Position-Specific Isotope Analysis of Xanthines: A 13C Nuclear Magnetic Resonance Method to Determine the 13C Intramolecular Composition at Natural Abundance

The natural xanthines caffeine, theobromine, and theophylline are of major commercial importance as flavor constituents in coffee, cocoa, tea, and a number of other beverages. However, their exploitation for authenticity, a requirement in these commodities that have a large origin-based price-range, by the standard method of isotope ratio monitoring by mass spectrometry (irm-MS) is limited. We have now developed a methodology that overcomes this deficit that exploits the power of isotopic quantitative 13C nuclear magnetic resonance (NMR) spectrometry combined with chemical modification of the xanthines to enable the determination of positional intramolecular 13C/12C ratios (δ13Ci) with high precision. However, only caffeine is amenable to analysis: theobromine and theophylline present substantial difficulties due to their poor solubility. However, their N-methylation to caffeine makes spectral acquisition feasible. The method is confirmed as robust, with good repeatability of the δ13Ci values in caffeine ...