The applicability of high-speed counter-current chromatography for preparative separation of biosynthesis products: Glycosylation products as example

Biosynthesis is a promising way to manufacture desired products, however, the purification of its final products is a tough work due to the huge amount of reaction matrix. Liquid stationary phase of high-speed counter-current chromatography could easily avoid the commonly disadvantages that occurred in traditional column chromatography in the field of biosynthesized products purification. This characteristic makes high-speed counter-current chromatography particularly applicable for final products separation in biosynthesis. In this study, the glycosylation products of Silybin B by one-pot glycosylation were successfully purified by high-speed counter-current chromatography to show the applicability of high-speed counter-current chromatography for preparative separation of biosynthesis products. An optimized n-hexane/ethyl acetate/methanol/water (2:5:2:3, v/v/v/v) system was applied in this study. As a result, four Silybin B glycosylation products, including 7 mg of Silybin B-5-O-beta-D-glucoside (SG-1), 12 mg of Silybin B-3-O-beta-D-glucoside (SG-2), 10 mg of Silybin B-7-O-beta-D-glucoside (SG-3), and 24 mg of Silybin B-20-O-beta-D-glucoside (SG-4), were simultaneously separated from 200 mg of glycosylation crude products, with the purity of 89.3, 95.2, 96.4, and 97.5%, respectively. Their structures were identified by spectroscopic analysis.