Study On The Secoiridoid Components And The Extract Processing Of Radix Gentianae

This paper separated partial Gentiana scabra Bunge components and identified their structures on traditional methods combining with modern plant chemical extracting and separating techniques.The extracting process of Gentiana scabra Bunge was established.Then, we established the Gentiana scabra Bunge extracts HPLC fingerprint. The research lays a theoretical and methodological foundation for effective exploitation and sustainable utilization of Radix Gentianae resource. The following several aspects were studied, the results are as follow:1.The components of Gentiana scabra Bunge were isolated and prepared using super-sonic, resins and (ordinary, middle-pressure) chromatography. We separated 10%,20%,50% ethanol part of resins using ordinary and middle-pressure chromatography. Six components Al, A2, A3, A4, A5, A6 were gained. Identified by physical-chemical and organic spectroscopy, Six components were confirmed. A1,Gentiopicrin. A2,A3 are isomer and perhaps one of the follows: 3'-O-β-D-Glucopyranosyl Gentiopicroside, 4'-O-β-D-Glucopyranosyl Gentiopicroside, 6'-O-β-D-Glucopyranosyl Gentiopicroside, A4,3-O-β-D-Glucopyranoside, 5-O-[4-hydroxy-E-cinnamoyl-(6)-β-D-glucopyranoside], A5,sweroside;4'-O-[β-D-Glucopyranosyl-(1→3)-2,3-dihyvdroxybenzoyl],2',3',6'-tri-Ac. A6, 2,3-Dihydroxybenzoic acid; 3-Me ether,β-D-glucopyranosyl ester.Comparing ordinary, midpressure chromatography and HSCCC in preparing gentiopicrin from the respects of time, energyconsuming, predisposal, we concluded that HSCCC was more effective method.2. Using chloroform-methanol-N-Butanol (5:4:2:4) as the solvent system, We separated Gentiopicrin from Gentiana scabra Bunge by HSCCC for the first time. The lower phase was the mobile phase. The flow-rate was 2.0mL/min;Apparatus rotate speed was 820r/min.The immobile phase retention was 38%;Retention time of Gentiopicrin was 320～380min.The purity of gentiopicrin was over 96.68%.3. The orthogonal test of L₉(3⁴) levels and facts was designed by varied ethanol solvent concentration, super-sonic time and liquid-material ratio. Result: the best extracting craftwork is the solvent concentration 50%, super-sonic time 30min and liquid-material ratio 1:8. With this process, the extract yield was 45.90%, Gentiopicrin content was 5.456%.4. HPLC fingerprint of Gentiana scabra Bunge extract was established. The HPLC method was, chromatogram column, Hypersil ODS5μm(4.6mm×250mm), mobile phase A methanol, B water, gradient elution, 0～30min, A 20～80%; 30～45min, A 80～20%; 45～55min, A 20～20%. Uvλ=270nm, flow rate, 1 mL/min, column temperature,35℃.According to the determining result of ten batch samples, we demarcated ten common fingerprint peaks. The similarity of fingerprint was over 0.90. Several innovations are as follow in this research1. For the first time middle-pressure chromatography was used to research components of Gentiana scabra Bunge. Meanwhile, we set up the gradient elution mode of dichloromethane-methanol by middle-pressure chromatography, providing technic reference for separating the component.2. For the first time.High-Speed Counter-Current Chromatography was used to separat and prepare Gentiopicrin, setting up a new method of isolating gentiopicrin.3. For the first time we established the HPLC finger print of Gentiana scabra Bunge extract, providing the quality standand for the extracting craft.