Preparation method of magnetic cottonseed protein nanospheres with high lipase loading capacity

Abstract

The invention discloses a preparation method of a magnetic cottonseed protein nanosphere with high lipase loading capacity, and relates to the technical field of enzyme engineering and lipase loading in the biological industry, and the preparation method comprises the following steps: coating nano Fe3O4 particles by using plant polyphenol and cottonseed protein by using an anti-solvent method to obtain the magnetic cottonseed protein nanosphere; adding the magnetic nano-protein spheres into a lipase solution, loading lipase on the magnetic cottonseed protein nano-spheres by utilizing covalence and secondary action between plant polyphenol and cottonseed protein, and separating, washing and drying to obtain the magnetic cottonseed protein nano-spheres with high lipase loading capacity. According to the preparation method, the magnetic cottonseed protein nanospheres are prepared by coating Fe3O4 nanoparticles with an anti-solvent method, lipase is immobilized on the surfaces of the nanospheres through covalence and secondary action between plant polyphenol and protein, parameters are optimized, and the magnetic cottonseed protein nanospheres with high lipase loading capacity are obtained. The magnetic cottonseed protein nanospheres obtained by the invention can realize efficient catalysis and cyclic utilization of lipase.

Description

Technical Field

[0001] The invention belongs to the technical field of enzyme engineering and lipase loading in the biological industry, and more specifically, relates to a method for preparing magnetic cottonseed protein nanospheres with high lipase loading. Background Art

[0002] With the rapid development of biotechnology, enzymes as biocatalysts have shown great application potential in the fields of food industry, medicine, environmental protection and chemical synthesis. Among them, lipase, as an important type of hydrolase, is scientifically known as triacylglycerol acyl hydrolase. It is an enzyme that can catalyze the hydrolysis of fats and oils. It is widely present in the tissues of animals, plants and microorganisms (such as molds, bacteria, etc.) containing fat; lipase can catalyze the hydrolysis of triglycerides into fatty acids and glycerol, and is widely used in food oil processing, detergent production, medicine and biochemical preparation. However, traditional lipases have problems such as easy inactivation and difficulty in recycling during use, which seriously limits their industrial application.

[0003] In order to overcome these challenges, researchers are committed to developing new enzyme immobilization technologies to improve the stability, activity and recyclability of enzymes. Among them, nanocarrier technology has become a popular research direction for enzyme immobilization due to its unique physicochemical properties, such as high specific surface area, good biocompatibility and easy surface modification. In particular, magnetic nanoparticles, such as Fe3O4, provide a new solution for the recycling of enzymes due to their easy separation and recycling properties under an external magnetic field.

[0004] Cottonseed protein is a natural and renewable protein resource with excellent biodegradability and biocompatibility. It is widely available and low-cost. In recent years, there has been an increasing number of studies on the use of cottonseed protein as a carrier material and the preparation of functionalized nanomaterials in combination with nanotechnology. However, directly immobilizing lipase on cottonseed protein nanospheres often has problems such as low loading efficiency and large enzyme activity loss during immobilization. Summary of the invention

[0005] An object of the present invention is to solve at least the above problems and / or disadvantages and to provide at least the advantages which will be described hereinafter.

[0006] In order to achieve these purposes and other advantages according to the present invention, a method for preparing magnetic cottonseed protein nanospheres with high lipase load is provided, comprising: using an anti-solvent method to coat nano Fe3O4 particles with cottonseed protein modified with plant polyphenols to obtain magnetic cottonseed protein nanospheres, adding the magnetic nanoprotein nanospheres to a lipase solution, utilizing the covalent and secondary effects between plant polyphenols and cottonseed protein to load the lipase on the magnetic cottonseed protein nanospheres, separating, washing, and drying to obtain magnetic cottonseed protein nanospheres with high lipase load.

[0007] Preferably, the method comprises the following steps:

[0008] Step 1: dispersing nano Fe3O4 in deionized water to obtain a nano Fe3O4 particle suspension; dissolving plant polyphenols and cottonseed protein powder in deionized water to obtain plant polyphenol solutions and cottonseed protein solutions respectively;

[0009] Step 2: using a plant polyphenol solution and a cottonseed protein solution to prepare cottonseed protein modified with plant polyphenols; adding a nano-Fe3O4 particle suspension to the cottonseed protein modified with plant polyphenols, and uniformly dispersing the suspension by ultrasonication to obtain a mixed system;

[0010] Step 3, stirring the mixed system, and simultaneously adding an anti-solvent to the mixed system, with the addition of the anti-solvent, the solubility of the cottonseed protein modified by plant polyphenols decreases, and the cottonseed protein gradually precipitates on the surface of the nano-Fe3O4 particles, and the surface of the nano-Fe3O4 particles is coated, and after standing for a period of time at a certain temperature, the solid-liquid separation is performed, and the magnetic cottonseed protein nanospheres are obtained by washing and drying;

[0011] Step 4: Add the magnetic cottonseed protein nanospheres to the lipase solution, utilize the covalent and secondary effects between plant polyphenols and cottonseed protein to load the lipase on the magnetic cottonseed protein nanospheres, perform solid-liquid separation, wash, and dry to obtain magnetic cottonseed protein nanospheres with high lipase load.

