Separation And Purification Of Saponins And Preparation Of Sapogenins From Chenopodium Quinoa Willd

The quinoa saponin mainly exists in the mastoid cells of quinoa husk.The toughness of cell wall makes that it difficult to separate and extract.The extraction rate of saponin is lower by traditional and single methods.And the use of acid,energy consumption and waste acid solutionof hydrolysis is large amount.Due to many kinds of quinoa sapogenins,little structural difference and lower content,it is very difficult to prepare high-purity sapogenin.So far,there is no effective method to preparing high-purity saponins.A method of multiple enzymes assisted with ultrasonic technology-purification by macroporous resin–acid hydrolysis and preparative high performance liquid chromatography(P-HPLC)toextract from quinoa husk has been established.The extraction rate of quinoa saponin was improved.The macroporous resin was selected topurify quinoa saponins to improve the purity of saponin,an effective method forthe preparation of sapogenin was established by acid hydrolysis,then the puresapogenin was obtained by P-HPLC?The main study work include as follows.1.Saponin was extracted from quinoa husk using single cellulase assisted with ultrasonic extraction,and the technology parameters were optimized by orthogonal experiment.And the optimum parameters were as follows: enzyme dosage(based on the mass of quinoa husk,the same below)of 1.0%,enzymatic hydrolysis temperature of 45?,pH of 5.0,enzymatic hydrolysis time of 15 min,and the yield under this condition was 81.56%.Saponin was extracted from quinoa husk using single pectinase assisted with ultrasonic extraction,and the technology parameters were optimized by orthogonal experiment.And the optimum parameters were as follows: enzyme dosage of 1.5%,enzymatic hydrolysis temperature of 50?,pH of 4.5,enzymatic hydrolysis time of60 min,and the yield under this condition was 82.20%.2.Based on the single enzyme assisted ultrasonic extraction saponin from quinoa husk,saponin was extracted from quinoa husk using multiple enzymes assisted with ultrasonic extraction,and the extraction was evaluated.According to the Box-Behnken experimental design principle,the response surface methodology was used to optimize the technical parameters.And the optimum parameters were as follows: total enzyme dosage of 1.5%,enzyme ratio(mass ratio of cellulase to pectinase)of 3:2,enzyme hydrolysis temperature of 50.5?,pH of 5.5,and enzyme hydrolysis time of17 min.Under the optimum conditions,the extraction design of quinoa saponins reached a maximum of 85.32%,which was 4.41% higher than using only cellulase(81.56%),3.66% higher than using only pectinase(82.20%),and 16.76% higher than using only ultrasonic extraction.3.Comparison of the adsorption and purification effects of S-8,D101,NKA-9,HPD-100,HPD-850 and AB-8 macroporous resins on quinoa saponins,the adsorption and purification effects of HPD-850 macroporous resin was the best,and the adsorption process conforms to the Pseudo first order kinetics equation,indicating that the adsorption process of HPD-850 macroporous resin on saponin is a reversible adsorption process.And the optimum parameters were as follows: flow rate of 2.5 mL/min;sample loading of 80 mL;ethanol concentration of 70%,elution flow rate of3.3 mL/min,the liquid volume of 70 mL.After purification by HPD-850 macroporous resin,the purity of quinoa saponin was increased by 52.44%,the content of impurities such as pigment,tannin and sugar was reduced.4.Quinoa saponins purified by macroporous resin were hydrolyzed by acid to prepare sapogenin.The effects of hydrochloric acid concentration,temperature and time on the extraction amount of sapogenin were studied.The technical parameters were optimized by response surface methodology.And the optimum parameters were as follows: hydrochloric acid concentration of 0.9 mol/L,temperature of 90?,enzymolysis time of 90 min,to reduce the use of acid,reduce environmental pollution,and improve the extraction amount of saponins.The purified sapogenin was prepared by preparative high performance liquid chromatography,and the optimal parameters for preparative chromatography were obtained.The mobile phases were acetonitrile whichis A and water which is B;gradient elution was performed at 0-20 min,70% A-85% A,20-40 min,90% A,flow rate was 10 mL/min,column temperature was room temperature.The retention time of oleanolic acid and hederagenin were 22.528 min and 10.230 min respectively.5.UPLC-MS was used to verify the structure of the sample and the comparison of TIC,EPI,XIC of standard and sample showed that the sample contained oleanolic acid and hederagenin.