One-step separation of nine structural analogues from Poria cocos (Schw.) Wolf. via tandem high-speed counter-current chromatography

• A tandem HSCCC separation system was first developed based on a six-port valve.
• Nine structural analogous including three isomers were isolated via tandem HSCCC.
• All separated compounds were separated by CCC from PC for the first time.
• Compound 9 with high retention ability was obtained by adopting extrusion elution.

A novel one-step separation strategy—tandem high-speed counter-current chromatography (HSCCC) was developed with a six-port valve serving as the switch interface. Nine structural analogues including three isomers were successfully isolated from Poria cocos (Schw.) Wolf. by one step. Compared with conventional HSCCC, peak resolution of target compounds was effectively improved in tandem one. Purities of isolated compounds were all over 90% as determined by HPLC. Their structures were then identified via UV, MS and 1H NMR, and eventually assigned as poricoic acid B (1), poricoic acid A (2), 3β,16α-dihydroxylanosta-7, 9(11), 24-trien-21-oic acid (3), dehydrotumulosic acid (4), polyporenic acid C (5), 3-epi-dehydrotumulosic acid (6), 3-o-acetyl-16α-hydroxydehydrotrametenolic acid (7), dehydropachymic acid (8) and dehydrotrametenolic acid (9) respectively. The results indicated that tandem HSCCC can effectively improve peak resolution of target compounds, and can be a good candidate for HSCCC separation of structural analogues.