Preparation method of soybean protein compound with high emulsibility and high oxidability

Abstract

The invention belongs to the technical field of food processing, and provides a preparation method of a high-emulsibility and high-oxidability soybean protein compound. The preparation method mainly comprises the following steps: respectively dissolving soybean protein and a flavonoid compound, mixing, adjusting the pH value, dialyzing, refrigerating, freezing, drying and the like to finally obtain flavonoid compound-soybean protein compound powder. The method is simple in operation process and relatively low in requirements on instruments and equipment, and the obtained compound has relatively high oxidation resistance and emulsibility.

Description

Technical Field

[0001] The invention belongs to the technical field of food processing, and particularly relates to a method for preparing a highly emulsifiable and highly oxidizable soybean protein complex. Background Art

[0002] In recent years, the safety and environmental issues of animal products have attracted the attention of consumers, leading to the increasing popularity of plant protein. Plant protein is protein extracted from plant foods, such as whole grains, beans and nuts. Plant protein usually lacks one or two essential amino acids and is therefore considered an "incomplete protein". Compared with other plant proteins, soy protein has a higher protein quality and is considered an important food source to meet the protein needs of the human body. Soybeans (legumes) contain a high protein content of about 40%. However, the problem of soy protein's easy oxidation and aggregation seriously reduces the functional properties of the protein.

[0003] Flavonoids are a type of polyphenolic compound that is widely found in herbs, vegetables, fruits and other plants. They have multiple physiological functions such as anti-inflammatory, anti-cancer and anti-oxidation, as well as reducing the risk of Alzheimer's disease. With the increasing demand for physical health, composite functional foods have gradually come into the public's view, and the interaction between bioactive flavonoids and dietary proteins has also attracted the attention of some experts and scholars. Some people have studied the covalent interaction between zein and three polyphenols: chlorogenic acid, gallic acid and caffeic acid. The results showed that the covalent binding of the three polyphenols significantly increased the solubility of the protein-polyphenol complex. Others have studied the effect of the covalent interaction between ECGG and WPI on the functional properties of WPI. The results showed that the addition of ECGG can reduce the interfacial adsorption capacity and surface hydrophobicity of whey protein and change the secondary structure of the protein, thereby enhancing the emulsification and interfacial properties.

[0004] Therefore, there is an urgent need for a method for modifying the functional properties of soybean protein using flavonoid compounds. Summary of the invention

[0005] In order to solve the above technical problems, the present invention proposes a method for preparing a highly emulsifiable and highly oxidizable soybean protein complex.

[0006] The technical solution of the present invention is:

[0007] A method for preparing a highly emulsifiable and highly oxidizable soybean protein complex mainly comprises the following steps:

[0008] (1) dissolving soybean protein in distilled water and stirring to obtain solution I;

[0009] (2) adding the flavonoid compound to distilled water, heating and stirring to obtain solution II;

[0010] (3) After solution II is cooled to room temperature, solution I and solution II are mixed together, stirred evenly, and the pH is adjusted to obtain a soybean protein complex solution. After dialyzing, refrigeration, freezing, and drying, a flavonoid compound-soy protein complex powder is obtained.

[0011] Preferably, the flavonoid compound is selected from any one of luteolin, quercetin and rutin.

[0012] Preferably, the concentration of soybean protein in solution I is 1.5-2.5 g / 100 mL; the solution is stirred at 500 r / min for 1.5-2.5 h.

[0013] Preferably, the concentration of flavonoids in solution II is 0.1-0.3 g / 100 mL; the reaction conditions are heating at 80-98° C. and stirring at 500 r / min for 10-20 min.

[0014] Preferably, in (3), solution I and solution II are mixed in equal volumes.

[0015] Preferably, in (3), stirring is performed at 500 r / min for 25 to 35 min, and the pH is adjusted to 7.

[0016] Preferably, the dialyzate in (3) is performed for 48 hours, with the water being changed every 6 hours.

[0017] Preferably, the mixture in (3) is refrigerated at 4° C. for 10 to 15 h and freeze-dried for 3 days.

