| Free radicals which impair the cells induced to diseases, are generated constantly in the process of natural metabolism of organism.Recently studies had shown that the occurrence and development of typeⅡdiabetes is related to the generation of free radicals. Antioxidants had the function of preventing diseases and slowing the aging process by eradicating free radicals in vivo, protecting cells and tissues from oxidative damage. In recent years, the natural antioxidant peptides had aroused people’s universal attention, while there is still little further research especially on the structure of antioxidant peptides. Therefore our study of the enzymolysis,isolation, purification, antioxidant activity and structure of cottonseed protein, as well as the therapeutical effect of cottonseed protein monomer to type Ⅱ diabetes helps to produce cottonseed protein monomer with high inoxidizability, evaluate the therapeutic effect to typeⅡdiabetes. We also can explain the therapeutic function of cottonseed protein monomer to typeⅡdiabetes as the point of antioxidant.We selected the cottonseed Protein as experimental materials, hydrolyzed by protease,then crude separated by centrifuging, filtering and ultrafiltration, finally collected the hydrolysate of cottonseed protein under 3000 Da. Subsequently we purified the product by DEAE Sephadex A-25 ion exchange chromatography and Sephadex G-10 gel filtration chromatography. Each component of chromatography had high inoxidizability by detecting in vitro. We finally collected 3 kinds of cottonseed peptides monomers by LC-MS/MS and identification of amino acids sequences. We used those cottonseed peptides monomers in the model of typeⅡdiabetes mouse induced by high fat diet combined with STZ in small doze,detected the Fasting plasma glucose(FPG)、serum insulin(INS)、glycosylated hemoglobin(Hb A1c)、triglycerides(TG)、total cholesterol(TC)、malondialdehyde(MDA)、superoxide dismutase(SOD) and glutathione(GSH) of mice. We then observed the pathological symptom of pancreases and kidneys of typeⅡdiabetes mice reacting with cottonseed peptides monomers by HE staining method.This study indicated that the optical temperature of protease was 50℃, the optical p H was8.0. Hydrolysis time was 100 min and the degree of hydrolysis was 23.6%. The structures of three cottonseed peptides monomers are Thr-Asp-Gln-Leu(MW:475 Da), Gly-Asp-Leu-Leu(MW:416 Da) and Leu-Leu-Glu-Pro-Ala(MW:541 Da). The model of typeⅡdiabetes mice had high rate of estabilishment(80%) which could induce typical symptom of typeⅡdiabetes such as hyperglycaemia、hyperlipidemia、hypoinsulinism and insulin resistance. cottonseed peptides monomers could reduce obviously the levels of FPG and Hb A1 c in serum of typeⅡdiabetes mice, enhance the level of INS in serum and inhibited insulin resistance. cottonseed peptides monomers could reduce the level of MDA, enhance the levels of SOD and GSH in serum and the ability of antioxidant, as well as inhibit the stress response. cottonseed peptides monomers could also alleviate the lesion of pancreas and kidney in type Ⅱdiabetes mouse, enlarge the volume of pancreas islet and increase the number of Islet cell.In conclusion, we getted three cottonseed peptides monomers with high inoxidizability by the method of enzymolysis, the structures of which are Thr-Asp-Gln-Leu(MW:475 Da),Gly-Asp-Leu-Leu(MW:416 Da), Leu-Leu-Glu-Pro-Ala(MW:541 Da). Cottonseed peptides monomers could obviously reduce the levels of blood glucose、blood fat of typeⅡdiabetes mice,regulate the metabolism of glycolipid, increase the level of INS in serum and develop the resistence of insulin. Cottonseed peptides monomers could enhance the ability of antioxidant,reduce lipid peroxidation and the generation of oxygen free radicals, protect the pancreas 、kidney of type Ⅱ diabetes mice on the basis of regulating the metabolism. Accumulated the results, the important information on the extensive application of Cottonseed peptides monomers to the treatment of typeⅡdiabetes. |
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