Enzyme cross-linked whey protein-quercetin compound as well as preparation method and application thereof

Abstract

The invention discloses an enzyme crosslinking whey protein-quercetin compound as well as a preparation method and application thereof, and belongs to the technical field of food processing. In order to solve the technical problem of poor technical effect of whey protein modification in the prior art, the invention provides a preparation method of an enzyme crosslinked whey protein-quercetin compound, and whey protein is modified through synergism of three different modification means of ultrasonic treatment, enzymatic crosslinking and polyphenol combination. According to the preparation method, ultrasonic treatment and enzymatic cross-linking have a synergistic effect, so that covalent interaction and combination of quercetin and protein are promoted, and the modification effect on whey protein is improved; the enzyme cross-linked whey protein-quercetin compound provided by the invention has relatively strong solubility, emulsification stability, oxidation resistance, uniformity and stability, and can be applied to the field of food processing.

Description

Technical Field

[0001] The invention belongs to the technical field of food processing, and particularly relates to an enzyme-crosslinked whey protein-quercetin complex and a preparation method and application thereof. Background Art

[0002] Whey protein is a protein present in whey after separation and precipitation of bovine milk. It is composed of a variety of substances, mainly including: β-lactoglobulin, α-lactalbumin, immunoglobulin and bovine serum albumin; protein products made from whey include: whey protein concentrate (protein content is 70%) and whey protein isolate (protein content is 90%); the above whey protein concentrate and whey protein isolate have good functional properties in food systems, such as: foaming, emulsification, gelling and other properties; among them, whey protein has good gel hardness, but still faces great challenges in application; for example: whey protein has poor water retention, is more sensitive to environmental conditions, is unstable, and has great limitations in food industry applications.

[0003] Flavonoids and polyphenols have excellent anti-inflammatory, anti-cancer and antibacterial effects. Quercetin, as a natural plant active substance widely found in fruits, vegetables and herbs, is widely used in food research.

[0004] The single chemical or physical treatment modification method in the prior art has a poor effect on improving the functional properties of whey protein, resulting in poor covalent or non-covalent interaction between polyphenols and proteins, making the secondary, tertiary and quaternary structures of the proteins unstable, thereby affecting the functional properties of the proteins after binding.

[0005] Therefore, those skilled in the art are eager to seek a modification technology to improve the functional properties of whey protein and expand its application range in the food industry, for example, as an excellent food ingredient. Summary of the invention

[0006] The invention aims to solve the technical problem that the whey protein modification technology in the prior art has a poor effect, and provides an enzyme-crosslinked whey protein-quercetin complex and a preparation method and application thereof.

[0007] One of the purposes of the present invention is to provide a method for preparing an enzyme-crosslinked whey protein-quercetin complex, the preparation method comprising the following steps:

[0008] S1: Dissolve whey protein in deionized water to prepare a whey protein solution, and adjust the pH value of the whey protein solution to 7.0 using a 0.2 mol / L NaOH solution; perform ultrasonic treatment on the whey protein solution in an ice water bath, with an ultrasonic power of 200 W, an ultrasonic time of 5 min, and an ultrasonic frequency of 25 Hz to obtain an ultrasonically treated whey protein solution;

[0009] S2: The whey protein solution after ultrasonic treatment in S1 is placed in a 50°C water bath for preheating for 15 minutes, and then horseradish peroxidase HRP, glucose oxidase and D-glucose are added, and water is added to mix and then enzyme cross-linking reaction is carried out. After the reaction, the solution is taken out and immediately inactivated, and cooled at room temperature to obtain whey protein after ultrasonic treatment and enzyme cross-linking reaction;

[0010] S3: adding quercetin solution to the whey protein obtained in S2 through ultrasonic treatment and enzyme cross-linking reaction, adjusting the pH value to 9.0, reacting with stirring at 25° C. for 24 h, dialyzing, and drying to obtain an enzyme-cross-linked whey protein-quercetin complex.

[0011] In a preferred embodiment of the present invention, the concentration of the whey protein solution obtained by the configuration described in S1 is 60 g / L.

