Human Urinary Metabolite Quantification in Nuclear Magnetic Resonance-based Metabonomics Using Electronic Internal Reference

We report a complete assessment for urinary quantitative analysis by using an electronic signal as an internal reference, which was generated via decoupling channel on a Varian NMR spectrometer with its amplitude, frequency and transverse relaxation time digitally programmable according to a decoupling shape file. The electronic signal with excellent stability (standard deviation 0.4%) avoided possible problems such as insolubility, interaction and spectral overlap for a traditional internal reference. Concentration determination using the electronic signal proved to be reliable by a series of quantitative experiments. The relative error of quantification was about 1% and 5% when the analyte concentration was above 20 mmol/L and below 2 mmol/L respectively. We further measured the concentration of human urinary metabolites in a simultaneous fashion using this electronic internal reference and obtained consistent results with the values measured by the biochemical analysis instrument used in hospitals.