Efficient acquisition of high-purity cyanidin-3-O-glucoside from mulberry fruits: An integrated process of ATPS whole-cell transformation and semi-preparative HPLC purification

Abstract

As a nutritious fruit, mulberry is an ideal source of high-quality cyanidin-3-O-glucoside (C₃G) with various biological activities. However, the difficult separation process of high-purity C₃G leads to its high price. To rapidly prepare high-purity C₃G, cyanidin-3-O-rutinoside is converted to C₃G by direct hydrolysis of rhamnose bond using a whole-cell catalyst containing α-rhamnosidase. Combined with an aqueous two-phase system, a coupling reaction separation system was established. Two monomers were successfully separated by semi-preparative high performance liquid chromatography (semi-preparative HPLC). The conversion of C₃G catalyzed by whole-cells in the PEG/Na₂SO₄ system increased from 47.11 % to 66.56 %, compared with the EtOH/(NH₄)₂SO₄ system, and the whole-cell activity remained above 50 % after five rounds of reuse. Meanwhile, the purity of C₃G was increased to 99 % via the semi-preparative HPLC purification and identified by MS. Thus, an integrated process of whole-cell-catalyzed conversion and product peak cutting partition collection provides a novel strategy for efficient biomanufacturing of high-purity C₃G.

Keywords: Aqueous two-phase system; Cyanidin-3-O-glucoside; Cyanidin-3-O-rutinoside; Mulberry red pigment; Semi-preparative HPLC; Whole cell.