title: Method for determining molar concentrations of metabolites in complex solutions from two-dimensional 1H-13C NMR spectra.
abstract: One-dimensional (1D) (1)H nuclear magnetic resonance (NMR) spectroscopy is used extensively for high-throughput analysis of metabolites in biological fluids and tissue extracts. Typically, such spectra are treated as multivariate statistical objects rather than as collections of quantifiable metabolites. We report here a two-dimensional (2D) (1)H-(13)C NMR strategy (fast metabolite quantification, FMQ, by NMR) for identifying and quantifying the approximately 40 most abundant metabolites in biological samples. To validate this technique, we prepared mixtures of synthetic compounds and extracts from Arabidopsis thaliana, Saccharomyces cerevisiae, and Medicago sativa. We show that accurate (technical error 2.7%) molar concentrations can be determined in 12 min using our quantitative 2D (1)H-(13)C NMR strategy. In contrast, traditional 1D (1)H NMR analysis resulted in 16.2% technical error under nearly ideal conditions. We propose FMQ by NMR as a practical alternative to 1D (1)H NMR for metabolomics studies in which 50-mg (extract dry weight) samples can be obtained.
authors.affiliation: Department of Biochemistry, University of Wisconsin-Madison, 433 Babcock Drive, Madison, Wisconsin 53706, USA.
authors.forename: Ian A
authors.initials: IA
authors.lastname: Lewis
authors.forename: Seth C
authors.initials: SC
authors.lastname: Schommer
authors.forename: Brendan
authors.initials: B
authors.lastname: Hodis
authors.forename: Kate A
authors.initials: KA
authors.lastname: Robb
authors.forename: Marco
authors.initials: M
authors.lastname: Tonelli
authors.forename: William M
authors.initials: WM
authors.lastname: Westler
authors.forename: Michael R
authors.initials: MR
authors.lastname: Sussman
authors.forename: John L
authors.initials: JL
authors.lastname: Markley
mesh_terms: D017360:Arabidopsis; D001826:Body Fluids / Q000737:chemistry*; D002247:Carbon Isotopes; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D000455:Medicago sativa; D008660:Metabolism; D008722:Methods; D015203:Reproducibility of Results; D012441:Saccharomyces cerevisiae; D012996:Solutions
chemical_list: D002247:Carbon Isotopes; D012996:Solutions