Improved high performance liquid chromatographic separation of anthocyanin compounds from grapes using a novel mixed-mode ion-exchange reversed-phase column

A novel mixed mode HPLC method using a column combining both ion-exchange and reversed-phase separation mechanisms has been developed to facilitate analysis of anthocyanins in grapes. Chromatographic performance and subsequent analysis of anthocyanidin diglucosides and acylated compounds are significantly improved using the new column, compared to those associated with conventional C18 reversed-phase methods. The mixed mode column produces a distinctive eluting pattern for the different anthocyanin subgroups, avoiding overlaps found with C18 columns. The enhanced chromatographic resolution provides nearly complete separation of 37 anthocyanin types, and permits detection of delphinidin 3-O-(6″-O-caffeoyl) β-d-glucoside for the first time in extracts of skins from Concord grapes.