Separation and Identification of Anthocyanins Extracted from Blueberry Wine Lees and Pigment Binding Properties toward β-Glucosidase.

Abstract

Anthocyanins were isolated from blueberry wine lees using Sephadex LH-20 column chromatography and semipreparative high-performance liquid chromatography (semipreparative HPLC) and then identified by HPLC-DAD-ESI-MS/MS. Our results show that malvidin-3-hexose (Mv-3-hex) and malvidin-3-(6'acetyl)-hexose (Mv-3-ace-hex) are the major components in the anthocyanin extracts of blueberry wine lees (>90%). The binding characteristics of Mv-3-hex and Mv-3-ace-hex with β-glucosidase were investigated by fluorescence spectroscopy, circular dichroism (CD) spectroscopy, and molecular docking. Spectroscopic analysis revealed that β-glucosidase fluorescence quenched by Mv-3-hex and Mv-3-ace-hex follows a static mode. Binding of Mv-3-hex and Mv-3-ace-hex to β-glucosidase mainly depends on electrostatic force. The result from CD spectra shows that adaptive structure rearrangement and increase of β-sheet structure occur only in the presence of Mv-3-ace-hex. A molecular docking study suggests that Mv-3-ace-hex has stronger binding with β-glucosidase than Mv-3-hex.