Study On Extracting And Purifying Diosgenin From Dioscorea Zingiberensis C.H.Wright Through Enzymatic Zymotechnics

Dioscorea Zingiberensis C. H. Wright(Chinese yam is a special kind of plant, which also named HuoTou geng or KuLiang jiang. The diosgenin is white crystal with stimulating smell. It's hard to dissolve in water, but easy to dissolve in some organic solvent like benzene, chloroform and so on. The structural formula of diosgenin is C₂₇H₄₂O₃, and the molecular weight is 414.61.The diosgenin is the hydrolysate of Dioscorea Zingiberensis C. H. Wright, which has tremendous medical value and marketable potential. The diosgenin has many physiological functions which plays an important role in human health, the world of biology calls it as "the key of life", the field of medication calls it as "the gold in medication, the mother of hormone". This paper studied the component of Dioscorea Zingiberensis C. H. Wright and the different methods of conservation. We judged the diosgenin's properties of physical chemistry, compared the extractation methods, including the tradition method and enzymatic zymotechnics. Finally, confrimed the best technics conditions. Ultrasonic may influence the structure of Dioscorea Zingiberensis C. H. Wright, so we contrasted ultrasonical extractation and ultrasonic with enzyme. The optimized conditions for the extraction and purification of diosgenin were obtained. The resultsindicated:1. Starch (26.99%) and coarse fiber (57.7%) content were the most among all the components of Diascorea Zingiberensis C. H. Wright which also had 5.24% dioscin. So we chose amylase, cellulose enzyme and compound enzyme to use in the experiment. Refrigeration was a good way to preserve the fresh Dioscorea Zingiberensis C. H. Wright and had no adverse effect on the extraction of diosgenin. 2. In the diosgenin determination experiment, we used two methods. One was Colorimetry of vanillin-acetic acid, another was Colorimetry of vitriol-methanol.The latter one was better with single apex appears at 414nm, which had small relatively standard deviation. We can find the best technics from orthogonal experiment by acid hydrolyzing directly: acid concentration is 3 mol/L, the time of acid hydrolyzing was 4 hours and the fermentation time with 60ml water was 24 hours. Acid concentration leads the first factor in the method and the water amount was the last one.3. Monofactorial experiment helped select out 4 factors and 3 levels which influenced thermoduric amylase and gave the best processing parameters as A₂B₃C₃D₂ which can accept 0.4935g diosgenin, 20.13% higher than traditional method, namely the Parameters just like the pH was 6, the dosage of emzyme was 0.1mL, the temperature was 50℃, and the time of fermenting was 36 hours. Of the 4 factors, temperature affected most while action time of amylase was the least. In other words, pH affected greater than amylase dosage while smaller than temperature.4. Slected four factors which effected more on thermostable amylase, we got the best technics parameters as A₃B₃C₃D₂, which can accept 0.4999g diosgenin, 21.69% higher than traditional method, the Parameters just like the pH was 6, the dosage of emzyme was 0.1mL, the temperature was 50℃, and the time of fermenting was 24 hours. Of these four factors, we found that the tempreture effected most on amylase, and the dosage of emzyme was the least. Comparing the two experiments with amylase or not, we found the output of diosgenin adding amylase was more than without amylase, also we can get more green sugar by adding amylase athough the total sugar was almost same. Analysing the relationship among the output of diosgenin, the weight of residue and sugar, we came to a conclusion that using little acid can decrease pollute by adding amylase which saving 33.33% acid.5. Considering the influence of ultrasonic on the structure of Dioscorea Zingiberensis C. H. Wright, we have compared the diosgenin extraction processing.We cannot get diosgenin only using ultrasonic extraction without fermentation. If we used ultrasonic to assist amylase extraction, the output of diosgenin were not higher obviously than amylase extraction, it would decrease if the ultrasonic power was too high or too low. If using ultrasonic instead of adding amylase with fermentation, we can get 0.4977g diosgenin which was 21.15% higher than taditional method while get 0.4994g diosgenin which was 21.56% higher by using ultrasonic with amylase fermentation. That was to say, ultrasonic can instead of amylase in a way, but the weight of sugar we got has decreased but the total sugar was invariability. Finally, considering the side effect to people and the higher cost of ultrasonic, we tought the amylase method was better.6. The diosgenin sample got from enzymolysis extraction was better than that got from direct fermentation in terms of sample melting point and purity. The melting point of the sample from enzymolysis extraction was 202-210℃and purity was over 88% while the latter from 198℃to 205℃while purity was 86%.7. Resin ADS-7 can adsorb and desorb diosgenin well with 94.35% absorption capability in a short adsorption and a short desorption. Ethanol had a strong ability both in adsorption and desorption, 95% of ethanol could almost desorb all the absorption. Add 8g ADS-7 into 200mL diosgenin solution with a concentration of 0.094mg/mL, then pump into ADS-7 with a constant flow of 1.0mL/min while collect effusion. After completed adsorption, taking 1 mL effusion to determine the content of diosgenin,drying the resin, adding 250mL desorption solvent into it, desorption ratio can come to 87.76%. Considering the cost, to use 250mL desorption solvent to determine the content of diosgenin, we got diosgenin purity was 94.12% which increased by 4.5% than before.