Use of Selective TOCSY NMR Experiments for Quantifying Minor Components in Complex Mixtures: Application to the Metabonomics of Amino Acids in Honey

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Abstract
The application of the traditional methods of multivariate statistics, such as the calculation of principle components, to the analysis of NMR spectra taken on sets of biofluid samples is one of the central approaches in the field of metabonomics. While this approach has proven to be a powerful and widely applicable technique, it has an inherent weakness, in that it tends to be dominated by those chemical species present at relatively higher concentrations. Using a set of commercial honey samples, a comparison of this classical metabonomics approach to one based on the use of the selective TOCSY experiment is presented. While the NMR spectrum of honey and its classical metabonomic analysis is completely dominated by a very few chemical species, specifically α-glucose and fructose, the statistical signal carried by minor honey components, such as amino acids, may be accessed using a selective TOCSY-based approach. This approach has the intrinsic virtue that it focuses the statistical analysis on a set of predefined chemical species, which might be chosen for their metabolic significance, and could be composed of either major or minor mixture constituents. Furthermore, the selective TOCSY method allows for more certain chemical identification, acquisition times of ∼1 min, and accurate quantification of the species contributing to the statistical discriminatory signal.