[0012] In a preferred embodiment of the present invention, the whey protein concentration after the water mixing in S2 is 50 g / L, and the added amounts of horseradish peroxidase HRP, glucose oxidase and D-glucose are 200 U / g, 6 U / g and 0.05 mmol respectively per gram of whey protein.

[0013] In a preferred embodiment of the present invention, the enzyme cross-linking reaction conditions in S2 are: placing the enzyme cross-linking reaction at 37° C. for 3 hours.

[0014] In a preferred embodiment of the present invention, the enzyme inactivation treatment condition in S2 is: placing in a water bath at 80-150° C. for 5-20 min to inactivate the enzyme.

[0015] In a preferred embodiment of the present invention, the whey protein concentration obtained in the ultrasonic treatment-assisted enzyme cross-linking reaction in S2 is 1% (w / w).

[0016] In a preferred embodiment of the present invention, the mixing mass ratio of the whey protein to the quercetin solution in S3 is 20:1.

[0017] In a preferred embodiment of the present invention, the dialysis condition in S3 is: using a 3500Da dialysis bag for 48 hours at 4°C.

[0018] The second object of the present invention is to provide an enzyme-crosslinked whey protein-quercetin complex, which is obtained by the above-mentioned preparation method.

[0019] The third object of the present invention is to provide the application of the above enzyme-crosslinked whey protein-quercetin complex in food processing.

[0020] Beneficial effects of the present invention: The present invention provides a method for preparing an enzyme-crosslinked whey protein-quercetin complex, wherein the whey protein is modified synergistically by three different modification methods, namely, ultrasonic treatment, enzymatic crosslinking, and combination with polyphenols, to prepare the enzyme-crosslinked whey protein-quercetin complex.

[0021] The present invention uses glucose oxidase to catalyze the oxidation of D-glucose to produce H2O2 and gluconic acid through an enzymatic cross-linking reaction. Horseradish peroxidase can oxidize tyrosine residues in proteins in the presence of H2O2 to form dityrosine residues, resulting in covalent cross-linking of proteins. The main components of whey protein: α-lactalbumin and β-lactoglobulin are dense globular proteins, which prevent the reaction between the dual oxidase system and whey protein. Ultrasonic waves are an environmentally friendly technology that does not contain exogenous chemical additives. The cavitation implosion of ultrasonic waves can generate huge energy. The non-thermal process of mechanical waves and acoustic cavitation can quickly destroy the physical structure of proteins and expose the target area, thereby promoting enzymatic cross-linking reactions and improving the functional properties of proteins.

[0022] The protease reacts with the whey protein to form a protein-like reaction and cross-links, transferring amino acids from one peptide chain to another peptide chain to achieve modification of the peptide chain, thereby converting a small molecular weight peptide chain into a high molecular weight peptide chain and enhancing rheological properties. The present invention synergizes ultrasonic treatment and enzymatic cross-linking to improve the solubility, emulsification stability, antioxidant properties, uniformity and stability of the whey protein, causing polyphenols (quercetin) to bind to the protein through covalent interactions, causing the secondary, tertiary and quaternary structures of the protein to change, thereby promoting the modification effect on the whey protein, and significantly improving the solubility, emulsification stability, antioxidant properties, uniformity and stability of the enzyme-crosslinked whey protein-quercetin complex compared to a single modification method.

[0023] The enzyme-crosslinked whey protein-quercetin complex prepared by ultrasonic treatment, enzymatic crosslinking and polyphenol combination has good functional properties of whey protein, expands the application of whey protein in the food industry, and can be used as an excellent food ingredient in the field of food processing. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 It is the solubility statistics of WPI-Q, CWPI-Q and UCWPI-Q;

[0025] Figure 2 It is the statistical diagram of emulsion stability of WPI-Q, CWPI-Q and UCWPI-Q;

[0026] Figure 3 It is the statistical diagram of ferric iron reducing power of WPI-Q, CWPI-Q and UCWPI-Q;

