| Walnut,as a homologous product of medicine and food,has a large planting area in China.However,its sales price is in a downward trend according to statistics over the years,while the development of walnut industry in China is still in the initial stage,and there are fewer applications in food.In this study,walnuts were used as raw materials to study the changes of nutrient components,antioxidant enzymes and antioxidant activities of walnuts at different germination time by soaking and germination at constant temperature.The appropriate germination time was selected.The extraction process of walnut protein was designed by alkali and buffer design response surface methodology.The DPPH free radical scavenging capacity,reducing power and ferrous ionization of walnut protein in different extraction methods were also studied.The results are as follows:1.During the germination of walnut seeds,the water content of walnut seeds increased with the prolongation of germination time.The contents of N,P and K increased with the prolongation of germination time.The content of soluble sugar and soluble protein increased in general,and the content of phenolic substances was the highest when untreated.During the germination of walnut seeds,the contents of protease,SOD and POD increased,while the contents of CAT increased first and then decreased.DPPH had the highest scavenging capacity and reducing power in 0 d,with the scavenging power of 32.28%and the absorbance value of reducing power of1.1397.The chelating ability of ferrous ions and SOD-like activity reached their peak at 15 days,96.54%and 174.09U/mg,respectively.The best germination time was 21days.2.In alkali extraction method,based on single factor results,15 groups of three-factor and three-level experiments were designed by Box-Behnken central combination.The results showed that the sequence of factors affecting the extraction process of walnut seed protein was:material-liquid ratio X3>extraction time X1>Na OH concentration X2.The extraction conditions were extraction time 1.25h,concentration 87.81m M and material-liquid ratio 1:62.6.The protein yield was adjusted by actual conditions.28.22%.The DPPH radical scavenging rate of walnut seed powder protein extracted by alkali ranged from 1.08 to 3.63%,the reducing power of seed protein extracted by alkali ranged from 0.212 to 0.328,and the chelating ability of ferrous ion of seed protein extracted by alkali ranged from 63.04to 89.19%.The extraction time X3>ratio of material to liquid X2>concentration of Na OH X1.The extraction conditions were Na OH concentration 83.64 m M,extraction time 9.14 min and ratio of material to liquid 1:13.80.The extraction yield of protein was 18.14%after adjusting the parameters.The DPPH radical scavenging rate of protein extracted from walnut germinating seeds by alkali extraction ranged from 0.02to 4.68%,and the protein reducing power was 0.02 to 4.68%.The range is0.218-0.364,and the chelating ability of protein ferrous ions is 98.59%-99.95%.3.Based on the single factor results,27 groups of four-factor and three-level experiments were designed by Box-Behnken central combination.The results showed that the sequence of factors affecting the extraction process of walnut seed protein was p H value X1>extraction time X4>solid-liquid ratio X3>ion concentration X2.The extraction conditions were p H 9.18,ion concentration 82.55m M liquid-liquid ratio 1:51.72 and extraction time 4.39h.The protein yield of parametric extraction was 28.30%.The DPPH radical scavenging rate of buffer solution for walnut seed protein extraction ranged from 3.94%to 10.98%.The protein reducing power ranged from 0.133%to 0.527,and the protein ferrous ion chelating ability ranged from 15.80%to 56.28%.The sequence of factors affecting the extraction process of walnut germinating seed protein was as follows:solid-liquid ratio X2>p H X3>ion concentration X1>extraction time X4.The extraction conditions of buffer solution were p H 9.98,ion concentration 72.20M liquid ratio 1:13.33,extraction time13.23min.The extraction yield of protein was 16.11%,DPPH radical scavenging rate was 1.14-10.06%,protein reducing power was 0.180-0.409,and protein ferrous ion chelating ability was 90.15%-98.59%.In summary,the utilization rate of storage materials and enzyme activity of walnut seeds and germinated seeds increased during germination.Then,the relationship between protein yield and extraction parameters was studied by alkali and buffer extraction methods,and the appropriate parameters and process of protein extraction were obtained.The antioxidant activities of extracts were compared and analyzed.The results showed that there was no significant difference in protein yield between buffer extraction and alkaline extraction,but the DPPH radical scavenging rate of protein components extracted by buffer extraction was significantly higher than that obtained by alkaline extraction,but the chelating ability of ferrous ions extracted by alkaline extraction was better than that extracted by buffer extraction.These results indicated that the establishment of extraction methods,parameters and processes guided by protein yield and antioxidant activity would be helpful to screen and isolate antioxidant proteins or polypeptides from walnut seeds,and provide a reference for the development and utilization of bioactive substances in walnut seeds,as well as for the development and utilization of other forest seed protein resources.Law and basis. |
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