The Study On The Isolation Of Tannins From Cyclobalanopsis Glauca And Its Anti-fatigue And Anticancer Activities

Cyclobalanopsis glauca (Thunb.) Oerst, is a member of the Fagaceae family. It contains a series of bio-compounds, including tannins, polysaccharide, alkaloids, polyphenols, VB1, VB2, Vc etc. Among them, the tannins are responsible pharmacological active substance. The tannins from cyclobanopsis have antifatigue, anticancer, anti-aging, antibacterial, antivirus activities as well as cardio-protective properties.The research focused on the tannins form Cyclobalanopsis glauca fruit (CGFT). Tannins was extracted by ultrasonic technique, the extraction process was optimized; Then products were separated and purified through macroporous adsorbent resins. And anti-fatigue and anticancer of the purified products were studied. The main results were as follows:(1) The main polyphenolic substances, including total phenol (2.433%), total flavones (3.25%) and tannins (1.821%) were determined.(2) On the base of the single factor experiments of ultrasonic-assisted extraction technique of tannins from Cyclobalanopsis glauca fruit (CGF), the method of response surface analysis (RSA) with four factors and three levels was adopted and the factors influencing the technological parameter were determined by means of regresss analysis. The results showed that the factors influencing extract yields were not linear, and the intensity of the impacts on the yields of CGFT were as follows:ultrasonic power> ethanol concentration> extraction temperature> the ratio of material to solvent. And the optimum conditions were:50℃of extraction temperature,61%of the ethanol concentration,313W of the ultrasonic power,31:1of the ratio of solvent to material, under such condition the extraction rate of tannins from CGF was1.978%.(3) Six kinds of macro porous resins were selected to compare their performances in absorbing and desorbing tannins from CGF. Experimental results show that SP-700resin possesses higher absorption (87.32%) and desorption capacity (85.69%). The optimum parameters for separation were as follows:sample concentration was0.4mg/mL; Flow rate of the adsorption was1.0mL/min; The optimal solution for the desorption was70%ethanol at the flow rate of1.0mL/min. The purity of CGFT was from7.71%to28.76%.(4) Five different chemical models in vitro were used to evaluate the antioxidant activities of CGFT. The actual EC50value of the1,1-diphenyl-2-picrylhydrazyl (DPPH) and superoxide anion radical scavenging activity of CGFT were1.73mg/mL and1.03mg/mL, which was lower than that of glutathione (GSH)(3.34mg/mL and3.67mg/mL). The reductive potential activity of GCFT was a little bit less than that of GSH. There was no significant difference in their reducing power, when the mass concentration of CGFT was2fold of that of GSH. The ferrous ion chelating activity of CGFT increased with increasing concentration, and it showed an excellent Fe2+-chelating ability of79.82+2.03%at5mg/ml. CGFT showed a certain degree of inhibition of erythrocyte oxidative hemolysis induced by H2O2, and the IC50value of H2O2-inhibition was18.76mg/mL. CGFT could improve the activity of the endogenous cellular antioxidant enzymes in mice. It could increase the activities of SOD, CAT and GSH-Px by7.4%-15.3%,27.7-56.1%and26.9%-43.2%, respectively. The results indicate that CGFT have potential antioxidant activities.Animal models were used to evaluate the anti-fatigue effects of CGFT. Results showed that CGFT could significantly increase the swimming time of mice by30%-45%, could delay or decrease the consumption of glucose and liver glycogen and improve the elimination of lactic acid and blood urea nitrogen. These results indicate that CGFT have potential anti-fatigue effects. The results in the present work clearly indicate that CGFT has significant antioxidant activity against various antioxidant systems in vitro. And data also suggested that CGFT possesses direct and potent anti-fatigue activities through multiple mechanisms. Especially, the antioxidant activities of CGFT had good relationship with its anti-fatigue effect.(5) The inhibitory of CGFT on human cervical Hela, human liver HepG2and human lung H460cancer cells were studied by MTT colorimetric assay in vitro, and the mouse tumor model was used to investigate the effects of CGFT on tumor growth in vivo. Results showed that CGFT had significant inhibition effects on the growth of human cervical Hela, human liver HepG2and human lung H460cancer cells with the IC50value of2.420mg/mL,12.365mg/mL and14.240mg/mL, respectively. CGFT also showed good inhibitory effect on the growth of Si so sarcoma-transplanted mice and increased with increasing concentration. The inhibition rate of Sigo sarcoma reached57.49%,41.79%and28.17%at the dose of100mg/kg,50mg/kg and10mg/kg, respectively.