[0018] Compared with the prior art, the present invention has the following advantages and effects:

[0019] (1) Easy to operate, only heating and stirring are required during the process;

[0020] (2) The requirements for instruments and equipment are relatively low, and the freeze-dried sample powder is easy to test in subsequent experiments;

[0021] (3) The composites have high antioxidant and emulsifying properties. The antioxidant activity of the SPI-Ru composite is as high as 8.685 μmol / mL, the absolute value of the potential is 29.57 mV, and the average particle size is 23.60 μm. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 The foaming and emulsifying properties of the composite in Example 6;

[0023] Figure 2 is the particle size and potential of the composite in Example 6;

[0024] Figure 3 is the total antioxidant capacity of the complex in Example 6;

[0025] Figure 4 is the fluorescence intensity of the complex in Example 6;

[0026] Figure 5 The effect of Lu, Qu and Ru on the interfacial tension of SPI in Example 6;

[0027] Figure 6 is the Fourier transform infrared spectrum of the soy protein complex with different flavonoid compounds in Example 6;

[0028] Figure 7 Flow chart for the preparation of soy protein complex. DETAILED DESCRIPTION

[0029] In order to enable those skilled in the art to better understand the present invention, the present invention is further described in conjunction with specific implementation methods.

[0030] 1. Experimental materials and equipment

[0031] 1.1 Experimental Materials

[0032] Table 1 Experimental materials

[0033]

[0034]

[0035] 1.2 Instruments and Equipment

[0036] Table 2 Main instruments and equipment

[0037]

[0038] Example 1 Preparation of flavonoid compound-soy protein complex

[0039] A method for preparing a highly emulsifiable and highly oxidizable soybean protein complex mainly comprises the following steps:

[0040] (1) Accurately weigh 2 g of (soy protein) SPI, dissolve it in 100 mL of distilled water, and stir it with a magnetic stirrer at 500 r / min for 2 h to obtain solution I, i.e., the soybean protein solution;

[0041] (2) 0.2 g of luteolin was added to 100 mL of distilled water, heated at 95 °C and stirred at 500 r / min for 15 min to obtain solution II, i.e., flavonoid solution;

[0042] (3) After the flavonoid solution was cooled to room temperature, solution I and solution II were mixed together in equal volumes, stirred at 500 r / min for 30 min to mix evenly, and the pH was adjusted to 7 to allow the soy protein and flavonoids to fully combine, thereby obtaining a soy protein complex solution containing 1% SPI and 0.1% flavonoids, and dialyzed for 48 h, with the water changed every 6 h. After the dialysis, the soy protein complex was placed in a refrigerator at 4°C for 12 h, and then placed in a freeze dryer for freeze drying for 3 days to obtain a flavonoid-soy protein complex powder (soy protein-luteolin, SPI-Lu).

[0043] Example 2 Preparation of flavonoid compound-soy protein complex

[0044] A method for preparing a highly emulsifiable and highly oxidizable soybean protein complex mainly comprises the following steps:

[0045] (1) Accurately weigh 1.5 g of (soy protein) SPI, dissolve it in 100 mL of distilled water, and stir it with a magnetic stirrer at 500 r / min for 1.5 h to obtain solution I, i.e., the soybean protein solution;

[0046] (2) 0.1 g of luteolin was added to 100 mL of distilled water, heated at 80 °C and stirred at 500 r / min for 10 min to obtain solution II, i.e., a flavonoid solution;

[0047] (3) After the flavonoid solution was cooled to room temperature, solution I and solution II were mixed together in equal volumes, stirred at 500 r / min for 25 min to mix evenly, and the pH was adjusted to 7 to allow the soy protein and flavonoids to fully combine, thereby obtaining a soy protein complex solution containing 0.75% SPI and 0.05% flavonoids, and dialyzed for 48 h, with the water changed every 6 h. After the dialysis, the soy protein complex was placed in a refrigerator at 4°C for 10 h, and then placed in a freeze dryer for freeze drying for 3 days to obtain a flavonoid-soy protein complex powder (soy protein-luteolin, SPI-Lu).