[0012] Preferably, in step 1, the mass volume ratio of nano-Fe3O4 to deionized water is 1-10 g: 20-50 mL; the mass volume ratio of plant polyphenols to deionized water is 1-2 g: 20 mL; and the mass volume ratio of cottonseed protein powder to deionized water is 3-5 g: 50-100 mL.

[0013] Preferably, in the step 2, the specific method for preparing cottonseed protein modified with plant polyphenols includes: stirring the cottonseed protein powder solution at a speed of 200 to 800 rpm, then dripping glutaraldehyde drop by drop, stirring continuously and standing to obtain a cross-linked system; stirring the cross-linked system at a speed of 600 to 1200 rpm, dripping the plant polyphenol solution into the cross-linked system drop by drop, stirring continuously, heating to 60 to 80° C. and keeping warm for 1 to 2 hours after the addition is completed, stopping stirring, and cooling to obtain cottonseed protein modified with plant polyphenols; wherein the volume ratio of the plant polyphenol solution to the cottonseed protein solution is 1:1, and the mass ratio of glutaraldehyde to the plant polyphenols is 1 to 3:5 to 10.

[0014] Preferably, in step 1, the plant polyphenol is one of tea polyphenol, apple polyphenol, grape polyphenol, resveratrol and bayberry tannin.

[0015] Preferably, in step 2, the volume ratio of the plant polyphenol solution, the cottonseed protein solution and the nano-Fe3O4 particle suspension is 1:1:2-4; the ultrasonic dispersion frequency is 40-60 kHz, and the dispersion time is 1-3 hours.

[0016] Preferably, in the step 3, the anti-solvent is one of petroleum ether, chloroform or benzene; and the volume ratio of the nano-Fe3O4 particle suspension to the anti-solvent is 2-4:5-8.

[0017] Preferably, in step three, the standing temperature is 30-60° C. and the standing time is 12-24 hours.

[0018] Preferably, in step 4, the concentration of the lipase solution is 1-1.6 g / mL, and the mass volume ratio of the magnetic cottonseed protein nanospheres to the lipase solution is 5-10 mg:1 mL.

[0019] Preferably, the nano-Fe3O4 in step 1 is replaced by modified nano-Fe3O4, wherein the preparation method of the modified nano-Fe3O4 comprises: selecting nano-Fe3O4 with a particle size distribution of 20 to 500 nm, ultrasonically dispersing it in anhydrous ethanol, the ultrasonic frequency is 30 to 60 kHz, and the ultrasonic time is 20 to 40 minutes to obtain a nano-Fe3O4 suspension; adding polyvinyl pyrrolidone and γ-aminopropyltriethoxysilane to the nano-Fe3O4 suspension, magnetically stirring at a speed of 1200 to 2400 rpm for 1 to 2 hours, and heating it to 80 to 90°C within 5 to 10 minutes. 0℃, keep warm for 15-60min, and then cool to room temperature; add sodium dodecylbenzene sulfonate, ultrasonic dispersion, the ultrasonic frequency is 40-80kHz, the ultrasonic time is 20-60min, the temperature is raised to 60-80℃ and then kept warm for 1-2h, solid-liquid separation, washing and drying to obtain modified nano-Fe3O4, wherein the mass volume ratio of nano-Fe3O4, anhydrous ethanol, polyvinyl pyrrolidone, sodium dodecylbenzene sulfonate and γ-aminopropyltriethoxysilane is 0.5-1g:10-30mL:0.2-0.5g:0.05-0.2g:0.1-0.2g.

[0020] The present invention at least includes the following beneficial effects: The present invention discloses a method for preparing a magnetic cottonseed protein nanosphere with a high load of lipase. The magnetic cottonseed protein nanosphere is prepared by coating nanoparticles Fe3O4 with an antisolvent method, and the lipase is fixed on the surface of the nanosphere by covalent and secondary interactions between plant polyphenols and proteins. The parameters are optimized to obtain the magnetic cottonseed protein nanosphere with a high load of lipase. The magnetic cottonseed protein nanosphere obtained by the present invention can realize the efficient catalysis and recycling of lipase.

