Studies On Preparation, Purication And Biological Activity Of Actinidia Arguta Peptide

The northeast wild Actinidia arguta is an important wild fruit resource. Actinidia argutaSieb.et Zucc belongs to Actinidiaceae of actinidia. Actinidia arguta is named that cane jujube,cane malon and actinidia pear. This wild plant distributes in the North China, Northeast China,Northeast China and Yangtze river, and chiefly concentrated in the Changbai mountains in theNortheast China. Its root, stem, leaves and fruit are all useful for us in their medical value andhealthy function. The root has the effect on the Promotingdigestion,clear heat and diuresis,the leaves contains lots of chlorophyll, carotene, potassium, sodium and etc. the fruitscontains lots of SOD, amino acid, sugars and etc. meanwhile, the fruits has the effect on thehyperlipidemia, hypertension, and stenocardia.Polypeptides is the study of a hot spot, at present, The researchers have no furtherresearch and study in the actinidia arguta. We can only search the reports of fruit SOD,Amino acids, and Enzymes. This article offer the important value on the total exploitation ofactinidia arguta.In this paper, preparation, purification and biological activity of actinidia argutapolypeptide was systematic studied. The extraction yield index as index, the actinidia argutaprotein extraction was used by alkali method, the optimum extraction conditions weredetermined through the response surface analysis method. The degree of hydrolysis as index,the actinidia arguta polypeptides preparation process was used by enzyme method, theoptimum conditions were determined through the response surface analysis method.Then, separate and purify the polypeptides of actinidia arguta by using the ultrafiltration,Gel chromatography of Sephadex G-25, ion-exchange column chromatography of DEAE-32and RP-HPLC. Identify the purity by using the cellulose acetate membrance electrophoresis.The antioxidant capacity of actinidia arguta polypeptide was comprehensive evaluated by invivo and vitro test.in the next step, and then have researched the stability of actinidia arguta.The Restrain bacterial capacity of actinidia arguta was comprehensive evaluated bydetermine the Antibacterial spectrum.The function of immune enhancement of actinidiaarguta polypeptide were evaluated by lymphocyte transformation test, determination of halfhemolysis value, phagocytic index, immune organ index and toes degree of mice.Use the multihole starch as the core materials, we encase the polypeptides of actinidiaarguta individual by gelatin and β-cyclodextrine. In the Safety evaluation on actinidia arguta.we use the Acute toxicological method, Ames method, PCE method, comets method and soon.ensuring the safety of the polypeptides of actinidia arguta.This paper makes a study of the conclusions are as follows:1.The protein content is1.25%in the fresh fruit of actinidia arguta.in the dried fruits,thecontent is7.5%.The optimum extraction conditions of alkali method are as follows: solid-liquid ratio of1:19, pH7.9, temperature50℃, time2.3h, protein extraction rate is53.23%.2.Use the degree of hydrolysis as index, we enzymolysis the protein of actinidia argutaby trypsin,papain and Neutral protease, the result showed that trypsin is better than the other.the optimum enzymolysis conditions were determined through the response surface analysismethod, the results as follows: pH7.14, enzyme activity7500u/g material, temperature56.43℃, Substrate concentration1.86%, under these conditions for hydrolysis of actinidia argutaprotein, degree of hydrolysis is23.32%, peptide yield is60.36%. there is17kinds of aminoacid by the automatic analyzer analysis, Contains7kinds of human body essential aminoacids.3．Ultrafiltration were carried out to separate actinidia arguta protein hydrolysate,thebetter conditions are:pressure0.06Mpa, temperature30℃, rate of velocity2.25ml/s.Theboth compontes of polypeptides has function on antioxidant.But the less than10KDamolecular weight is stronger SephadexG-25column chromatography was used to separatelow molecular weight of actinidia arguta protein hydrolysate, three separate peaks were got,of which the third peak with the strongest antioxidant activity. DEAE-32cellulose columnwas used to separate the active component purified by the SephadexG-25, four separate peakswere got, of which the fourth peak with the strongest antioxidant activity. RT-HPLC was usedto separate the active component purified by the DEAE-32cellulose column, two peaks wereisolated, of which the first has higher antioxidant activity. After the series of purification, thecontent of peptide is98.36%.4. Bacteriostatic experiment shows that the polypeptides of actinidia arguta has thebroadly function on restrain the activity of bacteria,but except the yeast and mold.5. The immune experiment shows that polypeptides of actinidia arguta can prolong theswimming time of weighted mice, increase the content of glycogen, enhance the enduranceof mice and easing the fatigue of mice.6. The Stability experiments showed that the polypeptides of actinida arguta keep thestability at pH6-8, The higher of the temperature,the more stability. The glycerin, sucrose,lactose, mannitol all can enhance the activity of stability.of which the glycerin is better. In theexperiments of Simulation gastric juice to digest, both the pepsin and trypsin can reduce theactivity of antioxidant.7. The Microcapsule experiments shows: the better process is Use the multihole starch asthe core materials and use the β-cyclodextrine as the material of wall.The capsule maintainedthe activity of antioxidant and keep dried.8. Toxicity test show that actinidia arguta peptide in mice acute toxicity test, LD50>20g/kg, is non-toxic class. Results of mouse bone marrow micronucleus test, Ames test, thechromosome aberration test of mouse primary spermatocyte and comet assay were negative. Actinidia arguta peptide is a non-toxic material. The present studies thus provide preliminaryproof of actinidia arguta peptide for the application of health food safety.