[0048] Example 3 Preparation of flavonoid compound-soy protein complex

[0049] A method for preparing a highly emulsifiable and highly oxidizable soybean protein complex mainly comprises the following steps:

[0050] (1) Accurately weigh 2.5 g of (soy protein) SPI, dissolve it in 100 mL of distilled water, and stir it with a magnetic stirrer at 500 r / min for 2.5 h to obtain solution I, i.e., the soybean protein solution;

[0051] (2) 0.3 g of luteolin was added to 100 mL of distilled water, heated at 98 °C and stirred at 500 r / min for 20 min to obtain solution II, i.e., flavonoid solution;

[0052] (3) After the flavonoid solution was cooled to room temperature, solution I and solution II were mixed together in equal volumes, stirred at 500 r / min for 35 min to mix evenly, and the pH was adjusted to 7 to allow the soy protein and flavonoids to fully combine, thereby obtaining a soy protein complex solution containing 1.25% SPI and 0.15% flavonoids, and dialyzed for 48 h, with the water changed every 6 h. After the dialysis, the soy protein complex was placed in a refrigerator at 4°C for 15 h, and then placed in a freeze dryer for freeze drying for 3 days to obtain a flavonoid-soy protein complex powder (soy protein-luteolin, SPI-Lu).

[0053] Example 4 Preparation of flavonoid compound-soy protein complex

[0054] Different from Example 1, in this example, 0.2 g of quercetin was added to 100 mL of distilled water, heated at 95° C. and stirred for 15 min to obtain Solution II (soy protein-quercetin, SPI-Qu).

[0055] Example 5 Preparation of flavonoid compound-soy protein complex

[0056] Different from Example 1, in this example, 0.2 g of rutin was added into 100 mL of distilled water, heated at 95° C. and stirred for 15 min to obtain solution II (soy protein-rutin, SPI-Ru).

[0057] Example 6 Experimental Analysis

[0058] The control group experiment differed from the steps in Examples 1 to 5 in that no flavonoid reagent was used to compound the soybean protein, and an equal volume of distilled water was used to adjust the pH, thereby finally obtaining a soybean protein solution (SPI).

[0059] Origin 2022 software was used for drawing, SPSS software was used for data analysis (P<0.05), and uppercase and lowercase letters were used as labels to correspond to differences within and between groups. The experiments were conducted in triplicate, and the average value of the experimental data was selected for data analysis.

[0060] 6.1 Foaming and emulsifying properties

[0061] (1) Foaming characteristics

[0062] The flavonoid compound-soy protein complex (SPI-Lu, SPI-Qu, SPI-Ru, SPI) obtained in Example 1, Example 4, and Example 5 was dispersed at a speed of 10000 rpm for 2 min using a high-speed disperser. After the dispersion was completed, the generated foam was immediately transferred to a 100 mL graduated cylinder, and the initial volume of the foam was recorded. The foaming capacity was then calculated, and the foam volume of the complex solution was recorded again with a 100 mL graduated cylinder after 10 min. The calculation formulas for foaming capacity (FA) and foam stability (FS) are as follows:

[0063]

[0064] V 0 : Foam volume of flavonoid-soy protein solution at 0 min;

[0065] V 10 : Foam volume of flavonoid-soy protein solution after standing for 10 min.

[0066] Table 3 Foaming ability and foam stability of different flavonoids-soy protein complexes

[0067]

[0068] The foaming property and foam stability of soy protein are closely related to its own spatial structure and are easily affected by external factors such as temperature and pH. In addition, the combination with other ingredients usually causes changes in foaming property and foam stability.

[0069] Depend on Figure 1 (Left) Combined with Table 3, it can be seen that the addition of flavonoids significantly improved the foaming property of soy protein, among which SPI-Ru>SPI-Qu>SPI-Lu>SPI, while the change trend of foam stability was the opposite. This may be because Ru is more likely to change the surface properties of soy protein than other flavonoids, making it difficult to form a stable membrane structure, thereby causing a decrease in foam stability.

[0070] The reason for the change of the foaming properties of soy protein by flavonoids may be that flavonoids bind to proteins through non-covalent interactions (hydrogen bonds or hydrophobic interactions), resulting in reduced hydrophobicity and surface tension on the protein surface, which promotes the formation of bubbles and oil-water equilibrium interfaces. Reduced hydrophobicity on the protein surface will enhance its foaming ability. In addition, the foaming property of protein is related to its secondary and tertiary structures. The addition of flavonoids can change the structure of protein, making it looser and promoting the formation of foam. In the flavonoid-soy protein composite system, the structures of flavonoids are different, and there are certain differences in the changes in the interaction force, secondary structure, hydrophobicity, etc. between proteins, and the effects on the functional properties of soy protein are also different. The reduction in foam stability after adding flavonoids may be due to the fact that the hydrophobic interaction of the system is weaker than the electrostatic repulsion, and the gas leakage cannot be completely blocked by the formed film. The foam gradually becomes larger and then breaks, and the foam stability is reduced.