[0027] Figure 4 It is the statistical graph of ABTS free radical scavenging rate of WPI-Q, CWPI-Q and UCWPI-Q;

[0028] Figure 5 It is the statistical diagram of free thiol content of WPI-Q, CWPI-Q and UCWPI-Q;

[0029] Figure 6 It is the particle size statistics of WPI-Q, CWPI-Q and UCWPI-Q;

[0030] Figure 7 Zeta potential statistics of WPI-Q, CWPI-Q and UCWPI-Q;

[0031] Figure 8 It is the solubility statistics of UCWPI and UCWPI-Q;

[0032] Fig. 9 It is the statistical diagram of ferric iron reducing power of UCWPI and UCWPI-Q;

[0033] Fig.10 The ABTS free radical scavenging rate statistics of UCWPI and UCWPI-Q;

[0034] Fig.11 Statistical chart of free thiol content of UCWPI and UCWPI-Q. DETAILED DESCRIPTION

[0035] Those skilled in the art can refer to the content of this article and appropriately improve the process parameters. It should be particularly noted that all similar replacements and modifications are obvious to those skilled in the art, and they are all considered to be included in the present invention. The method and application of the present invention have been described through preferred embodiments, and relevant personnel can obviously modify or appropriately change and combine the methods and applications described herein without departing from the content and scope of the present invention to implement and apply the technology of the present invention.

[0036] In order to make the purpose, technical scheme and advantages of the present invention more clear, the present invention is further described in detail below in conjunction with specific embodiments and the accompanying drawings of the specification. The test methods used in the following examples are conventional methods unless otherwise specified, and the materials, reagents, methods and instruments used are conventional materials, reagents, methods and instruments in the art unless otherwise specified, and can be obtained by those skilled in the art through commercial channels.

[0037] Example 1: A whey protein-quercetin complex with ultrasonic treatment and enzyme cross-linking reaction

[0038] S1: Dissolve whey protein in deionized water to prepare a whey protein solution with a concentration of 60 g / L, and adjust the pH value of the whey protein solution to 7.0 with a 0.2 mol / L NaOH solution; perform ultrasonic treatment on the whey protein solution in an ice water bath, with an ultrasonic power of 200 W, an ultrasonic time of 5 min, and an ultrasonic frequency of 25 Hz. After the ultrasonic treatment, obtain a whey protein solution after ultrasonic treatment, which is recorded as UWPI;

[0039] S2: The whey protein solution after ultrasonic treatment in S1 is placed in a 50°C water bath for preheating for 15 minutes, and then horseradish peroxidase HRP, glucose oxidase and D-glucose are added, and the mixture is placed at 37°C for enzyme cross-linking reaction for 3 hours after adding water. After the reaction, the mixture is taken out and immediately placed in an 85°C water bath for enzyme inactivation for 10 minutes, and cooled at room temperature. The whey protein concentration after the water mixing is 50g / L, and the horseradish peroxidase HRP, glucose oxidase and D-glucose are added in an amount of 200U / g, 6U / g and 0.05mmol per gram of whey protein, respectively; the whey protein subjected to ultrasonic treatment and enzyme cross-linking reaction is obtained, which is recorded as UCWPI;

[0040] S3: Adding quercetin solution to the whey protein obtained in S2 through ultrasonic treatment and enzyme cross-linking reaction, wherein the whey protein concentration is 1% (w / w), and the mixing mass ratio of the whey protein to the quercetin solution is 20:1; adjusting the pH value of the mixed solution to 9.0, stirring and reacting at 25°C for 24 hours, dialyzing at 4°C for 48 hours using a 3500Da dialysis bag, and drying to obtain a whey protein-quercetin complex through ultrasonic treatment and enzyme cross-linking reaction, recorded as UCWPI-Q.