[0021] The invention also uses organic polymer polyvinyl pyrrolidone to coat nano Fe3O4, thereby increasing the surface area of ​​nano Fe3O4, and uses gamma-aminopropyl triethoxysilane to increase the tightness of the combination between polyvinyl pyrrolidone and nano Fe3O4, and introduces amino groups to increase the active sites on the surface of nano Fe3O4 coated with polyvinyl pyrrolidone, thereby promoting the combination of cottonseed protein and nano Fe3O4 coated with polyvinyl pyrrolidone; subsequently, sodium dodecylbenzene sulfonate is used as a dispersant to disperse the nano Fe3O4 coated with polyvinyl pyrrolidone, thereby reducing the agglomeration between the nano Fe3O4 coated with polyvinyl pyrrolidone, and preparing modified nano Fe3O4 with good dispersibility, small agglomeration and large surface area; the modified nano Fe3O4 with good dispersibility, small agglomeration and large surface area can coat more cottonseed protein modified with plant polyphenols, thereby further increasing the loading amount of the magnetic cottonseed protein nanospheres on lipase.

[0022] Other advantages, objectives and features of the present invention will be embodied in part through the following description, and in part will be understood by those skilled in the art through study and practice of the present invention. DETAILED DESCRIPTION

[0023] The present invention is described in further detail below so that those skilled in the art can implement it according to the description.

[0024] It should be understood that the terms such as “having”, “including” and “comprising” used herein do not exclude the existence or addition of one or more other elements or combinations thereof.

[0025] Example 1

[0026] This embodiment provides a method for preparing magnetic cottonseed protein nanospheres with high lipase loading, comprising the following steps:

[0027] Step 1: Disperse 1g of nano-Fe3O4 in 20mL of deionized water to obtain a nano-Fe3O4 particle suspension; dissolve 1g of bayberry tannin in 20mL of deionized water, and dissolve 3g of cottonseed protein powder in 50mL of deionized water to obtain bayberry tannin solution and cottonseed protein solution respectively;

[0028] Step 2, stirring the cottonseed protein powder solution at a speed of 600 rpm, then dripping 0.2 g of glutaraldehyde drop by drop, stirring continuously and then standing to obtain a cross-linked system; stirring the cross-linked system at a speed of 600 rpm, dripping 20 mL of bayberry tannin solution drop by drop into the cross-linked system (the volume of the bayberry tannin solution and the cottonseed protein solution is the same), stirring continuously, heating to 80° C. and keeping warm for 2 h after the addition is completed, stopping stirring, and cooling to obtain cottonseed protein modified with bayberry tannin; adding 40 mL of nano-Fe3O4 particle suspension to the cottonseed protein modified with bayberry tannin, ultrasonically dispersing evenly, the ultrasonic dispersion frequency is 60 kHz, and the dispersion time is 3 h to obtain a mixed system;

[0029] Step 3, stirring the mixed system, and at the same time, adding 100 mL of petroleum ether as an anti-solvent to the mixed system, with the addition of the anti-solvent, the solubility of the cottonseed protein modified with bayberry tannin decreases, and gradually precipitates on the surface of the nano-Fe3O4 particles, and coats the surface of the nano-Fe3O4 particles. After standing at a temperature of 30 to 60° C. for 12 to 24 hours, the solid-liquid separation is performed, and the magnetic cottonseed protein nanospheres are obtained by washing and drying;

[0030] Step 4: Add 10 mg of magnetic cottonseed protein nanospheres to 1 mL of 1.2 g / mL lipase solution, utilize the covalent interaction between bayberry tannin and cottonseed protein to load the lipase on the magnetic cottonseed protein nanospheres, perform solid-liquid separation, wash, and dry to obtain magnetic cottonseed protein nanospheres with high lipase load.

