Absolute quantification of metabolites in tomato fruit extracts by fast 2D NMR
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Quantitative NMR metabolomics is a powerful tool to have access to valuable information on metabolism. Unfortunately, the quantitative analysis of metabolic samples is often hampered by peak overlap. Two-dimensional (2D) spectroscopy offers a promising alternative and quantitative results can be obtained provided that a calibration approach is employed. However, the duration of 2D NMR experiments is barely compatible with the metabolomic study of a large number of samples. This drawback can be circumvented by relying on “ultrafast” experiments capable of recording 2D spectra in a single-scan. While such experiments are not sensitive enough to match the concentrations of metabolic samples, a compromise can be reached by hybrid strategies capable of recording 2D NMR spectra of extracts in a few minutes only. The purpose of this study is to demonstrate that these multi-scan single-shot experiments can be successfully applied to the absolute quantification of major metabolites in plant extracts. Fast COSY experiments are recorded in 5 min on tomato fruit pericarp extracts at different stages of development. The concentration of eight major metabolites is determined with a trueness better than 10 % and a technical repeatability of a few percent. The experiments performed at two magnetic fields lead to similar quantitative results, in coherence with the metabolism of tomato fruit. The results show that fast 2D NMR methods form a promising tool for fast targeted metabolomics, and open promising perspective towards the automated and high-throughput quantitative analysis of large groups of plant and other samples for metabolomics and for the modelling of metabolism.
KeywordsNMR Quantitative analysis Ultrafast 2D NMR COSY Tomato Extracts
The authors are grateful to Dr Estelle Martineau, Prof Serge Akoka and Prof Gérald Remaud for fruitful discussions, and acknowledge Michel Giraudeau for linguistic assistance. They thank Pierre Gaillard and Jacques Longuesserre from INVENIO for help in organizing the greenhouse experiment and Emilie Labadie for technical help in the greenhouse. They also thank Dr Camille Bénard, Dr Benoit Biais, Patricia Ballias and other colleagues of the Fruit Biology and Pathology Research Unit (UMR1332) for their crucial contribution during culture follow up, harvests and sample preparation, and Daniel Jacob for help with spectra and metadata uploading into MetaboLights. This research was supported by a young investigator starting grant from the “Agence Nationale de la Recherche” (ANR grant 2010-JCJC-0804-01), the European project Eranet EraSysBio + “FRuit Integrative Modelling”, the CORSAIRE metabolomics facility, the Bordeaux Metabolome Facility and MetaboHUB (ANR-11-INBS-0010 project).
Conflict of interest
The authors declare that there are no conflicts of interest.
Compliance with ethical requirements
This article does not contain any studies with human or animal subjects.
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