Study On The Relationship Between Molecular Structure And Function Of Soy 7S And 11S Globulins In Reverse Micelles Extraction Process With Chaotropic Agents

As an excellent plant protein resource,soybean has the advantages of wide cultivation,high yield,low price and high protein.The types of amino acids of soy protein are similar to those of milk,and they belong to the same high-quality protein as egg protein,and have good functional properties,so they are widely used in the food industry.As the main components of soy protein,7S and 11S globulins play a decisive role in the structure,physicochemical properties and functional properties of soy protein.The reverse micelles extraction method is a new technology suitable for the separation and purification of biological macromolecules.It has broad application prospects in the extraction of plant proteins.However,due to its low extraction rate,reverse micelles cannot be used in actual industrial production.In order to increase the protein extraction rate of reverse micelles and improve the functional properties of soy protein,this paper uses reverse micelles with chaotropic agents method to extract soybean 7S and 11S globulins,and further improves the reverse micelles through single factor experiment and response surface optimization test design.The protein extraction rate of the bundle,and the differences in the structure,physicochemical properties and functional properties of soybean 7S and 11S globulins extracted by the reverse micelles method and the isoelectric precipitation method with chaotropic agents are compared.The main research results are as follows:（1）Sucrose and sucralose were selected as the chaotropic agents for extracting soybean7S and 11S globulins before reverse micelles.The results showed that when the concentration of sucrose was 0.05 mol/L,the forward extraction rate of 7S globulin was increased from37.19%to 71.82%;when the concentration of sucrose was 0.01 mol/L,the forward extraction rate of 11S globulin was increased from 37.19%to 73.61%.Through the response surface optimization test design,the optimal extraction process for extracting 7S globulin from reverse micelles with chaotropic agents was sucrose concentration 0.04 mol/L,W₀22,pH value 8.7,electrolyte concentration 0.2 mol/L,extraction temperature 55℃,extraction time50 min,sample addition amount is 1.5 g/L conditions,the forward extraction rate of 7S globulin can reach up to 83.21%;Under the condition of electrolyte concentration 0.8 mol/L,pH 12,extraction temperature 55℃and 20 min extraction time,the backward extraction rate of 7S globulin can reach up to 90.5%.Through response surface optimization test design,the optimal extraction process for extra（2）The structural properties of 7S and 11S globulins extracted by the reverse micelles method and the isoelectric point precipitation method with a solvent are significantly different.The results of SDS-PAGE gel electrophoresis showed that the purity of 7S and 11S globulins extracted by isoelectric point precipitation method were 92.01%and 93.27%,respectively,and the purity of 7S and 11S globulin extracted by reverse micelles containing the solvent was96.17%and 96.81%,respectively.Amino acid composition analysis showed that the 7S and11S globulins obtained by the two extraction methods contained 17 amino acids,but the PER₃value of the 7S globulin extracted from the reverse micelles with chaotropic agents was greater than 2,which indicated that the reverse micelles with chaotropic agents had improve the quality of protein.X-ray diffraction analysis results showd that the protein skeleton structure extracted by the two methods is almost the same,but the reverse micelles with chaotropic agents could modify the amorphous structure of 7S and 11S globulins.Fourier infrared spectroscopy analysis showed that the absorption peak intensity of 7S and 11S globulin extracted by isoelectric point precipitation method was higher than that of 7S and11S globulin extracted by reverse micelles with chaotropic agents.Compared with isoelectric point precipitation method,it contained Theβ-sheet content of 7S globulin prepared reverse micelles by chaotropic agents is significantly less,but the content ofβ-turn structure,random coils andα-helices is more;the 11S globulin prepared reverse micelles by chaotropic agents,the content ofβ-sheet,random coil andα-helix is lower than that of 11S globulin prepared by isoelectric point precipitation method,but the content ofβ-turn has increased.Scanning electron microscopy results show that the 7S globulin extracted by isoelectric point precipitation method is scattered and spongy,and the protein surface has a large number of discontinuous holes and a small number of spherical structures;the 7S globulin extracted by the reverse micelles by chaotropic agents is mainly composed of flakes.The 11S globulin extracted by isoelectric point precipitation method has a compact sponge-like structure,and the protein surface is rough and has pores.And a large number of spherical structures;11S globulin extracted from reverse micelles with chaotropic agents was composed of rod-shaped and sheet-shaped structures,and the protein surface is smooth,without pores and spherical structures.（3）Study the surface hydrophobicity,Zeta potential,particle size distribution,thermodynamic properties of the four proteins.Surface hydrophobicity analysis showed that the H₀of the 7S and 11S globulins extracted by isoelectric point were 145.7 and 154.86,respectively;the H₀of the 7S and 11S globulins extracted by the reverse micelles with chaotropic agents were 59.7 and 96.79,respectively;the H₀of the 7S and 11S globulins extracted with chaotropic agents was 59.7 and 96.79,respectively.Zeta potential analysis showed that the absolute value of Zeta potential of 7S and 11S globulin extracted by isoelectric point precipitation method was lower than that of reverse micelles method with chaotropic agents,which indicated that the surface of 7S and 11S globulin extracted by isoelectric point precipitation method.The number of charges is less than that of the reverse micelles method with chaotropic agents,and the degree of molecular structure expansion is more significant.The results of particle size distribution analysis showed that the average particle diameters of 7S and 11S globulins extracted by isoelectric precipitation method were131.3 and 81.79 nm,respectively;the average particle diameters of 7S and 11S globulins extracted reverse micelles with chaotropic agents were 47.41 and 64.6 nm,respectively.Differential scanning calorimetry analysis showed that the denaturation temperature and denaturation enthalpy of 7S and 11S globulin obtained by isoelectric point precipitation method were 70.50℃,8.97 J/g and 89.01℃,17.68 J/g,respectively The denaturation temperatures of the 7S and 11S globulins obtained by the beam method are 92.14℃,0.84 J/g and 101.87℃,1.734 J/g,respectively.The analysis of flavor components showed that 218and 263 compounds were detected in the 7S and 11S globulins extracted from the reverse micelles with chaotropic agents.Compared with the isoelectric point precipitation method,the7S and 11S globules extracted from the reverse micelles with chaotropic agents were detected.The protein content of aldehydes,esters and alcohols is lower,and the content of hydrocarbons and ketones is higher.（4）Comparative study on the nitrogen solubility index,emulsification and stability,foamability and stability,water absorption and oil absorption of 7S and 11S globulin extracted by the reverse micelle with chaotropic agents and the isoelectric point precipitation method.The difference.At different pH,the solubility,emulsification,foaming and foam stability of7S and 11S globulins extracted by reverse micelles with chaotropic agents were significantly higher than those extracted by isoelectric point precipitation method;in different Na Cl At different concentrations,the nitrogen solubility index,emulsification and stability,foaming and stability of the 7S and 11S globulin extracted by reverse micelles with chaotropic agents showed different changes.The water absorption and oil absorption of the 7S globulin extracted by the isoelectric point precipitation method and the reverse micelles with chaotropic agents were 0.73 g/g,1.18 g/g and 2.16 g/g,4.03 g/g,respectively.The water absorption and oil absorption of 11S globulin extracted by the isoelectric point precipitation method and the reverse micelles with chaotropic agents were divided into 2.11 g/g,1.91 g/g,2.80 g/g,4.78 g/g.The water and oil absorption capacity of 7S and 11S globulins was significantly higher than that of 7S and 11S globulins extracted by isoelectric point precipitation method.