| Glycopeptide is linked by short sugar chains and short peptides,it is a compound which has a variety of biological activity,such as anti-oxidation activity,anti-tumor activity,antilipidemic activity and hypoglycemic activity.At present,glycopeptide is extracted from a variety of plants,such as soybean,dandelion,polygonatum and camellia oleifera.High-temperature soybean meal is a by-product of the extraction of soybean oil and has a huge yield.It is rich in nutritive value,in which protein content up to 50%.However,the high-temperature desolvation step in the oil refining process causes the heat denaturation of protein in high-temperature soybean meal,resulting in the decrease of some functional properties,which limits its application in the food industry.Therefore,in order to broaden the comprehensive utilization of high-temperature soybean meal resources,its hydrolyzate was selected as raw materials in this study,preparation,extraction,isolation,purification,properties characterization and antioxidant activity evaluation of glycopeptide were performed.The main results were as follows:1.Study on preparation technology of glycopeptide by glycosylation of high-temperature denatured soybean meal hydrolyzateThe effects of ultrasonic power,ultrasonic time,ultrasonic temperature,mass ratio of protein and glucose,protein concentration and pH value on degree of grafting and browning index were studied in the grafting reaction.Taking the significant factors as independent variables and the grafting degree as the response value,the response surface optimization experiment was carried out to determine the optimum parameters of preparation of glycopeptide by glycosylation of high-temperature soybean meal hydrolyzate.The results showed that the power was 150 W,the temperature was 75 ?,the time was 75 min,the mass ratio of protein and glucose was 1:2,the protein concentration was 5%,the pH value was 9.0,the degree of grafting and browning index were 26.41±0.27% and 0.419±0.013,respectively.2.Study on extraction technology of high-temperature denatured soybean meal glycopeptideThe effects of extraction time,the ratio of raw material to water and extraction temperature on yield of high-temperature soybean meal crude glycopeptide were researched.The optimum extract process of high-temperature soybean meal glycopeptid was determined through the single factor experiments and response surface optimization experiments.The results showed that the ratio of raw material to water was 1:22 g/mL,extraction time was 2.8 h and extraction temperature was54.8?,the yield of glycopeptide was 14.71±0.31%.3.Study on isolation and purification technology of high-temperature denatured soybean meal glycopeptideFirstly,HSG-1 semi-pure product,HSG-2 semi-pure product and HSG-3 semi-pure product were obtain after the high-temperature soybean meal crude glycopeptide were eluted by DEAE-52 ion exchange chromatography.Then glycopeptide HSG-2 was obtained after HSG-2 semi-pure product was purified by Sephadex G-50 gel chromatography,the peptide and sugar content of that were 23.86±0.31% and 73.28±0.29%,respectively.4.Characteristic evaluation and structure characterization of high-temperature denatured soybean meal glycopeptide HSG-2SDS-PAGE electrophoresis detection showed that the molecular weight of high-temperature denatured soybean meal glycopeptide HSG-2 was 12149 Da.The rheological analysis shows that HSG-2 solution is a typical non-Newtonian fluid.The differential scanning calorimetry of HSG-2showed that the thermal denaturation temperature and denaturation enthalpy value were 162.30?and 328.50 J/g,respectively.High performance liquid chromatography analysis of the monosaccharides showed that the monosaccharide of the HSG-2 were consisted of mannose,ribose,rhamnose,glucuronic acid,galacturonic acid,glucose,galactose,xylose,arabinose and fucose,simultaneously,the mass of each monosaccharide was 9024.76,209.96,655.22,524.27,612.77,197055.71,29525.82,636.27,5108.70 and 506.38 mg/kg,respectively.?-elimination reaction analysis showed that HSG-2 contained O-glycopeptide bond.Infrared spectrum analysis showed that HSG-2 had both the characteristic absorption peak of polysaccharide and the characteristic absorption peak of polypeptide,and the type of glycosidic bond was ? type.Amino acid composition analysis showed that HSG-2 contained 17 kinds of amino acids,of which the three most abundant amino acids were glutamic acid,aspartic acid and arginine,and the contents were8.34 g/100 g,4.19 g/100 g and 2.94 g/100 g respectively.Circular dichroism analysis and secondary structure showed that the ?-helix accounted for 0.3% of the secondary structure,?-sheet accounted for 25.4%,?-folds accounted for 23.1%,and random folding accounted for 51.2%.5.Antioxidant activity evaluation of high-temperature denatured soybean meal glycopeptide HSG-2Antioxidant experiments in vitro showed that HSG-2 had some scavenging capacity for DPPH radical,hydroxyl radical and superoxide anion radical,and it also had good iron reduction ability. |
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