Studies On Separation And Purification Of Active Component From Traditional Chinese Medicinal Herbs (Rheum Officinale Baill., Psoralea Corylifolia L., Etc) By High-speed Counter-current Chromatography

High-speed counter-current chromatography (HSCCC) is a form of liquid-liquid partition chromatography, which was developed in the 1980's. Compared with conventional chromatographic methods, HSCCC does not use any solid matrix. Therefore, it eliminates irreversible adsorption, decomposition, contamination and many other disadvantages owing to chemical reactions of samples with solid support. In addition, it has many advantages over traditional chromatographic methods such as high efficiency, high loading capacity and high recovery, so it is very suitable for separation and purification of fragile compounds from traditional Chinese herbs and other natural products. With the development of nearly 30 years, it has made great progress in theory and technique. HSCCC has gained extensive application in the fields of biology, medicament, foodstuff, material, agriculture, environment protection and so on. Especially in the aspect of preparative separation and purification of active components from traditional Chinese herbs, it has become one of the most preponderant separating and analytical techniques. As an important part of world medicine treasure, Chinese traditional medicine is the quintessence of China. With the history of more than 2000 years, it has contributed to the existence and propagation of our Chinese people. Under the influence of the thought of "return to the nature", it is becoming more and more popular with people all around the world. Compared with western medicine, it has many strong points such as abundant resource, unique curative effect, lesser side effect and toxicity. But the most serious disadvantage is that the quality control of traditional Chinese herbs and its products lacks clear standard which is the biggest obstacle to the modernization of Chinese traditional medicine. In addition, the reason why it can treat diseases is not very clear and the research and development on pharmacological effect progress slowly, which limits its development to some extent. Therefore, it has become an urgent task to investigate Chinese traditional medicine with modern technological instrument, clarify the reason why it can cure ailments and establish integrate quality appraise system. At the present time, methods of separating and purifying active components from traditional Chinese herbs are preparative high performance chromatography, column elution, HSCCC and so on. Applying HSCCC to the study on Chinese traditional medicine can realize preparation of high purity standard materials, preparation of active parts or active components associated with pharmacological test, and large-scale production. And it can also supply more abundant and exact information for the establishment of fingerprinting of traditional Chinese herbs and correlative products. In the present paper, under the optimized experiment conditions, active components from common traditional Chinese herbs(Rheum officinale Baill.,Angelica dahurica (Fisch. ex Hoffm) Benth, et Hook. f ,Psoralea Corylifolia ,Epimedium koreamum Nakai and Aucklandia lappa Decne)were separated and purified. The purity of the obtained compounds after HSCCC separation was above 98% determined by HPLC analysis and their chemical structures were identified by 1H-NMR and 13C-NMR. It can supply some reference for pharmaceutical test and identification and quality control of the above herbs and the correlative products. â… ,Preparative isolation and purification of 5 hydroxyanthraquinone active components and cinnamic acid from Rheum officinale Baill. using HSCCC Crude extract from Rheum officinale Baill. was isolated and purified by HSCCC using pH gradient elution. Aether was used as the stationary phase. 1% NaH2PO4 and 1% NaOH solution was used as the mobile phase to perform pH gradient elution. The method yielded 18.8 mg of rhein, 18.4 mg of emodin, 14.2 mg of aloe-emodin, 10.1 mg of chrysophanol, 5.5 mg of physcion and 19.0 mg of cinnamic acid from 120.5 mg of crude sample. The purity of the 6 components was above 98% determined by HPLC analysis, and their chemical structures were identified by 1H-NMR and 13C-NMR. The method can separate and purify 6 compounds in 500 min and the result was gratifying. â…¡,Preparative isolation and purification of 3 coumarin active components from Angelica dahurica (Fisch. ex Hoffm) Benth, et Hook. f using HSCCC Aether extract of Angelica dahurica (Fisch. ex Hoffm) Benth, et Hook. f was separated using HSCCC. In the method the upper phase of n-hexane-methanol-water (5:5:5, v/v) was used as the stationary phase of HSCCC. The mobile phase used in HSCCC was the lower phase of n-hexane-methanol-water (5:5:5, v/v) and n-hexane-methanol-water (5:7:3, v/v) that was changed in gradient. 29.1 mg of imperatorin, 28.4 mg of isoimperatorin and 35.2 mg ofoxypeucedanine were yielded from 100.2 mg of crude sample, each at over 98% purity as determined by HPLC analysis. The peak fractions of HSCCC were identified by 1H-NMR and 13C-NMR. Using gradient elution method, 3 compounds with similar chemical structures and properties were purified within 6 h. It can be seen that gradient elution is an efficient approach for complicated samples. â…¢,Preparative isolation and purification of 2 coumarin active components from Psoralea corylifolia L. using HSCCC Light petroleum extract of Psoralea corylifolia L. was purified using HSCCC. n-hexane-ethyl acetate-methanol-water (5:5:4.5:5.5, v/v) was used as two-phase solvent system. The upper phase was used as the stationary phase and the lower phase was used as the mobile phase. 39.6 mg of psoralen and 50.8 mg of isopsoralen were yielded from 100.8 mg of crude extract after one-step HSCCC separation. The purity of them was above 99% determined by HPLC analysis and their chemical structures were identified by 1H-NMR and 13C-NMR. Using this method a pair of position isomers with similar properties obtained purification in 4 h and the result was satisfying. Thus it can be seen that separation efficiency of HSCCC is more remarkable. â…£,Preparative isolation and purification of 3 flavonoid glycoside active components from Epimedium koreamum Nakai by HSCCC Ethanol extract of Epimedium koreamum Nakai was purified using HSCCC. Chloroform-methanol-water (4:3.5:2, v/v) was used as two-phase solvent system. The upper phase was used as the stationary phase and the lower phase was used as the mobile phase. 46.5 mg of icariin, 11.4 mg of epimedokoreanoside I and 17.7 mg of icariside II were yielded from 200.3 mg of crude extract. The purity of them was over 98% determined by HPLC analysis and their chemical structures were identified by 1H-NMR and 13C-NMR. â…¤,Preparative isolation and purification of 2 sesquiterpene lactone active components from Aucklandia lappa Decne by HSCCC Costunolide and dehydrocostuslactone were separated and purified from Aucklandia lappa Decne by HSCCC. A two-phase solvent system composed of light petroleum-methanol-water (5:6.5:3.5, v/v) was used for HSCCC separation, and the upper phase was used as the stationary phase. 35.7 mg of costunolide and 43.6 mg ofdehydrocostuslactone were obtained from 110.5 mg of crude extract after HSCCC separation. The purity of them was above 99% as determined by HPLC and their chemical structures were identified by 1H-NMR and 13C-NMR.