Separation of Polyphenols and Caffeine from the Acetone Extract of Fermented Tea Leaves (Camellia sinensis) Using High-Performance Countercurrent Chromatography

Leaves from Camellia sienensis are a popular natural source of various beverage worldwide, and contain caffeine and polyphenols derived from catechin analogues. In the current study, caffeine (CAF, 1) and three tea polyphenols including (−)-epigallocatechin 3-O-gallate (EGCg, 2), (−)-gallocatechin 3-O-gallate (GCg, 3), and (−)-epicatechin 3-O-gallate (ECg, 4) were isolated and purified by flow-rate gradient high-performance countercurrent chromatography (HPCCC) using a two-phase solvent system composed of n-hexane–ethyl acetate–methanol–water (1:9:1:9, v/v). Two hundred milligrams of acetone-soluble extract from fermented C. sinensis leaves was separated by HPCCC to give 1 (25.4 mg), 2 (16.3 mg), 3 (11.1 mg) and 4 (4.4 mg) with purities over 98%. The structures of 1–4 were elucidated by QTOF-MS, as well as (1)H- and (13)C-NMR, and the obtained data were compared to the previously reported values.