[0041] Comparative Example 1: Whey protein-quercetin complex without ultrasonic treatment and enzyme cross-linking reaction

[0042] S1: Dissolve whey protein in deionized water to prepare a whey protein solution with a concentration of 60 g / L, and adjust the pH value of the whey protein solution to 7.0 with a 0.2 mol / L NaOH solution to obtain a whey protein solution, which is recorded as WPI;

[0043] S2: The 1% (w / w) whey protein solution obtained in S1 was mixed with the quercetin solution in a mass ratio of 20:1, the pH value was adjusted to 9.0, and the mixture was stirred at 25°C for 24 hours. The mixture was dialyzed at 4°C for 48 hours using a 3500Da dialysis bag and dried to obtain a whey protein-quercetin complex, which was recorded as WPI-Q.

[0044] Comparative Example 2: Enzyme-crosslinked whey protein-quercetin complex without ultrasonic treatment

[0045] S1: Dissolve whey protein in deionized water to prepare a whey protein solution with a concentration of 60 g / L, and adjust the pH value of the whey protein solution to 7.0 with a 0.2 mol / L NaOH solution to obtain a whey protein solution, which is recorded as WPI;

[0046] S2: The whey protein solution obtained in S1 is placed in a 50°C water bath for preheating for 15 minutes, and then horseradish peroxidase HRP, glucose oxidase and D-glucose are added, and the mixture is placed at 37°C for enzyme cross-linking reaction for 3 hours. After the reaction, the mixture is taken out and immediately placed in an 85°C water bath for enzyme inactivation for 10 minutes, and cooled at room temperature. The whey protein concentration after the water mixing is 50 g / L, and the horseradish peroxidase HRP, glucose oxidase and D-glucose are added in an amount of 200 U / g, 6 U / g and 0.05 mmol per gram of whey protein, respectively; the whey protein after the enzyme cross-linking reaction is obtained, which is recorded as CWPI;

[0047] S3: Adding quercetin solution to the whey protein after enzyme cross-linking reaction in S2 with a concentration of 1% (w / w), the mixing mass ratio of the whey protein to the quercetin solution is 20:1; then adjusting the pH value of the mixed solution to 9.0, placing it at 25°C for stirring and reacting for 24 hours, dialyzing it at 4°C for 48 hours using a 3500Da dialysis bag, and drying to obtain an enzyme-cross-linked whey protein-quercetin complex, recorded as CWPI-Q.

[0048] Effect experiment:

[0049] (1) Emulsification determination method

[0050] The sample solution with a concentration of 1 mg / mL was mixed with soybean oil in a ratio of 3:1 (v / v), and homogenized for 2 min at 10000 rpm using an IKA homogenizer to obtain an emulsion; 50 μL of the obtained emulsion was taken out from the bottom of the container at 0 and 10 min after homogenization, and mixed with 5 ml g / L SDS to obtain a mixed emulsion; the absorbance value of the mixed emulsion was read at 500 nm using a UV-visible spectrophotometer, and 1 g / L SDS solution was used as a blank control to calculate the emulsion stability index ESI.

[0051]

[0052] Where: A0 and A 10 Represent the absorbance of the sample at 0 min and 10 min, respectively.

[0053] (2) Determination of thiol content

[0054] Take 1.25 mL of each sample solution with a concentration of 6 mg / mL, mix them with Tris-glycine buffer (3.75 mL) containing 8 M urea, then add 50 μL of EIIman reagent, react at 40°C for 25 min, measure the absorbance at 412 nm, use deionized water as a blank control, and calculate the thiol content -SH content.

[0055]

[0056] In the formula, 73.53 = 10 6 / (1.36×10 4 ), 1.36×10 4 is the molar extinction coefficient; A 412 is the absorbance value of the sample at 412 nm; D is the dilution factor (D=100); C is the protein concentration of the sample (mg / mL).

[0057] (3) Method for determining the reducing ability of ferric iron

[0058] Take 1 mL of each sample solution with a concentration of 4 mg / mL, add 2.5 mL of water and 2.5 mL of potassium ferricyanide solution (10 mg / mL), mix well and react in a 50°C water bath for 20 min, add 2.5 mL of trichloroacetic acid solution (100 mg / mL) respectively, centrifuge at 3000 r / min for 10 min, take 2.5 mL of supernatant, add 2.5 ml of deionized water and 0.5 mL of FeCl3 solution (1 mg / ml), mix well and let stand for 10 min, and measure its absorbance A 700 , with deionized water as the control; the reducing ability of ferric iron is represented by absorbance, and the larger the absorbance value, the stronger its reducing power.

