| Juglans sigllata Dode is a kind of wild walnut with some few nuts quite difficult to take out.The kernel rate of Juglans sigllata Dode is only 25%~30%,and the oil rate could reach 70%.Walnut is rich in protein,fatty acids and various trace elements such as a variety of beneficial to human nutrition.Furthermore,wild walnutexhibites advances compored with ordinary walnuts,such as more natural,organic,pollution-free and including more nutrients.Wild walnut cake obtained after oil extraction was generally thrown away,which resulted in a waste of high quality protein.In order to improve the utilization of the Juglans sigllata Dode cake,the high purity protein and the soluble polypeptide from Juglans sigllata Dode were prepared.The process conditions were optimized by response surface methodology,and amino acid composition and molecular weight distribution of the products were detected.The main research contents and conclusions are listed as follows:(1)Optimization of extraction separation protein response surface method of cold pressed Juglans sigllata Dode: using the cold pressed walnut cake as raw material,the single factor based on single factor screening by Plackett-Burman test,response surface methodology and Box-Behnken.Optimize the separation process optimization of protein extraction by response surface method to be alkali extraction p H of 9.6 cold pressing wild walnut cake,the temperature of 51? alkali extraction and acid precipitation p H4.5.Under these parameters,the protein extraction rate was 75.25%.(2)Purification of walnut protein isolate glucoamylase: the optimal purification process was optimized on the basis of single factor and orthogonal test.The conditions were the enzymatic hydrolysis temperature of 45 ?,enzymolysis p H 4.5,enzymolysis time of 120 min,enzyme d osage 0.4%,and solid-liquid ratio 1:10.Under these conditions,the enzyme purified wild walnut protein purity was 95.79%.The purified walnut protein was rich in amino acid composition and the sum of amino acids was up to 98.07%.SDS-PAGE electrophoresis analysis showed that the most abundant relative molecular mass of Juglans sigllata Dode protein was 40.7k Da and 21.6k Da.(3)Preparation of wild walnut polypeptide by two-step enzymatic hydrolysis(neutral protease and alkaline protease): according to the results of single factor,the fixed parameters of the neutral protease were optimized by response surface method Central-Composite test Design considering the interaction of alkaline protease,then the conditions was optimized to be neutral protease p H7.0,the temperature 45?,the amount of enzyme 6000u/g.After the hydrolysis of 3h,the enzyme inactivation conditions were alkaline protease p H9.1,enzymolysis temperature 51?,the amount of enzyme 6000u/g,and hydrolysis time 3.7 h.The highest degree of hydrolysis of Juglans sigllata Dode polypeptide was about 34%.Amino acid composition analysis and molecular weight distribution exhibited that the essential amino acid content was high and the molecular weight distribution is mainly concentrated in 180~500Da.(4)Comparison of different degree of hydrolysis of Juglans sigllata Dode polypeptide functional properties: the increased degree of hydrolysis could improve the functional properties of Juglans sigllata Dode polypeptide,but the excessive hydrolysis would result in the lack of some physicochemical properties.The comparison of functional characteristics among Juglans sigllata Dode defatted protein powder,Juglans sigllata Dode isolated protein and Juglans sigllata Dode indicated that enzyme hydrolysis can improve the functional properties of protein,protein solubility,emulsifying and foaming;the solubility of the peptide was better and more stable in the acid or base environment. |
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