| Pueraria is an important constituent for drug and food since it has a lot of nutriments and particular meditative effects and has been widely used. As the main active component of Pueraria,Puerarin has been paid more and more attention and known for its meditative health protection function and clinic cure effect. Flavnonoids are eagerly needed in either domestic or the international markets. The extraction and purification techniques are needed to improve at present. This paper systematical studied on the process of extraction,purification and clathration of the main component- the isoflavones. The content and result of this study as follows:1. L9(34) Orthogonal design was adopted to research the factors: concentration of ethanol,extraction time,extraction temperature and ratio of ethanol to materials. The results of variance analysis showed that concentration of ethanol﹥extraction temperature﹥ratio of ethanol to materials﹥extraction time,and according to the calculation results of the orthogonal design, it indicated that the best extracting condition was: 60% ethanol, ratio of ethanol to materials was 15, extract 1 hour under the temperature of 70℃. With the best extracting condition, the extraction rate of total flavonoids was 93%.2. Effect of ultrasonic frequency on extraction rate was studied in ultrasonic extraction research, the results indicate:the higher the ultrasonic frequency was, the greater the extraction rate of total flavonoids was. Under the 32kHz ultrasonic frequency and other conditions remain the same as reflux extraction with ethanol, it could reach 95% of the extraction rate after half an hour, significantly higher than the reflux extraction. In the kinetic research of the ultrasonic extraction, the equation of the curve simulation results show that the Cubic curve reflected the dynamic processes of the ultrasonic extraction on total flavonoids.3. The static and dynamic adsorption/desorption methods were used to research the effect of different ADS series macroporous resin, and the separation and purification process was evaluated by measuring the concentration of total flavonoid in the fractions with UV spectrophotometer. Results indicated the ADS-8 macroporous resin was the most effective compared with other macroporous resins. The optimum conditions were screened, which were 6.8 mg/ml corresponding to crude drug for concentration of extract, pH 5–6 with the adsorption speed 4BV/h; water volume for removing impurities 400 ml; and the concentration of the eluant ethanol 70% (V/V), volume of ethanol 60 ml with desorption speed 2 BV/h. By this method, the final contents of total flavonoids exceeded 94%.4. The AKTA Prime system was used to separate flavnonoids with the methods of TLC and HPLC. Results indicated that the system could separate puerarin from the impurity, the content of puerarin was about 70% after sepatating. Puerarin in the sample was not interfered from the chromatogram of example compared to that of pueraria standard.5. Method of sequential change of molar concentration has been uesd to ensure the molar ratio of the inclusion complex (PU andβ-CD,HP-β-CD,M-β-CD,SPE7-β-CD) were all 1:1, and then the four kinds of inclusion complex were prepared by using ultrasonic method. The phase solubility method was used to prove the solubility-improvement and the stability constants were determined under the temperature of 25℃, 37℃and 45℃. The results indicated: the solubility of PU increased linearly with the increase of CD concentration, which showed typical AL type phase solubility profile, it also proved the molar ratio was 1:1; the solubility of PU was 1～3 times more in a series of concentrations of cyclodextrin; the stability constants of 25℃is higher than any others, and that of 37℃is the lowest. In addition, the results of stability investigation indicated that the inclusion complex was more stable than the physical mixture. |
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