title: High-speed counter-current chromatography coupled online to high performance liquid chromatography-diode array detector-mass spectrometry for purification, analysis and identification of target compounds from natural products.
abstract: A challenge in coupling high-speed counter-current chromatography (HSCCC) online with high performance liquid chromatography (HPLC) for purity analysis was their time incompatibility. Consequently, HSCCC-HPLC was conducted by either controlling HPLC analysis time and HSCCC flow rate or using stop-and-go scheme. For natural products containing compounds with a wide range of polarities, the former would optimize experimental conditions, while the latter required more time. Here, a novel HSCCC-HPLC-diode array detector-mass spectrometry (HSCCC-HPLC-DAD-MS) was developed for undisrupted purification, analysis and identification of multi-compounds from natural products. Two six-port injection valves and a six-port switching valve were used as interface for collecting key HSCCC effluents alternatively for HPLC-DAD-MS analysis and identification. The ethyl acetate extract of Malus doumeri was performed on the hyphenated system to verify its efficacy. Five main flavonoids, 3-hydroxyphloridzin (1), phloridzin (2), 4',6'-dihydroxyhydrochalcone-2'-O-β-D-glucopyranoside (3, first found in M. doumeri), phloretin (4), and chrysin (5), were purified with purities over 99% by extrusion elution and/or stepwise elution mode in two-step HSCCC, and 25mM ammonium acetate solution was selected instead of water to depress emulsification in the first HSCCC. The online system shortened manipulation time largely compared with off-line analysis procedure and stop-and-go scheme. The results indicated that the present method could serve as a simple, rapid and effective way to achieve target compounds with high purity from natural products.
authors: Research Institute of Chinese Medicine, Hunan Academy of Chinese Medicine, Changsha 410013, China.
authors: Xuejuan
authors: X
authors: Liang
authors: College of Chemistry and Chemical Engineering, Central South University, Changsha 410083, PR China.
authors: Yuping
authors: Y
authors: Zhang
authors: Research Institute of Chinese Medicine, Hunan Academy of Chinese Medicine, Changsha 410013, China.
authors: Wei
authors: W
authors: Chen
authors: Research Institute of Chinese Medicine, Hunan Academy of Chinese Medicine, Changsha 410013, China.
authors: Ping
authors: P
authors: Cai
authors: Research Institute of Chinese Medicine, Hunan Academy of Chinese Medicine, Changsha 410013, China. Electronic address: zhangshuihan0220@126.com.
authors: Shuihan
authors: S
authors: Zhang
authors: Research Institute of Chinese Medicine, Hunan Academy of Chinese Medicine, Changsha 410013, China.
authors: Xiaoqin
authors: X
authors: Chen
authors: College of Chemistry and Chemical Engineering, Central South University, Changsha 410083, PR China. Electronic address: shuyshi@gmail.com.
authors: Shuyun
authors: S
authors: Shi
mesh_terms: D001688:Biological Products / Q000737:chemistry* / Q000302:isolation & purification; D002623:Chemistry Techniques, Analytical / Q000295:instrumentation / Q000379:methods*; D002851:Chromatography, High Pressure Liquid; D003377:Countercurrent Distribution; D005419:Flavonoids / Q000032:analysis / Q000302:isolation & purification; D027845:Malus / Q000737:chemistry; D013058:Mass Spectrometry; D010936:Plant Extracts / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification
chemical_list: D001688:Biological Products; D005419:Flavonoids; D010936:Plant Extracts; C043561:chrysin
keywords: Flavonoid; HSCCC–HPLC–DAD–MS; Malus doumeri; On-line purity monitor