Preparation Of Ginkgo Active Peptide And The Research Of Functional Peptide Product

In this dissertation,ginkgo seed was used as raw material to study the preparation process of ginkgo hypoglycemic peptide.Then one kind of functional peptide product with hypoglycemic effect was developed.Further,the hypoglycemic activity of ginkgo active peptide and functional peptide product in vivo was verified through animal test.The research would provide a theoretical basis for the comprehensive utilization of ginkgo peptide.The main findings were as follows:(1)The ?-glucosidase inhibitor is an oral hypoglycemic agent that could treat diabetes by delaying the absorption of intestinal carbohydrates.it could inhibit various ?-glucosidases located in the small intestine competitively,so that the starch into glucose slowed down,thereby slowing the intestinal absorption of glucose,reduce postprandial hyperglycemia.In the previous studies,the ?-glucosidase inhibitory activities was often acted as an evaluated index Of the hypoglycemic activity.In this study,?-glucosidase inhibitory peptide was prepared by enzymatic hydrolysis of ginkgo biloba protein,test with ?-glucosidase inhibition activities as index,by comparing the enzymatic hydrolysis effect of five kinds of commercial proteases,the inhibition activities of ?-glucosidase of papain was 13.01%,papain was chosen as the best enzyme source.The factorial design experiment and response surface experiment were conducted to optimize the conditions of enzymatic hydrolysis processing,results showed that the optimum enzyme conditions were: p H7.36,extraction time of 4.4 h,E/S ratio of 5%(w/w)and temperature of 57.9?,The highest ?-glucosidase inhibitory activities was 17.18%.(2)In order to obtain a higher active peptide,first the ginkgo crude peptide was divided into six components by ultrafiltration,and ?-glucosidase inhibition activities of the two components of <3 k Da and 3~10 k Da were 50.43% and 43.59%,respectively.Which indicated that the small molecule peptide has a higher activity.Then,the two components of <3 k Da and 3~10 k Da were further separated and purified by HPLC.The four components of A1 and A2,B1 and B2 were obtained respectively.The molecular weight was 927.80 Da,1063.80 Da,3107.00 Da,3129.60 Da,respectively.The results showed that the four peptide components contained hypoglycemic-related amino acids.(3)Studies have shown that amino acids such as glutamic acid,aspartic acid and proline are related to the hypoglycemic effect of peptides.The amino acid composition of A1,A2,B1 and B2 was analyzed by amino acid automatic analyzer.The main amino acids of A1 were cysteine(Cys)which content was 9.02 g/100 g,glutamic acid(Glu)20.47 g/100 g and aspartic acid(Asp)8.29 g/100 g,the major amino acids of A2 were glutamic acid(Glu)19.13 g/100 g,aspartic acid(Asp)9.93 g/100 g and arginine(Arg)9.93 g/100 g,the main amino acids of B1 were valine(Val)8.24 g/100 g,methionine(Met)9.71 g/100 g,cystine(Cys)12.24 g/100 g,tyrosine(Tyr)8.47 g/100 g of,Glutamic acid(Glu)18.82 g/100 g and aspartic acid(Asp)9.47 g/100 g,the main amino acids of B2 were glutaric acid(Glu)20.35g/100 g,aspartic acid(Asp)13.31 g/100 g and arginine(Arg)8.84 g/100 g.The results of the test showed that the four peptide components contained hypoglycemic-related amino acids.(4)The functional peptide products were prepared by using the <3 k Da peptide that prepared by the previous experiment,and the taste,efficacy and stability of the functional peptide products were evaluated.The flavor formula and stabilizer formula of the functional peptide product were studied through the factorial design experiment and response surface experiment.Considering the taste,efficacy and stability of functional peptide products,the optimum process of developing product as follows: the addition amount of xylitol was 5%,the addition amount of citric acid was 0.3%,the amount of glycine was 1.2%,the addition amount of xanthan gum and sodium carboxymethyl cellulose were 0.02% and 0.04%(m/v),respectively.(5)In order to verify the hypoglycemic activity of ginkgo biloba active peptide and functional polypeptide product,mice induced by streptozotocin were used for 30 days feeding test,the body weight,blood glucose level,triglyceride level and total cholesterol level were monitored.Ginkgo active peptide and the functional peptide product had no effect on diabetic mice weight loss symptoms;the total hypoglycemic rates of the low,middle,high dose group of ginkgo active peptide and functional peptide product were 68.66%,73.23%,70.19%,69.80%,65.30% and65.56%,respectively,Which idicated ginkgo biloba active peptide and functional peptide products have a good hypoglycemic effect;the reduction rates of the triglyceride level of the low,middle,high dose group of ginkgo active peptide and functional peptide product were 8.60%,12.74%,12.80%,6.73%,12.75% and22.47%,respectively;the reduction rates of total cholesterol of the low,middle,high dose group of ginkgo active peptide and functional peptide product were 6.73%,12.75%,22.47%,15.00%,23.04% and 26.96%,respectively,Which indicated that ginkgo biloba active peptide and functional peptide products can effectively reduce the triglyceride and total cholesterol of diabetic mice.