Study On Antioxidant Peptides From Walnut Meal By Solid-state Fermentation

To make full use of the protein resources in walnut meal, solid-state fermentation process was adopted for the preparation of walnut peptides(WPs) from walnut meaL In this paper, the fermentation process, the antioxidant activity of WPs, The molecular weight distribution and the amino acids of peptide were studied. Fermentation strain was selected, optimum solid-state fermentation conditions, as well as antioxidant activity of WPs, molecular weight distribution and amino acid composition of WPs were measured. The main conclusions are as followings:Selecting fermentation strainsFive strains including Aspergillus niger, Aspergillus oryzae, Bacillus subtilis, Bacillus licheniformis and Mucor were used for solid-state fermentation of walnut meal. According to the influence of different strains on peptide yield, Bacillus subtilis was proved to be the best fermentation strains.Effects of polyphenols in walnut meal on solid-state fermentationIt was found that the contents of polyphenols in walnut meal had great impact on the solid-state fermentation. After removing polyphenols from walnut the peptides yield improved15.2%. Nine polyphenols were identified from walnut meal by UPLC-ESI-MS, which were gallic acid, vanillic acid glucoside, digalloylglucose, rutin, cumaroylquinic acid, glansreginin B, glansreginin A, chlo ro genic acid, and ellagic acid. These polyphenols extracted from walnut meal can be used as raw material of drugs. Optimization of solid-state fermentation processBased on the single factor experiments, response surface experiments were adopted to optimize of solid fermentation process. The effects of three critical structuring components, namely fermentation time, inoculum size, and water content on the degree of hydrolysis (DH) and antioxidant activity (FRAP) were investigated. Experimental models were as follows:DH:Y1=-445.42+9.19X1+6.08X2+89.58X3-0.034X1X2-0.201X1X3-0.168X2X3-0.051X12-0.138X22-22.661X32FRAP:Y2=-2.178+0.064X1+0.018X2+0.478X3-0.0002X1X2-0.0003X1X3-0.001X,X3-0.0004X120.00015X22-0.1664X32The optimal fermentation conditions obtained by two equations were fermentation time82.01h. inoculum size10.40%and water content1.50mL/g. The models were significant regression (P<0.05) and the correlation coefficient were0.9639and0.8779for DH and FRAP. These suggested that there was a good correlation between variable and dependent variables. The error between actual value and the predicted value were only1.65%and2.6%by experimental verification, which indicated the models can successfully predict the experiments.Ultrafiltration and molecular weight(Mw) distribution of WPs After ultrafiltration, the WPs were divided into three parts. Mw<5000Da account for90.2%,5000Da<Mw<10000Da account for8.6%, while Mw>10000Da account for only1.2%of while peptides. The results of molecular weight distribution of WPs showed that the molecular weight of walnut protein decreased significantly compared with those not fermented. In parts Mw<5000Da, Mw<1500Da account for 91.49%. Amino acid analysis results showed that the content of Arg, Asp, Glu, Tyr and His in parts Mw<5000Da are relatively high than other parts, the component may have other components do not have biological activity. The special amino acid composition of Mw<5000Da may lead to the different biological activity.Determination of antioxidant activity of WPsAccording to the (Ferric ion reducing antioxidant power, FRAP) of different peptides, the part of Mw<5KDa showed highest FRAP activity. So it was selected as peptides for further determination. The antioxidant activity of peptides including Ferric ion reducing antioxidant power, DPPH radical scavenging activity, Hydroxyl radical scavenging activity, Superoxide anion radical scavenging activity and Fe2+ion-chelating activity. Vc and GSH were used as positive reference. The results showed that the antioxidant activity of WPs was much higher than GSH at same concentration while a little lower than Vc. Fe2+ion-chelating activity results showed that WPs was much lower than EDTA but higher than GSH, The same concentration of GSH has almost no Fe2+chelating activity. These results indicated that WPs have potential applications as food and pharmaceutical.