[0059] (4) ABTS free radical scavenging rate determination method

[0060] Mix equal volumes of ABTS aqueous solution (7 mmoL / L) and potassium persulfate aqueous solution (2.45 mmoL / L) of the sample solution, and store it at 4°C in the dark for 12-16 h as a ready-to-use reagent; dilute the reagent with anhydrous ethanol and adjust the OD value at 734 nm to 0.70±0.02, take 300 μL of sample (1 mg / mL) and mix with 3 mL of ABTS, let it stand at room temperature in the dark for 6 min, and measure the absorbance at 734 nm. Use anhydrous ethanol instead of the sample group as a blank control.

[0061] ABTS=(A0-A1) / A0×100%

[0062] Where A0 is the absorbance value of the blank group; A1 is the absorbance value of the sample.

[0063] (5) Solubility determination method

[0064] The 1 mg / mL sample solution was centrifuged at 10,000×g for 25 min, the supernatant was taken for later use, and the protein content of the supernatant was determined by the BCA method; the solubility of the sample was expressed as the ratio (%) of the protein content in the supernatant to the total protein content in the solution.

[0065] (6) Particle size and Zeta determination method

[0066] The sample was diluted to 1 mg / mL and its particle size and zeta potential were measured using a particle size and potential analyzer (Malvern Instruments Ltd, UK).

[0067] Result analysis:

[0068] The solubility of UCWPI-Q prepared in Example 1, WPI-Q prepared in Comparative Example 1 and CWPI-Q prepared in Comparative Example 2 was measured. The results are as follows: Figure 1 As shown; it can be seen that compared with WPI-Q without ultrasonic treatment and enzyme cross-linking reaction, and CWPI-Q without ultrasonic treatment alone, the UCWPI-Q prepared by the synergistic effect of ultrasonic treatment and enzyme cross-linking reaction in the present invention has stronger solubility, with a solubility of up to 90%; and the solubility of UCWPI-Q is significantly different from that of WPI-Q and CWPI-Q.

[0069] The emulsion stability of UCWPI-Q prepared in Example 1, WPI-Q prepared in Comparative Example 1 and CWPI-Q prepared in Comparative Example 2 was measured. The results are as follows: Figure 2 As shown; it can be seen that compared with WPI-Q without ultrasonic treatment and enzyme cross-linking reaction, and CWPI-Q without ultrasonic treatment only, the UCWPI-Q prepared by the synergistic effect of ultrasonic treatment and enzyme cross-linking reaction in the present invention has higher emulsification stability, and the emulsification stability can reach 70%; and the emulsification stability of UCWPI-Q is significantly different from that of WPI-Q and CWPI-Q.

[0070] The UCWPI-Q prepared in Example 1, the WPI-Q prepared in Comparative Example 1 and the CWPI-Q prepared in Comparative Example 2 were tested for ferric iron reducing power. Figure 3 As shown; it can be seen that compared with WPI-Q without ultrasonic treatment and enzyme cross-linking reaction, and CWPI-Q without ultrasonic treatment alone, the UCWPI-Q prepared by the synergistic effect of ultrasonic treatment and enzyme cross-linking reaction in the present invention has a higher ferric iron reducing power, and the ferric iron reducing power of UCWPI-Q is significantly different from that of WPI-Q and CWPI-Q.

[0071] The ABTS free radical scavenging rate of UCWPI-Q prepared in Example 1, WPI-Q prepared in Comparative Example 1 and CWPI-Q prepared in Comparative Example 2 was measured. The results are as follows: Figure 4 As shown; it can be seen that compared with WPI-Q without ultrasonic treatment and enzyme cross-linking reaction, and CWPI-Q without ultrasonic treatment alone, the UCWPI-Q prepared by the synergistic effect of ultrasonic treatment and enzyme cross-linking reaction in the present invention has a stronger ABTS free radical scavenging ability, indicating that it has a stronger antioxidant ability.

