A method of microwave‐assisted extraction coupled with high‐speed counter‐current chromatography was established for separation and purification of isopimpinellin, pimpinellin and phellopterin from Toddalia asiatica (L.) Lam. The conditions of MAE including the extraction solvent, size of sample, solid/liquid ratio, extraction temperature and extraction time were optimized by a mono‐factor test. That is, 2.0 g dried powder of T. asiatica (L.) Lam of 0.30‐0.15 mm size was extracted with 20 mL (solid/liquid ratio of 1:10, g/mL) methanol under 50 °C for 1 min. The crude extract was separated and purified by high‐speed counter‐current chromatography with hexane–ethyl acetate–methanol–water (5:5:5.5:4.5, v/v/v/v) solvent system. 0.85 mg/g of isopimpinellin, 2.55 mg/g of pimpinellin and 0.95 mg/g of phellopterin were obtained from original sample in one‐step within 240 min, the purity determined by high performance liquid chromatography was 95.0%, 99.1% and 96.4%, respectively. Their chemical structures were further identified by mass spectroscopy and nuclear magnetic resonance spectroscopy. The results demonstrated that microwave‐assisted extraction coupled with high‐speed counter‐current chromatography was a feasible, economical and efficient technique for rapid extraction, separation and purification of effective compounds from natural products.