A novel microfluidic aqueous two-phase system with immobilized enzyme enhances cyanidin-3-O-glucoside content in red pigments from mulberry fruits

Abstract

Mulberry red pigments are safe, nontoxic and are one of the 48 natural pigments officially approved in China. Cyanidin-3-O-glucoside accounts for about 20 % of the total mulberry red pigments and has high physiological activity. Therefore, a novel microfluidic aqueous two-phase system with immobilized enzyme was constructed to enhance the cyanidin-3-O-glucoside content in red pigments under parallel-laminar flow condition for the first time. The effects of two-phase flow rates, temperature, pH and substrate concentration on cyanidin-3-O-rutinoside conversion and cyanidin-3-O-glucoside purity were investigated. The cyanidin-3-O-rutinoside conversion was 75.66 % within 8.59 s, and the purity of cyanidin-3-O-glucoside in the product was 88.78 % with an increase of 68 %. The immobilized enzyme could be reused seven times, maintaining a relative activity over 60 %. Compared with aqueous two-phase system, the overall reaction efficiency of mulberry red pigments in microfluidic aqueous two-phase system was higher. Cyanidin-3-O-rutinoside and cyanidin-3-O-glucoside were identified by HPLC-PDA-ESI-MS/MS. Overall, a new kind of high-quality red mulberry pigments with a cyanidin-3-O-glucoside purity close to 90 % was prepared using the innovative system quickly and effectively. This is of great significance for resource utilization.