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Five phenylethanoid glycosides (PhGs), forsythoside B, verbascoside, alyssonoside, isoverbascoside, and leucosceptoside B, were isolated and purified from Lamiophlomis rotata (Benth.) Kudo by high-speed counter-current chromatography (HSCCC) combined with macroporous resin (MR) column separation. In the present study, the two-phase solvent system composed of ethyl acetate/n-butanol/water (13:3:10, v/v/v) was used for HSCCC separation. A total of 27 mg of forsythoside B, 41 mg of verbascoside, 29 mg of alyssonoside, 23 mg of isoverbascoside, and 13 mg of leucosceptoside B with purities of 97.7, 99.2, 99.5, 99.3, and 97.3%, respectively, were obtained in a one-step separation within 4 h from 150 mg of crude extract. The recoveries of the five PhGs after MR-HSCCC separation were 74.5, 76.5, 72.5, 76.4, and 77.0%, respectively. The chemical structures of all five compounds were identified by (1) H and (13) C NMR spectroscopy.