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Int. J. Mol. Sci. 2016, 17(7), 1017; doi:10.3390/ijms17071017

Stable Isotope-Assisted Evaluation of Different Extraction Solvents for Untargeted Metabolomics of Plants

1
Center for Analytical Chemistry, Department of Agrobiotechnology (IFA-Tulln), University of Natural Resources and Life Sciences, Vienna (BOKU), Konrad-Lorenz-Strasse 20, 3430 Tulln, Austria
2
Institute for Biotechnology in Plant Production, Department of Agrobiotechnology (IFA-Tulln), University of Natural Resources and Life Sciences, Vienna (BOKU), Konrad-Lorenz-Strasse 20, 3430 Tulln, Austria
3
Luxembourg Centre for Systems Biomedicine, University of Luxembourg Campus Belval, Avenue du Swing 6, 4367 Esch-Belval, Luxembourg
These authors contributed equally to this work.
*
Author to whom correspondence should be addressed.
Academic Editor: Ute Roessner
Received: 18 May 2016 / Revised: 13 June 2016 / Accepted: 21 June 2016 / Published: 28 June 2016
(This article belongs to the Special Issue Metabolomics in the Plant Sciences)
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Abstract

The evaluation of extraction protocols for untargeted metabolomics approaches is still difficult. We have applied a novel stable isotope-assisted workflow for untargeted LC-HRMS-based plant metabolomics , which allows for the first time every detected feature to be considered for method evaluation. The efficiency and complementarity of commonly used extraction solvents, namely 1 + 3 (v/v) mixtures of water and selected organic solvents (methanol, acetonitrile or methanol/acetonitrile 1 + 1 (v/v)), with and without the addition of 0.1% (v/v) formic acid were compared. Four different wheat organs were sampled, extracted and analysed by LC-HRMS. Data evaluation was performed with the in-house-developed MetExtract II software and R. With all tested solvents a total of 871 metabolites were extracted in ear, 785 in stem, 733 in leaf and 517 in root samples, respectively. Between 48% (stem) and 57% (ear) of the metabolites detected in a particular organ were found with all extraction mixtures, and 127 of 996 metabolites were consistently shared between all extraction agent/organ combinations. In aqueous methanol, acidification with formic acid led to pronounced pH dependency regarding the precision of metabolite abundance and the number of detectable metabolites, whereas extracts of acetonitrile-containing mixtures were less affected. Moreover, methanol and acetonitrile have been found to be complementary with respect to extraction efficiency. Interestingly, the beneficial properties of both solvents can be combined by the use of a water-methanol-acetonitrile mixture for global metabolite extraction instead of aqueous methanol or aqueous acetonitrile alone. View Full-Text
Keywords: 13C-labelling; plant metabolomics; sample preparation; Triticum aestivum; wheat; untargeted metabolomics 13C-labelling; plant metabolomics; sample preparation; Triticum aestivum; wheat; untargeted metabolomics
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Doppler, M.; Kluger, B.; Bueschl, C.; Schneider, C.; Krska, R.; Delcambre, S.; Hiller, K.; Lemmens, M.; Schuhmacher, R. Stable Isotope-Assisted Evaluation of Different Extraction Solvents for Untargeted Metabolomics of Plants. Int. J. Mol. Sci. 2016, 17, 1017.

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