The Study Of Functional Characterstics And Antioxidant Activity On Enzyme Hydrolyzed Protein And Glutelin In Naked Oats

Take naked oats as raw material, used alkali extraction and acid precipitation method and Osborne method to extract the protein and glutelin firstly. Then selected appropriate protease to hydrolyze naked oats protein and glutelin to get antioxidant peptide, which followed by the sduty of functional properties of their hydrolysates. Five different antioxidant assays in vitro had been used to investigate the antioxidant activity of naked oats protein and glutelin hydrolysates. Naked oats glutelin hydrolysates purified by ion exchange chromatography and reversed phase liquid chromatography (RP-HPLC). Mass spectrometry was taken to conduct molecular weight and primary sequence identification to antioxidant peptide which got from RP-HPLC. Got amino acid sequence of naked oats glutelin antioxidant peptides combined with the analysis of amino acid composition. It provided theoretical basis for the comprehensive utilization of naked oats. The main research results were as follow:Took naked oats protein and glutelin as substrate, used Alcalase, Trypsin and Protamex to hydrolyze, and the hydrolysis was carried out in their respective optimum conditions. With degree of hydrolysis (DH) and hydroxyl free radical scavenging rate as index to investigate the effect of enzymatic hydrolysis. Results showed that with the hydrolysis of Alcalase, both naked oats protein and glutelin reached higher value of DH and hydroxyl free radical scavenging rate. Single factor and orthogonal test were taken to get the optimum hydrolysis condition. Optimum conditions for naked oats protein were as follow:substrate concentration 10g/L, enzyme dosage 10%, pHvalue9, temperature 60℃, the DH reached up to 40% and hydroxyl free radical scavenging rate reached up to 50% with these conditions. Optimum conditions for naked oats glutelin were as follow: substrate concentration lOg/L, enzyme dosage7.5%, pHvalue 8.5, temperature 55℃, the DH reached close to 40% and hydroxyl free radical scavenging rate reached up to 60% with these conditions.Through the investigation of water-holding capacity, oil-holding capacity, solubility, foam ability and foam stability, emulsifying ability and emulsion stability, discuss the functional properties of naked oats protein, glutelin and their hydrolysates. The results showed that water-holding capacity and oil-holding capacity of hydrolysates were decreased. Naked oats protein hydrolysates and glutelin hydrolysates had good solubility in weak alkaline environment. Appropriate NaCl concentration solusions could increase foaming ability and foaming stability of naked oats protein, glutelin and their hydrolysates. However, decreased the emulsifying ability and increase emulsion stability. Foaming and emulsifying abilities could decrease with sucrose solutions, while foam and emulsion stability increase with sucrose solutions.The antioxidant activity of naked oats protein and glutelin hydrolysates were evaluated using five different antioxidant assays in vitro. The results showed there was a certain does relationship with antioxidant acitivity and hydrolysates concentration of naked oats protein and glutelin. Naked oats protein and glutelin hydrolysates were good hydrogen and electron donator, with strong scavenging activity against·OH, O2-·, DPPH·and H2O2 radicals, had good reducing power.Naked oats protein hydrolysates had been isolated into five fractions after purified by ion exchange chromatography, and found that basic component E had the highest antioxidant activity, IC501.79mg/mL and 1.54mg/mL respectively. Then glutelin hydrolysates had been isolated into four fractions, and basic component D had the highest antioxidant activity, IC501.53mg/mL and 1.28mg/mL respectively. The fraction D of glutelin hydrolysates further purified by semi-preparing RP-HPLC, and got two components,the component D-1 had the higher antioxidant activity. Analytical RP-HPLC was taken to purify component D-1, and got main peak D-1a, it had strong hydroxyl scavenging activity, which the IC50 was 0.68mg/mL. Identified by RP-HPLC, and confirmed it was a single peak. Free Radical scavenging capacity of naked oats glutelin hydrolysates improved greatly after purified by ion exchange chromatography and RP-HPLC chromatography.Amino acid composition analysis of D-1a showed that this antioxidant peptide contain seven amino acides. It included hydrophobic amino acid residues His and Pro, aromatic amino acids Tyr, non polar aliphatic amino acid Ala, which played an important role on its antioxidant activity. Identified by MALDI-TOF-TOF mass spectrometry, the molecular weight of antioxidant peptides D-1a was 785.38Da, and the amino acid sequence was HYNAPAL (His-Tyr-Asn-Ala-Pro-Ala-Leu).