Combined HILIC and RP HPLC methods for the purification of glycosidic aroma precursors in grape juices

Wine aroma, a very important sensory parameter, is composed by a wide variety of compounds with different aromatic properties. A significant part of important flavour compounds is accumulated in grapes as non-volatile and flavourless glycoconjugates, which are known as glycosidic aroma precursors. The aglycone part of glycosides is often represented by monoterpenes, C13-norisoprenoids, benzene derivatives and long-chain aliphatic alcohols. The sugar moiety includes glucose (O-β-D-glucosides) or disaccharides (O-diglycosides). Flavorless glycoconjugates represent a high aroma potential as they can be hydrolyzed to release volatile flavorants which are involved in wine flavor. The component analysis of glycoconjugates required their preliminary esterification or enzymatic hydrolysis, to liberate the aglycones, and the subsequent identification by GC-MS. Glycosidic aroma precursors were fractionated by using countercurrent chromatography (MLCCC) but pure compounds could be obtained by reversed phase (RP) HPLC with C18 column only after per-acetylation of glycosides. Hydrophilic interaction chromatography (HILIC) coupled to RP-HPLC provides a particularly powerful method for the separation of polar compounds. We present here our preliminary results of this approach applied to the isolation of pure aroma precursors from juice of two grape varieties. The glycosides were studied by 1D- and 2D-NMR spectroscopy which led to the identification of geranyl 6-O-b-L-arabinofuranosyl-b-D-glucopyranoside, glucosyl (E)-2,6-dimethyl-6-hydroxyocta-2,7-dienoate, glycosides of linalool oxides and others.