Studies On Hypoglycemic Substances And Mechanism Of Onion Polypeptide

Diabetes mellitus(DM)is a metabolic disease characterized by hyperglycemia,which is due to the deficiency of insulin secretion or the damage of its biological function,or both of them.DM can lead to impairment or dysfunction of a variety of tissues,especially the kidney,heart,blood vessel,eye,and nerve,etc.At present,DM has become the third largest killer after cancer and leukemia,of which global prevalence population has exceeded 400 million.Onion is a traditional plant with medicine and food dual purposes,which is rich in nutrients,such as a variety of cellulose,protein,vitamins,saponins and other effective components,and which possessed many physiological effects,including the decrease of blood pressure,blood sugar,blood lipid,and anti-cancer.It has been proved that the onion contains a kind of peptide which can inhibit the fungi or prevent the loss of osteoporosis,but the research on the hypoglycemic effect of the onion peptide has not been reported.The research work of this paper mainly focused on the preparation,separation,purification of onion peptide,and the hypoglycemic effect and mechanism in vitro and in vivo were also studied.Firstly,the optimal enzymatic hydrolysis conditions prepared for onion peptide were determined by orthogonal test.Secondly,the crude onion peptide was separated by gel chromatography and HPLC technology.Thirdly,the hypoglycemic effect of the onion peptide components was carried out in vitro.Finally,the hypoglycemic effect of onion peptide and protein was performed in vivo,which the biochemical analysis and histological examination were utilized to study the hypoglycemic effect and mechanism.The main results of this work were as follows:(1)The optimal technological conditions for the preparation of onion peptide were:the given mass of acid protease was added to the 0.20 g/mL onion protein solution,ofwhich the weight was according to the ratio of protein to enzyme(0.6:1),mixed well,and hydrolyzed at 55 ℃ for 2 h in pH 3.5,inactivated the acid protease at 95 ℃ for 15 min,then according to the ratio of protein to enzyme(0.3:1)to add pepsin into the mixing solution,mixed well and hydrolyze at 37 ℃ for 2 h in pH 1.5,inactivated the pepsin at95 ℃ for 15 min,and cooled to room temperature followed by frozen centrifugation for20 min at 20000 r/min,frozen-dried supernatant fluid,and then obtained the onion polypeptide powder.The method was simple and environmental friendly.Additionally,the α-amylase inhibition activity of obtained onion peptide was 86.85%,and the hydrolysis degree was 30.12%.(2)The optimal separation conditions of the crude onion polypeptide by the gel chromatography were as follows: 0.01 mol/L(containing 0.05 Na Cl mol/L)phosphate buffer(PB)was as mobile phase,sample concentration was 8 mg/m L.Therefore,four kinds of onion polypeptide component(P1-P4)could be enriched conveniently,and the separation degree of each component was high under this conditions.(3)The P1,one of four onion polypeptide components,had higher α-amylase inhibitory activity and α-glucosidase inhibitory activity,the best inhibitory concentration of P1 was 20.0 mg/mL and 30.0 mg/m L,respectively,and the inhibition rate reached53.8% and 92.6%,shown the higher hypoglycemic activity in vitro.The optimal conditions for purified P1 by HPLC were as follows: mobile phase was water: acetonitrile(97:3,both containing 0.1% TFA),stationary phase was C18,injection volume was 20 μL,flow rate was 0.5 mL/min,and the detection wavelength was 270 nm.Under the above conditions,the main peak of the retention time was 4.618 min,the separation degree was 1.388,and the peak area was 85.03%.(4)Onion protein and polypeptide could effectively reduce blood glucose in T2 DM mice and increase their sugar tolerance,and had certain inhibitor effect on liver and kidney damage caused by hyperglycemia.In addition,both of onion protein andpolypeptide could recover the physiological indexes related DM to the normal level,such as fasting blood glucose(FBG)、 AST 、UREA 、TC 、TG,and the effect of onion polypeptide is better than that of protein,the reasons may be that P1 possessed higher inhibitor activity of α-amylase and α-glucosidase,but the hypoglycemic target and pathway need to be further studied.