Tannins are critically important molecules in viticulture and oenology because of the influential role that they play in determining the organoleptic properties of wine and their contribution to the health benefits associated with wine consumption. Despite the recognised importance of wine tannins, their complete chemical characterisation remains elusive, partly due to separation difficulties presented by their highly complex and diverse chemical structures. As a result, these compounds cannot be separated by conventional HPLC methods.

In this report, we demonstrate how on-line comprehensive hydrophilic interaction chromatography (HILIC) × reversed-phase liquid chromatography (RP-LC) coupled to fluorescence detection (FLD) and high resolution electrospray mass spectrometry (HR-ESI-MS) can be exploited as a powerful analytical method for detailed analysis of grape seed tannins. Combining HILIC and RP-LC resulted in greatly reduced co-elution, providing a well-structured elution pattern with separation on the basis of molecular size in the HILIC dimension and isomeric composition in the RP-LC dimension. HILIC×RP-LC-FLD-HR-ESI-MS delivered high resolution in a practical analysis time while providing selective and sensitive detection as well as greater confidence in compound identification. Experimental results allowed identification of a broad spectrum of proanthocyanidins, including procyanidins of a degree of polymerisation up to 16 and a degree of galloylation up to 8. The developed methodology represents the most powerful single analytical system for characterisation of grape seed tannins reported to date.