Separation and Purification of Ginsenosides and Flavonoids in From the Leaves and Stems of Panax quinquefolium by High‐Speed Countercurrent Chromatography and Online‐Storage Inner‐Recycling Countercurrent Chromatography

ABSTRACT Study aimed to isolate and purify compounds from the stems and leaves of Panax quinquefolius . By employing a highly innovative separation technique that combined multistage countercurrent chromatography (MRCC), high‐speed countercurrent chromatography (HSCCC), and an advanced online‐storage inner‐recycling countercurrent chromatography (OS‐IRCCC) mode for the first time, 12 compounds were successfully isolated, including 10 ginsenosides and 2 flavonoids. First, the crude extract was fractionated into five parts using D101 MRCC, with HPLC analysis revealing that 40% and 60% ethanol eluate contained the highest compound diversity. Overall, 40% ethanol eluate was separated using the solvent system of EtOAc/ n ‐BuOH/H 2 O (2:1:3, v/v), whereas 60% ethanol eluate underwent traditional countercurrent chromatography coupled with OS‐IRCCC separation using the solvent system of methyl tert‐butyl ether (MTBE)/ n ‐BuOH/ACN/H 2 O (4:2:3:8, v/v). Ultimately, various compounds were obtained, including kaempferol 3‐ O ‐ β ‐ d ‐glucopyranosyl‐(1 → 2)‐ β ‐ d ‐galactopyranosyl‐7‐ O ‐ α ‐ l ‐rhamnopyranoside (13.2 mg), ginsenoside Rc (7.4 mg), 20(R)‐ginsenoside Rh 1 (7.2 mg), ginsenoside Re (12.3 mg), kaempferol 3‐ O ‐ β ‐ d ‐glucopyranosyl‐(1 → 2)‐ β ‐ d ‐galactopyranoside (14.1 mg), ginsenoside Rb 1 (8.2 mg), ginsenoside Rb 2 (17.5 mg), ginsenoside Rb 3 (27.3 mg), ginsenoside Rg 1 (13.3 mg), ginsenoside Rg 2 (9.7 mg), ginsenoside Rd (11.4 mg), and pseudo‐ginsenoside F 11 (16.7 mg). This research highlights the efficacy of the novel separation technique in isolating and purifying valuable compounds from P. quinquefolius stems and leaves.