[0072] The free thiol content of UCWPI-Q prepared in Example 1, WPI-Q prepared in Comparative Example 1 and CWPI-Q prepared in Comparative Example 2 was measured. The results are as follows: Figure 5 As shown; it can be seen that compared with WPI-Q without ultrasonic treatment and enzyme cross-linking reaction, CWPI-Q without ultrasonic treatment and UCWPI-Q after ultrasonic treatment and enzyme cross-linking reaction both significantly reduced the free thiol content, among which the free thiol content in UCWPI-Q was slightly lower than that in CWPI-Q; indicating that ultrasonic treatment can promote the exposure of thiol structure of whey protein, thereby promoting the modification of whey protein; at the same time, the synergistic effect of ultrasonic treatment and enzyme cross-linking reaction has a better effect on promoting the exposure of thiol structure of whey protein.

[0073] The particle size and Zeta potential of UCWPI-Q prepared in Example 1, WPI-Q prepared in Comparative Example 1 and CWPI-Q prepared in Comparative Example 2 were measured. The results are as follows: Figure 6-7 As shown; it can be seen that compared with WPI-Q without ultrasonic treatment and enzyme cross-linking reaction, CWPI-Q without ultrasonic treatment and UCWPI-Q after ultrasonic treatment and enzyme cross-linking reaction both significantly reduced the particle size and Zeta potential, among which the particle size and Zeta potential of UCWPI-Q were slightly lower than those of CWPI-Q; indicating that ultrasonic treatment can effectively reduce the particle size and Zeta potential of whey protein, thereby improving the uniformity and stability of whey protein.

[0074] In summary, the present invention uses the synergistic effect of ultrasonic treatment and enzyme cross-linking reaction to prepare a whey protein-quercetin complex; the results show that the synergistic effect of ultrasonic treatment and enzyme cross-linking reaction helps to improve the solubility, emulsification stability, antioxidant properties, uniformity and stability of whey protein, so that the whey protein can be better combined with quercetin to prepare a complex.

[0075] The solubility of UCWPI and UCWPI-Q prepared in Example 1 was measured. The results are as follows: Figure 8As shown, it can be seen that through the preparation method provided by the present invention, after the whey protein UCWPI subjected to ultrasonic treatment and enzyme cross-linking reaction is mixed with quercetin, not only the solubility of the complex is not reduced, but on the contrary the solubility of the complex is improved, indicating that the addition of quercetin improves the solubility of the complex, thereby improving the utilization rate of whey protein in the field of food processing.

[0076] The UCWPI and UCWPI-Q prepared in Example 1 were tested for ferric iron reducing power. The results are as follows: Fig. 9 As shown, it can be seen that through the preparation method provided by the present invention, after the whey protein UCWPI subjected to ultrasonic treatment and enzyme cross-linking reaction is mixed with quercetin, the ferric iron reducing power of the complex is improved, indicating that the addition of quercetin improves the ferric iron reducing power of the complex.

[0077] The ABTS free radical scavenging rate of UCWPI and UCWPI-Q prepared in Example 1 was measured. The results are as follows: Fig.10 As shown, it can be seen that through the preparation method provided by the present invention, after the whey protein UCWPI subjected to ultrasonic treatment and enzyme cross-linking reaction is mixed with quercetin, the ABTS free radical scavenging rate of the complex is improved, indicating that the addition of quercetin improves the ABTS free radical scavenging rate of the complex.

[0078] The free thiol content of UCWPI and UCWPI-Q prepared in Example 1 was measured. The results are as follows: Fig.11 As shown, it can be seen that through the preparation method provided by the present invention, after the whey protein UCWPI subjected to ultrasonic treatment and enzyme cross-linking reaction is mixed with quercetin, the free thiol content is significantly reduced; it shows that the binding of quercetin affects the internal structure of whey protein, thereby changing the exposure of free thiol groups inside the whey protein, thereby proving that the whey protein UCWPI subjected to ultrasonic treatment and enzyme cross-linking reaction has a higher binding amount for quercetin, which is more conducive to the modification of whey protein.

