High-Pressure Liquid Chromatographic Separation of Anthocyanins of Vitis Vinifera

Abstract

High-pressure liquid chromatography (HPLC) was applied to separation and quantitation of the anthocyanin pigments of two varieties of Vitis vinifera. Reversed-phase columns and acidified water-methanol eluants were used to separate the pigments of Cabernet Sauvignon grapes into twenty discrete peaks. Easily resolved from one another were five anthocyanidin-3-monoglucosides, five anthocyanidin-3-monoglucoside-acetates, five anthocyanidin-3-monoglucoside-p-coumarates, and malvidin-3-monoglucoside-caffeoate. Also separated were three unknown pigments. Monoglucoside pigments accounted for 65% of the total pigment. The monoglucoside-acetates and monoglucoside-p-coumarates respectively represented 26.9% and 8% of the total pigment. The malvidin-derived anthocyanins were 70% of the total, and anthocyanins containing the aglycone cyanidin represented only 1.5%. Evidence was obtained that the acylated anthocyanins are preferentially formed from malvidin-3-monoglucoside rather than from delphinidin-3-monoglucoside. The combination of anthocyanin degradation procedures and HPLC analysis allowed rapid identification of the anthocyanin pigments. HPLC separation of pigment degradation products required less than 15 minutes. One hundred micrograms of pigment was sufficient for identification by degradaton and HPLC analysis.