[0071] (2) Emulsification properties

[0072] The method for determining the emulsification activity and stability is as follows: 4 mL of soybean oil is added to a certain concentration of flavonoid compound solution, and a high-speed disperser is used to disperse the solution at 10,000 rpm for 60 seconds. Immediately after the dispersion, 10 μL of fresh emulsion is taken from about 0.5 cm above the bottom of the beaker, and the solution is dropped into 9.9 mL of SDS solution with a mass concentration of 0.1%, and the solution is diluted 1,000 times. After the emulsion and SDS solution are vortexed and mixed, the absorbance at 500 nm is recorded using an enzyme reader and recorded as A. 0 After the emulsion has been allowed to stand for 10 minutes, repeat the absorbance measurement steps above and record the absorbance at this time as A 10 .

[0073] The calculation formulas for emulsifying activity (EAI) and emulsifying stability (ESI) are as follows:

[0074] EAI(m 2 / g)={(2×2.303) / [C×(1-φ)×10 4 ]}×A 500 ×D F

[0075] ESI(min)=A 0 ×Δt / A 0 -A 10

[0076] EAI: emulsifying activity, m 2 / g; ESI: emulsion stability, min; D F: dilution factor, taken as 1000; C: concentration of the complex in the emulsion, 1.00 g / 100 mL; φ: volume fraction of the oil phase used to prepare the emulsion.

[0077] Table 4 Emulsifying activity and emulsifying stability of different flavonoids-soy protein complexes

[0078]

[0079] like Figure 1 As shown in (right), flavonoids significantly improved the emulsification activity and emulsification stability of soy protein. The reason may be that the addition of flavonoids caused changes in the electrostatic charge on the surface of soy protein, reduced its hydrophobicity, and improved the balance of the oil-water interface.

[0080] The effect of different flavonoids on the emulsifying properties of protein is related to the structural changes of protein. When the protein structure changes, the emulsifying properties also change. The combination of soy protein and flavonoids can effectively improve the functional properties of protein. The combined flavonoid-soy protein complex has good emulsifying properties. At the same time, the balance between the hydrophilicity and lipophilicity of protein makes the emulsion system more stable. The combination of hydrophilic flavonoids and proteins can increase the irregular curling of proteins, adjust the balance of hydrophilic-hydrophobic groups on the surface of proteins, optimize adsorption behavior, and inhibit their self-aggregation tendency, thereby improving emulsifying activity. Different flavonoids have different structures. The combination with proteins can change the structure of proteins and thus enhance the emulsifying properties of proteins.

[0081] 6.2 Particle size and zeta potential

[0082] The flavonoid compound-SPI complex was dissolved in water to form a complex solution, and the particle size distribution of the emulsion was measured using a laser particle size distribution analyzer. The average diameter was used to represent the particle size. After the complex solution was diluted to a concentration of (0.1 mg / mL), the potential was measured using a British Malvern zeta potential meter.

[0083] Table 5 Average potential and particle size of different flavonoids-soy protein complexes

[0084]

[0085] Particle size and potential are key indicators for evaluating the stability of soybean protein emulsions compounded with flavonoids. Figure 2 The left combined table 5 shows that compared with soy protein, the addition of flavonoids significantly reduced the emulsion particle size, and the trend was SPI-Ru<SPI-Lu<SPI-Qu<SPI, among which the SPI in the SPI-Ru group was reduced by 27.63% compared with the control group.

[0086] Figure 2 Right, among all the emulsion samples, the absolute value of Zeta potential of the emulsion without flavonoids was the lowest, only 19.57mV. After the flavonoids were added, the absolute value of Zeta potential increased significantly, and the SPI-Ru group reached 29.57mV.

[0087] The possible reasons are: (1) the negative charge of soy protein increases with the combination of flavonoids; (2) the addition of flavonoids causes changes in the structure of soy protein, which increases the electrostatic repulsion and increases the absolute value of the emulsion Zeta potential.