[0031] Example 2

[0032] This embodiment provides a method for preparing magnetic cottonseed protein nanospheres with high lipase loading, comprising the following steps:

[0033] Step 1: Disperse 4 g of nano-Fe3O4 in 40 mL of deionized water to obtain a nano-Fe3O4 particle suspension; dissolve 2 g of apple polyphenol in 20 mL of deionized water, and dissolve 4 g of cottonseed protein powder in 60 mL of deionized water to obtain an apple polyphenol solution and a cottonseed protein solution, respectively;

[0034] Step 2: Stir the cottonseed protein powder solution at a speed of 800 rpm, then drop 0.4 g of glutaraldehyde, continue stirring and let stand to obtain a cross-linked system; stir the cross-linked system at a speed of 1200 rpm, drop 60 mL of apple polyphenol solution into the cross-linked system (the volume of the apple polyphenol solution and the cottonseed protein solution is the same), continue stirring, heat to 80°C and keep warm for 2 hours after the addition is completed, stop stirring, and cool to obtain cottonseed protein modified with apple polyphenol; add 60 mL of nano-Fe3O4 particle suspension to the cottonseed protein modified with apple polyphenol, disperse evenly with ultrasonic dispersion frequency of 60 kHz and dispersion time of 2 hours to obtain a mixed system;

[0035] Step 3, stirring the mixed system, and at the same time, adding 120 mL of chloroform as an anti-solvent to the mixed system, with the addition of the anti-solvent, the solubility of the cottonseed protein modified by apple polyphenols decreases, and gradually precipitates on the surface of the nano-Fe3O4 particles, coating the surface of the nano-Fe3O4 particles, standing at 60°C for 12 hours, separating the solid and the liquid, washing and drying to obtain magnetic cottonseed protein nanospheres;

[0036] Step 4: Add 10 mg of magnetic cottonseed protein nanospheres into 1 mL of 1.2 g / mL lipase solution, utilize the covalent and secondary effects between apple polyphenols and cottonseed protein to load the lipase on the magnetic cottonseed protein nanospheres, perform solid-liquid separation, wash, and dry to obtain magnetic cottonseed protein nanospheres with high lipase load.

[0037] Example 3

[0038] This embodiment provides a method for preparing magnetic cottonseed protein nanospheres with high lipase loading, comprising the following steps:

[0039] Step 1: Disperse 5 g of nano-Fe3O4 in 50 mL of deionized water to obtain a nano-Fe3O4 particle suspension; dissolve 2 g of grape polyphenol in 20 mL of deionized water, and dissolve 5 g of cottonseed protein powder in 100 mL of deionized water to obtain a grape polyphenol solution and a cottonseed protein solution, respectively;

[0040] Step 2: Stir the cottonseed protein powder solution at a speed of 800 rpm, then drop 0.6 g of glutaraldehyde, continue stirring and let stand to obtain a cross-linked system; stir the cross-linked system at a speed of 1200 rpm, drop 100 mL of grape polyphenol solution into the cross-linked system (the volume of the grape polyphenol solution is the same as that of the cottonseed protein powder solution), continue stirring, heat to 80° C. and keep warm for 2 h after the addition is completed, stop stirring, and cool to obtain cottonseed protein modified with grape polyphenol; add 80 mL of nano-Fe3O4 particle suspension to the cottonseed protein modified with grape polyphenol, disperse evenly with ultrasonic dispersion at a frequency of 60 kHz and a dispersion time of 3 h to obtain a mixed system;

[0041] Step 3, stirring the mixed system, and at the same time, adding 160 mL of benzene as an anti-solvent to the mixed system, with the addition of the anti-solvent, the solubility of the cottonseed protein modified by grape polyphenols decreases, and gradually precipitates on the surface of the nano-Fe3O4 particles, coating the surface of the nano-Fe3O4 particles, standing at 60°C for 12 hours, separating the solid and the liquid, washing and drying to obtain magnetic cottonseed protein nanospheres;

[0042] Step 4: Add 10 mg of magnetic cottonseed protein nanospheres to 1 mL of 1.2 g / mL lipase solution, utilize the covalent interaction between grape polyphenols and cottonseed protein to load the lipase on the magnetic cottonseed protein nanospheres, perform solid-liquid separation, wash, and dry to obtain magnetic cottonseed protein nanospheres with high lipase load.

[0043] Example 4

[0044] This embodiment provides a method for preparing magnetic cottonseed protein nanospheres with high lipase loading. Compared with Example 1, the nano Fe3O4 in step 1 is replaced with modified nano Fe3O4, and the process parameters of the remaining steps are the same as those in Example 1; wherein the preparation method of modified nano Fe3O4 includes:

[0045] 10 g of nano-Fe3O4 with a particle size distribution of 20 to 500 nm was selected and ultrasonically dispersed in 250 mL of anhydrous ethanol at an ultrasonic frequency of 60 kHz and an ultrasonic time of 40 min to obtain a nano-Fe3O4 suspension;

[0046] Add 2g polyvinyl pyrrolidone and 1g γ-aminopropyltriethoxysilane to the nano-Fe3O4 suspension, stir magnetically at 1200rpm for 2h, heat to 80°C within 5min, keep warm for 30min, and cool to room temperature; add 0.5g sodium dodecylbenzene sulfonate, ultrasonically disperse, the ultrasonic frequency is 60kHz, the ultrasonic time is 30min, heat to 80°C and keep warm for 2h, separate the solid and liquid, wash and dry to obtain modified nano-Fe3O4.

