Fast and precise quantitative analysis of metabolic mixtures by 2D 1H INADEQUATE NMR.
Quantitative analysis of metabolic mixtures by (1)H 1D NMR offers a limited potential for precise quantification of biomarkers, due to strong overlap between the peaks. Two-dimensional spectroscopy is a powerful tool to unambiguously and simultaneously measure a larger number of metabolite contributions. However, it is still rarely used for quantification, first because quantitative analysis by 2D NMR requires a calibration procedure due to the multi-impulsional nature of 2D NMR experiments, and above all because of the prohibitive experiment duration that is necessary to obtain such a calibration curve. In this work, we develop and evaluate a 2D (1)H INADEQUATE protocol for a fast determination of metabolite concentrations in complex mixtures. The 2D pulse sequence is carefully optimized and evaluated in terms of precision and linearity. Quantitative (1)H INADEQUATE 2D spectra of metabolic mixtures are obtained in 7 min with a repeatability better than 2% for metabolite concentrations as small as 100 μM and an excellent linearity. The method described in this work allows a fast and precise quantification of metabolic mixtures, and it forms a promising tool for metabonomic studies.
Martineau
Estelle
E
Université de Nantes, CNRS, CEISAM, UMR 6230, B.P. 92208, 2 rue de la Houssinière, F-44322 Nantes Cedex 03, France.
Giraudeau
Patrick
P
Tea
Illa
I
Akoka
Serge
S
D015415:Biomarkers / Q000737:chemistry; D002021:Buffers; D002138:Calibration; D045744:Cell Line, Tumor; D002623:Chemistry Techniques, Analytical; D002626:Chemistry, Pharmaceutical / Q000379:methods*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D055432:Metabolomics / Q000379:methods; D015203:Reproducibility of Results; D013696:Temperature; D013997:Time Factors
D015415:Biomarkers; D002021:Buffers