[0088] The average particle size of the soy protein emulsion decreased after adding flavonoids, which indicates that soy protein can produce a stable adsorption effect at the oil-water interface, which also improves the stability of the soy protein emulsion. The reason for the increase in the absolute value of the Zeta potential may be that the phenolic hydroxyl groups in the flavonoids undergo a deprotonation reaction to produce a large amount of negative charges. The combination of protein and flavonoids can also introduce negatively charged groups. These reasons increase the negative charge of the protein emulsion and increase the absolute value of the Zeta potential. The increase in the absolute value of the Zeta potential of the emulsion is consistent with the decrease in the average particle size of the droplets. This is because the surface charge of the protein droplets increases, and the droplets are not easy to aggregate, which is conducive to the reduction of the average particle size of the emulsion and the improvement of the stability of the emulsion, thereby forming an emulsion gel with a dense structure.

[0089] 6.3 Total antioxidant capacity

[0090] The total antioxidant capacity (T-AOC) of flavonoid-protein complex was determined by FRAP method. 0.1 g of freeze-dried flavonoid sample was added to 100 mL of distilled water and mixed thoroughly to obtain flavonoid-SPI sample solution. 1% SPI solution was used as the control sample solution of flavonoid-soy protein. Then, 5 μL of sample solution was added to 170 μL of FRAP reagent in the color developing solution in a 96-well plate, and then 15 μL of distilled water was added, and the mixture was repeatedly aspirated three times to ensure uniform mixing. FRAP reagent was prepared according to reagent I: II: III = 10:1:1, and it needs to be preheated at 37°C before use.

[0091] Reagent I: 1.5 mL, pH 3.6, 0.3 M acetate buffer solution;

[0092] Reagent II: 1.5 mL 10 mM TPTZ solution;

[0093] Reagent III: 1.5 mL 20 mM FeCl 3 Solution.

[0094] After mixing the three solutions of the solution sample, the chromogenic solution, and distilled water, react for 10 min at room temperature of 25 °C. Immediately after the reaction, measure its absorbance A at 590 nm using an enzyme-linked immunosorbent assay (ELISA) reader.

[0095]

[0096] V 1 : Sample volume in the reaction, 5 μL = 0.005 mL

[0097] D: Dilution factor, which is 1 if not diluted;

[0098] ΔA = A measurement - A blank;

[0099] Table 6 Total antioxidant capacity of different flavonoid-soy protein complexes

[0100]

[0101] As shown in Table 6, Figure 3 the addition of flavonoids significantly improved the antioxidant activity of soy protein. This may be due to the antioxidant properties of flavonoids themselves, and the stronger binding of Ru to the protein, resulting in the strongest total antioxidant activity in the SPI-Ru group. In addition, Ru and soy protein formed an interfacial film with better mechanical strength and toughness in the emulsion, which is another reason for the improvement of its antioxidant properties.

[0102] The improvement of the total antioxidant capacity of the complex is attributed to the large number of phenolic hydroxyl groups carried by flavonoids themselves. Adding flavonoids to soy protein introduced more phenolic hydroxyl groups, significantly enhancing the antioxidant properties of the complex. The improvement of antioxidant properties is also related to the interaction between the hydroxyl groups of flavonoids and protein groups, such as the carboxyl and hydroxyl groups of proteins. The formed complex has a unique structure, thereby enhancing the antioxidant properties.

[0103] 6.4 Fluorescence spectrum

[0104] Dilute the flavonoid-SPI sample to 0.2 mg / mL with distilled water. Measure the fluorescence spectrum using a fluorescence spectrophotometer and scan according to this method. Set the excitation wavelength to 295 nm (stimulating tryptophan fluorescence), and set the emission wavelength range to 300 - 500 nm. The scanning speed is 600 nm / min, and the slit width is 5 nm. Measure the fluorescence emission spectrum at 20 °C with distilled water as the blank.

[0105] As Figure 4As shown in the figure, by adding flavonoids, the fluorescence intensity of soy protein decreased, and the fluorescence intensity of the complex was SPI-Ru<SPI-Lu<SPI-Qu<SPI. Soy protein was quenched in a dose-dependent manner, which indicated that the two interacted with each other. The aromatic ring of flavonoids combined with the tyrosine and tryptophan residues of soy protein, and the hydrophilicity of protein increased and the hydrophobicity decreased. Protein has substances that emit intrinsic fluorescence, which is mainly due to the presence of tryptophan, tyrosine and phenylalanine residues. The addition of flavonoids caused the peak of the fluorescence spectrum to blue shift, and the wavelength moved to a shorter direction. The reason for the blue shift may be related to the interaction between the fluorescent molecules and water molecules, or to the change of the internal conformation of the fluorescent molecules. For example, water molecules bind to fluorescent molecules through hydrogen bonds, and the interaction between them will reduce the excited state lifetime of the fluorescent molecules, resulting in the blue shift of the fluorescence emission peak. In addition, the interaction between fluorescent molecules and other solvent molecules, such as van der Waals force and electrostatic interaction, may also cause the peak to blue shift. The peak blue shift may also be due to the change of the internal conformation of the fluorescent molecules, the change of the electron density, and the change of the fluorescence emission wavelength.