[0079] In summary, the present invention combines whey protein UCWPI prepared by the synergistic effect of ultrasonic treatment and enzyme cross-linking reaction with quercetin to prepare a whey protein-quercetin complex; the results show that the addition of quercetin has a certain promoting effect on the modification of whey protein, and its modification effect is significantly higher than that of a single treatment method.

[0080] The invention modifies whey protein by combining three different modification methods, namely ultrasonic treatment, enzymatic cross-linking and combination with polyphenols, and the solubility, emulsification stability, antioxidant property, uniformity and stability of the prepared enzyme-cross-linked whey protein-quercetin complex are significantly improved.

[0081] The contents not described in detail in the specification of the present invention are well-known technologies for those skilled in the art. Although the present invention has been disclosed as above with preferred embodiments, it is not intended to limit the present invention. Anyone familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the protection scope of the present invention should be based on the definition of the claims.

Claims

1. A method for preparing an enzyme-crosslinked whey protein-quercetin complex, characterized in that: The preparation method comprises the following steps: S1: Dissolve whey protein in deionized water to prepare a whey protein solution, and adjust the pH value of the whey protein solution to 7.0 using a 0.2 mol / L NaOH solution; perform ultrasonic treatment on the whey protein solution in an ice water bath, with an ultrasonic power of 200 W, an ultrasonic time of 5 min, and an ultrasonic frequency of 25 Hz to obtain an ultrasonically treated whey protein solution; S2: The whey protein solution after ultrasonic treatment in S1 is placed in a 50°C water bath for preheating for 15 minutes, and then horseradish peroxidase HRP, glucose oxidase and D-glucose are added, and water is added to mix and then enzyme cross-linking reaction is carried out. After the reaction, the solution is taken out and immediately inactivated, and cooled at room temperature to obtain whey protein after ultrasonic treatment and enzyme cross-linking reaction; S3: adding quercetin solution to the whey protein obtained in S2 through ultrasonic treatment and enzyme cross-linking reaction, adjusting the pH value to 9.0, reacting with stirring at 25° C. for 24 h, dialyzing, and drying to obtain an enzyme-cross-linked whey protein-quercetin complex.

2. The preparation method according to claim 1, characterized in that: The concentration of the whey protein solution obtained by the configuration described in S1 is 60 g / L.

3. The preparation method according to claim 1, characterized in that: The whey protein concentration after adding water and mixing in S2 is 50 g / L, and the added amounts of horseradish peroxidase HRP, glucose oxidase and D-glucose are 200 U / g, 6 U / g and 0.05 mmol respectively per gram of whey protein.

4. The preparation method according to claim 1, characterized in that: The enzyme cross-linking reaction conditions in S2 are: placing the enzyme cross-linking reaction at 37° C. for 3 hours.

5. The preparation method according to claim 1, characterized in that: The enzyme inactivation treatment conditions in S2 are: placing in a water bath at 80-150° C. to inactivate the enzyme for 5-20 minutes.

6. The preparation method according to claim 1, characterized in that: The whey protein concentration obtained in S2 for the ultrasonic treatment-assisted enzyme cross-linking reaction was 1% (w / w).

7. The preparation method according to claim 1, characterized in that: The mixing mass ratio of the whey protein to the quercetin solution in S3 is 20:

1.

8. The preparation method according to claim 1, characterized in that: The dialysis conditions described in S3 are: using a 3500Da dialysis bag for 48 hours at 4°C.

9. An enzyme-crosslinked whey protein-quercetin complex, characterized in that: The composite is obtained by the preparation method according to any one of claims 1 to 8.

10. Use of the enzyme-crosslinked whey protein-quercetin complex according to claim 9 in food processing.

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