, high-throughput techniques are required that enable screening for relative changes rather than absolute concentrations of compounds. Most analytical techniques for profiling small molecules consist of gas chromatography (GC) or high-performance liquid chromatography (HPLC) coupled to mass spectrometry. HPLC separations are better suited for the analysis of labile and high molecular weight compounds, and for the analysis of nonvolatile polar compounds in their natural form. Although GC- and HPLC-based profiling techniques are not truly quantitative, the compounds detecting and employing the acceptable standards may compare their relative amounts. We have demonstrated that reversed-phase monolithic capillary chromatography and hydrophilic chromatography can be successfully applied for sufficient plant crude extracts separations and metabolomics studies.
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References
Tolstikov, V. V., Lommen, A., Nakanishi, K., Tanaka, N., and Fiehn O. (2003) Monolithic silica-based capillary reversed phase liquid chromatography/electrospray mass spectrometry for plant metabolomics. Anal. Chem. 75, 6737–6740.
Tolstikov, V. V., Fiehn, O., and Shulaev, V. (2005) The influence of the extraction methodology on global LC-MS and GC-MS metabolite profiling of arabidopsis thaliana leaf tissues. 53rd ASMS Conference, San Antonio, TX, June 5–9, 2005.
Tolstikov, V. V., Lorence, A., Cortes, D. F., et al. (2005) Use of capillary monolithic columns for untargeted metabolite profiling of the mutant lines overexpressing Miox4, the gene involved in L-ascorbic acid biosynthesis in arabidopsis. 28th International Symposium On Capillary Chromatography and Electrophoresis. Las Vegas, NV, May 22–25, 2005.
Tolstikov, V. V. and Fiehn, O. (2002) Analysis of highly polar compounds of plant origin: combination of hydrophilic interaction chromatography and electrospray ion trap mass spectrometry. Anal. Biochem. 301, 298–307.
Tolstikov, V. V., Tanaka, N., and Fiehn O. (2003) Comprehensive metabolome analysis of crude Arabidopsis thaliana leaf extracts by LC/ESI-MSn/UV coupling. 2nd Plant Metabolomics Conference, Potsdam, Germany, April 25–28, 2003.
Tolstikov, V. V., Tanaka, N., and Fiehn, O. (2003) Metabolomics: LC-MS analysis development. Joint BTS/Cereal Chemistry Symposium, Adelaide, Australia, September 8–10, 2003.
Tolstikov, V. V., Zhang, B., Weckwerth, W., and Fiehn O. (2001) Structural investigation of O-glycans derived from plant material by the use of the HILIC HPLC separation and ESI-mass spectrometry. 49th ASMS conference on Mass Spectrometry and Allied Topics, Chicago, IL, May 27–31, 2001.
Tolstikov, V. V., Costisella, B., Weckwerth W., Zhang B., and Fiehn O. (2002) Accurate QTOF and MSn Ion trap measurements require additional NMR data for plant metabolites de-novo identification. 50th ASMS Conference on Mass Spectrometry and Allied Topics, Orlando, FL, June 2–7, 2002.
Tolstikov, V. V., Tanaka, N., and Fiehn, O. (2003) LC/MS analysis and development for plant metabolomic studies. 2003 LCMS Montreux Symposium. Savannah, GA, October 15–17, 2003.
Tanaka, N., Tolstikov, V., Weckwerth, W., Fiehn, O., and Fukusaki, H., (2003) Micro HPLC for metabolomics. In: Frontier of Metabolomic Research, Springer-Verlag, Tokyo, Japan, pp. 85–100.
Ikegami, T., Kobayashi, H., Kimura, H., Tolstikov, V., Fiehn, O., and Tanaka, N. (2005) HPLC for metabolomics: high efficiency separations utilizing monolithic silica columns. In: Metabolomics. The Frontiers of Systems Biology, Springer Verlag, Tokyo, Japan, pp. 107–126.
Nikiforova, V., Kopka, J., Tolstikov, V., Fiehn, O., Hesse, H., and Hoefgen, R. (2005) Systems rebalancing of metabolism in response to sulfur deprivation, as revealed by metabolome analysis of arabidopsis plants. Plant Physiology 138, 304–318.
Kobayashi, H., Kajiwara, W., Inui, Y., et al. (2004) Chromatographic properties of monolithic silica capillary columns for polar and nonpolar compounds in reversed-phase HPLC. Chromatographia 60, S19–S25.
Ikegami, T., Dicks, E., Kobayashi, H., et al. (2004) How to utilize true performance of monolithic silica columns? J. Sep. Sci. 27, 1292–1302.
Richardson, P. T. and Baker, D. A. (1982) The chemical composition of cucurbit vascular exudates. J. Exp. Bot. 33, 1239–1247.
Snyder, L. R., Glajch, J. L., and Kirkland, J. J. (1988) Non-ionic samples: reversed-and normal-phase HPLC. In: Practical HPLC Method Development. Wiley, New York, NY, pp. 233–291.
Snyder, L. R., Stadalius, M., and Quarry, M. A. (1983) Gradient elution in reversed-phase HPLC separation of macromolecules. Anal. Chem. 55, 1412A–1430A.
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Tolstikov, V.V., Fiehn, O., Tanaka, N. (2007). Application of Liquid Chromatography-Mass Spectrometry Analysis in Metabolomics. In: Weckwerth, W. (eds) Metabolomics. Methods in Molecular Biology™, vol 358. Humana Press. https://doi.org/10.1007/978-1-59745-244-1_9
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DOI: https://doi.org/10.1007/978-1-59745-244-1_9
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