[0106] The reason for the decrease in the fluorescence intensity of the flavonoid-soy protein complex may be that the luminescent groups (leucine and tyrosine) on the protein side chain are exposed to a hydrophilic environment. The addition of different flavonoids to soy protein caused a significant decrease in the fluorescence intensity of the complex, indicating that the complex underwent fluorescence quenching. The addition of flavonoids caused the maximum fluorescence emission wavelength of the complex to blue shift. This may be caused by the interaction between flavonoids and soy protein. The polarity of the environment around the aromatic amino acid residues has weakened, the hydrophobicity of the environment around the luminescent group has become greater, and the structure of the soy protein has changed and become more compact.

[0107] 6.5 Interfacial tension

[0108] The dynamic interfacial tension of flavonoid-SPI complex was measured by hanging drop method. A certain amount of complex powder was dissolved in distilled water so that the concentration of the complex in the solution was 10 mg / mL. A stainless steel needle syringe with an inner diameter of 1.19 mm connected to an electric injection controller was inserted into a glass tank containing purified soybean oil. The syringe was adsorbed at the oil-water interface for 180 minutes. Before the experiment, each sample solution needed to be balanced in the syringe for 10 minutes to reach 25°C. Use an electric injection controller to drop the complex solution, with a volume of 14 μL per drop. The complex solution was slowly dropped into soybean oil, and the CCD camera system was used to continuously record the droplet image within 5 minutes, for a total of 25 times. The Young-Laplace equation was selected to calculate the interfacial tension:

[0109]

[0110]

[0111] γ: dynamic interfacial tension of the oil-water interface; g: gravitational acceleration; C: capillary constant; Δρ: density difference between the oil and water phases; X and Z: horizontal and vertical coordinates of any point on the outer contour of the droplet; θ: contact angle at any point; b: radius of curvature of point P in the droplet.

[0112] Figure 5 After adding flavonoids, the interfacial tension of soy protein gradually decreased. From the change curve of the interfacial tension of the flavonoid-soy protein complex, it can be seen that the change curve of the interfacial tension conforms to the change characteristics of the protein, the rate of decrease of the interfacial tension value of the complex gradually slows down, and the interfacial tension continues to decrease with increasing time. The interaction between flavonoids and soy protein usually causes the internal structure of the protein molecule to partially unfold, exposing the hydrophobic residues inside the protein molecule to the protein surface, so that the protein molecules are quickly adsorbed to the interface, reducing the interfacial tension, forming a stable and elastic interfacial film, and finally improving the emulsification of soy protein.

[0113] The interaction between soy protein and rutin can reduce the interfacial tension of the oil-water interface. This phenomenon is attributed to the conformational change of the formed complex, which reduces the surface hydrophobicity of soy protein and increases its hydrophilicity, increases the adsorption rate of the oil-water interface, makes it difficult for protein to aggregate, reduces the interfacial tension of the emulsion system, and improves the emulsification activity and stability.

[0114] 6.6 Infrared spectrum

[0115] Infrared test (tablet method): The freeze-dried powder of flavonoid compound-soy protein complex was mixed with potassium bromide powder and ground evenly, and pressed into transparent particles. The functional groups of the sample were analyzed using a Fourier transform infrared spectrometer. The wave number range of the infrared spectrum was 4000cm -1 ~400cm -1 The composite sample was scanned by transmission, 64 times, with a resolution of 4 cm -1 , and the infrared spectra of the samples were analyzed using peak fitting software.

[0116] Table 8 Secondary structures of soybean protein complexes with different flavonoids

[0117]

[0118] Note: Different lowercase letters in the same column of data indicate significant differences (P<0.05).

