Preparation By Enzymatic Hydrolysis, Microencapsulation And Bioactivity Assement Of The Antioxidant Peptides From Egg White Proteins

Egg white is a good source of high quality proteins for food nutrition and bioactive peptides.The peptides from hydrolyzed egg protein by one or several proteases have the antioxidant activity.Antioxidant peptide of egg white proteins?APEWP?plays a great role in food industry and medicine industry because of its nutrition and healthy function.Moreover,the allergenicity of egg white proteins will be mostly reduced because linear epitopes and conformational epitope will be destroied when egg white protein were hydrolyzed.To produce the antioxidant peptides from egg white proteins,the enzymes and the enzymatic hydrolysis conditions were optimized firstly.The peptide was chelated by four kinds of metal ions including(K+,Ca²⁺,Mg²⁺,Zn²⁺).Antioxidant peptides were microcapsuled by piercing-concreting method,followed by analyzing the controlled-release properties and antioxidant activity.Both IgE and IgG binding capacity and cell cytotoxicity of antioxidant peptides were detected.The protective effect against cell oxidative stress injury and transfer-absorption in vitro were evaluated in a Caco-2 oxidative stress damage cell model and Caco-2 monolayer cell model,respectively.The main methods,results and conclusions are presented as follows.1.Chymotrypsin combined with pepsin were selected as the optimal prtoteolytic enzymes for producing antioxidant pepetide from egg whitle based on the FRAP test of their hydrolysates.The hydrolysis condition was optimized by response surface methodology,and the optimized parameters of the hydrolysis condition were E1/E2 at 1.7:1,preheating time for 10 min,the volume ratio of material to water at 10 %,E/S at 0.4 % and hydrolysis time for 3.16 h.The FRAP value of hydrolysate predicted by response surface methodology model was 13.33 ?mol AAE/g DW.To verify the accuracy of the prediction,verification test was repeated for three times and the results was 13.46 ± 0.456 ?mol AAE/g DW which was in good agreement with the prediction value,suggesting the response surface model was suitable for optimization of enzymatic hydrolysis conditions.2.APEWP were preparaed through ultrafiltration with molecular weight cut off?MWCO?of 3000 u?Millipore?,and the FRAP value and ·OH radical scavenging activity of the APEWP were 30.86 ?mol AAE/g DW and 43.2 %,respectively,indicating that the APEWP has a good antioxidant activity.The peptides were chelated with K+,Ca²⁺,Zn²⁺ and Mg²⁺,respectively,and the FRAP value of metal-chelating peptides were significantly decreased and OH radical scavenging activity were not significantly different when compared to the APEWP.3.The APEWP was microcapsuled by piercing-concreting method,and the embedding rate was 80.17 %,the particle diameter was among 500-1000 ?m.The microcapsuled APEWP was slowly released in simulated gastric fluid and simulated intestinal fluid.The antioxidant activity of APEWP could be protected by microencapsulation.4.The IgG and IgE binding to APEWP tested by indirect competitive ELISA assay was decreased in compare with the heated egg white?95?,10 min?and its hydrolysate.The cell cytotoxicity of APEWP and digested APEWP was determinated by CCK-8 kits,and the APEWP was no cell cytotoxicity in the range of the concentration of 100-1600 ?g/mL,Regarding to the digested APEWP,the valibity was 88.80 % when the concentration was 800 ?g/m L,which is low than the reference value of 95%,indicating the existed cell cytotoxicity.5.The oxidative-stress damage induced by H2O2 in a Caco-2 cell model was established.It was shown that the APEWP could effectively protect Caco-2 cell from oxidative damage,and there was positive correlated within 100-800 ?g/m L of APEWP and the digested APEWP.Moreover,the Caco-2 monolayer cell model was established to test the Papp and transport rate of APEWP and its digestived products.The Papp values of APEWP and its digestived products were more than 1×10-5 cm/s,suggesting the transport of APEWP and the its digestive products in vitro were all well absorpted.Moreover,the transport of APEWP and its digestive products in 2 h were 6.92 % and 8.64 %,respectively.