Comprehensive analysis of chestnut tannins by reversed phase and hydrophilic interaction chromatography coupled to ion mobility and high resolution mass spectrometry

Abstract

In this study, we report a methodology based on reversed phase LC (RP-LC) and hydrophilic interaction chromatography (HILIC) separations coupled to ion mobility (IM) and high resolution mass spectrometry (HR-MS) for the detailed analysis of hydrolysable tannins. The application of this approach to the analysis of an industrial chestnut (Castanea sativa, wood chips) tannin extract is demonstrated. A total of 38 molecular species, including a large number or isomers, were identified in this sample based on HR-MS(E) and UV absorption spectral information as well as retention behaviour in both separation modes. In total, 128 and 90 isomeric species were resolved by RP- and HILIC-LC-IM-TOF-MS, respectively. The combination of low- and high collision energy mass spectral data with complementary chromatographic separations allowed tentative and putative identification of twenty molecular species, comprising 78 isomers, in chestnut for the first time. Ion mobility resolved six new dimeric and trimeric vescalagin conformers with unique arrival (drift) times, including new conformers of roburin A-D which were not separated using either RP-LC or HILIC. HILIC was found to be the preferred separation mode for the analysis of vescalagin derivatives, while RP-LC is preferred for the analysis of ellagitannins with a cyclic glucose core. For the complete separation of the galloyl glucose species, comprehensive HILIC × RP-LC separation would be required.