[0119] Figure 6Combined with Table 8, it shows that different structural regions of proteins produce different characteristic absorption bands, which can be analyzed to determine the secondary structure of proteins. The main components of this secondary structure are β-folding and α-helix, which is the most important aspect of protein structure. The wave number range of α-helix is ​​1645~1664cm -1 , β-folding is 1615~1664cm -1 and 1682~1700cm -1 , 1664~1681cm -1 Represents the β-turn, 1637-1645 cm -1 The β-sheet content decreased after adding flavonoids, while the α-helix content and random coil content increased. Amide A band (3200~3440cm -1 ) represents the NH stretching vibration. The characteristic absorption wave number of the amide I band is usually between 1600 and 1700 cm -1 The amide I band is the most important part of the secondary structure of proteins. The strongest absorption peak of SPI-Ru is at 1649 cm -1 At the position, it forms hydrogen bonds with adjacent chains through stretching vibration. Amide II band (1500~1600cm -1 ) is caused by NH bending vibration and CN stretching vibration, indicating the extent to which the NH group participates in hydrogen bonding with the adjacent α chain. The shift of amide bands I and II to lower wavenumbers is associated with the increase in hydrogen bonding between C=O and NH groups.

[0120] The amide I band and amide II band correspond to the bonding information of C=O and NH, respectively. The amide I band (1700-1600 cm -1 ) and amide A band (3100~3500cm -1 ) have characteristic peaks and change slightly, which may be due to the cross-linking of flavonoids and soy protein to form hydrogen bonds. The absorption peak of amide I band represents the stretching vibration of C=O, and the absorption peak of amide A band represents the stretching vibration of OH bond. The β-folding content decreases, while the α-helix content and random coil content increase. This may be because the addition of flavonoids affects the hydrogen bonds of soy protein, resulting in the breakage of polypeptide chains and changes in conformation. The changes in the secondary structure of soy protein indicate that soy protein is cross-linked with flavonoids, the conformation of soy protein is rearranged, and the orderliness of the molecule is significantly reduced.

[0121] In summary, the preparation method of the soybean protein complex provided by the present invention is to dissolve the soybean protein and the flavonoid compound separately, mix them, adjust the pH, and finally obtain the flavonoid compound-soy protein complex powder through the steps of dialysis, refrigeration, freezing and drying. The obtained soybean protein complex has high emulsification and high oxidation properties.

[0122] The above are only preferred embodiments of the present invention, and are not intended to limit the patent scope of the present invention. All equivalent changes and modifications made within the scope of the present invention should still fall within the scope of the present invention.

Claims

1. A method for preparing a highly emulsifiable and highly oxidizable soybean protein complex, characterized in that: The main steps include: (1) Dissolve soybean protein in distilled water and stir to obtain solution I; (2) adding the flavonoid compound to distilled water, heating and stirring to obtain solution II; (3) After solution II is cooled to room temperature, solution I and solution II are mixed together, stirred evenly, and the pH is adjusted to obtain a soybean protein complex solution. After dialysis, refrigeration, freezing, and drying, a flavonoid compound-soy protein complex powder is obtained.

2. The method for preparing a highly emulsifiable and highly oxidizable soybean protein complex according to claim 1, characterized in that: The flavonoid compound is selected from any one of luteolin, quercetin and rutin.

3. The method for preparing a highly emulsifiable and highly oxidizable soybean protein complex according to claim 1, characterized in that: The concentration of soy protein in the solution I is 1.5-2.5 g / 100 mL; the solution is stirred at 500 r / min for 1.5-2.5 h.

4. The method for preparing a highly emulsifiable and highly oxidizable soybean protein complex according to claim 1, characterized in that: The concentration of the flavonoid compound in the solution II is 0.1-0.3 g / 100 mL; the reaction conditions are heating at 80-98° C. and stirring at 500 r / min for 10-20 min.

5. The method for preparing a highly emulsifiable and highly oxidizable soybean protein complex according to claim 1, characterized in that: In the above (3), solution I and solution II are mixed in equal volumes.

6. The method for preparing a highly emulsifiable and highly oxidizable soybean protein complex according to claim 1, characterized in that: In the step (3), the mixture is stirred at 500 r / min for 25-35 min and the pH value is adjusted to 7.

7. The method for preparing a highly emulsifiable and highly oxidizable soybean protein complex according to claim 1, characterized in that: In the above (3), the dialysis was performed for 48 hours, and the water was changed every 6 hours.

8. The method for preparing a highly emulsifiable and highly oxidizable soybean protein complex according to claim 1, characterized in that: The mixture in (3) was refrigerated at 4°C for 10-15 hours and freeze-dried for 3 days.

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