[0047] Example 5

[0048] This embodiment provides a method for preparing magnetic cottonseed protein nanospheres with high lipase loading. Compared with Example 1, the nano Fe3O4 in step 1 is replaced with modified nano Fe3O4, and the process parameters of the remaining steps are the same as those in Example 1; wherein the preparation method of modified nano Fe3O4 includes:

[0049] 10g of nano-Fe3O4 with a particle size distribution of 20-500nm was selected and ultrasonically dispersed in 200mL of anhydrous ethanol at an ultrasonic frequency of 60kHz and an ultrasonic time of 40min to obtain a nano-Fe3O4 suspension;

[0050] Add 3g polyvinyl pyrrolidone and 2g γ-aminopropyltriethoxysilane to the nano-Fe3O4 suspension, stir magnetically at 2400rpm for 2h, heat to 90°C within 10min, keep warm for 60min, and cool to room temperature; add 1g sodium dodecylbenzenesulfonate, ultrasonically disperse, the ultrasonic frequency is 80kHz, the ultrasonic time is 60min, heat to 80°C and keep warm for 1h, separate the solid and liquid, wash and dry to obtain modified nano-Fe3O4.

[0051] Comparative Example 1

[0052] The present comparative example provides a method for preparing magnetic cottonseed protein nanospheres with high lipase load. Compared with Example 1, the operation of modifying cottonseed protein with bayberry tannin in step 2 is not performed, and cottonseed protein modified with bayberry tannin is not used to coat nano-Fe3O4 in step 3. Instead, 20 mL of bayberry tannin solution and 20 mL of cottonseed protein solution are directly mixed with 40 mL of nano-Fe3O4 particle suspension, and ultrasonic dispersion is uniform. The ultrasonic dispersion frequency is 60 kHz and the dispersion time is 3 h to obtain a mixed system. The mixed system is then stirred, and 120 mL of chloroform is dripped into the mixed system as an anti-solvent. With the dripping of the anti-solvent, the solubility of cottonseed protein modified with apple polyphenols decreases, and it is gradually precipitated on the surface of nano-Fe3O4 particles to coat the surface of nano-Fe3O4 particles. After standing at 60°C for 12 h, solid-liquid separation is performed, and the magnetic cottonseed protein nanospheres are obtained by washing and drying.

[0053] 10 mg of magnetic cottonseed protein nanospheres were added to 1 mL of 1.2 g / mL lipase solution. The lipase was loaded on the magnetic cottonseed protein nanospheres by utilizing the covalent and secondary effects between bayberry tannins and cottonseed protein. After solid-liquid separation, washing and drying, magnetic cottonseed protein nanospheres with high lipase loading were obtained.

[0054] Comparative Example 2

[0055] This comparative example provides a method for preparing magnetic nanospheres loaded with lipase. Compared with Example 1, 10 mg of nano Fe3O4 particle suspension is directly added to 1 mL of 1.2 g / mL lipase solution, solid-liquid separation is performed, and the magnetic nanospheres loaded with lipase are obtained by washing and drying.

[0056] The lipase loading of the magnetic cottonseed protein nanospheres with high lipase loading prepared in Examples 1 to 5, the magnetic cottonseed protein nanospheres with high lipase loading prepared in Comparative Example 1, and the lipase-loaded magnetic nanospheres prepared in Comparative Example 2 were measured respectively, and the following table was obtained:

[0057] Lipase loading (mg / g) Example 1 205.2 Example 2 194.0 Example 3 188.5 Example 4 252.6 Example 5 248.9 Comparative Example 1 155.7 Comparative Example 2 124.6

[0058] As can be seen from the above table, the magnetic cottonseed protein nanosphere samples with high lipase loading prepared in Examples 1 to 5 all achieved a high loading amount of lipase, and were significantly better than Comparative Example 1 and Comparative Example 2; among them, compared with Examples 1 to 3, the magnetic cottonseed protein nanosphere samples in Examples 4 and 5 had further improved the lipase loading amount.

[0059] The number of devices and processing scales described here are used to simplify the description of the present invention. Applications, modifications and variations of the present invention will be obvious to those skilled in the art.

[0060] Although the embodiments of the present invention have been disclosed as above, they are not limited to the applications listed in the specification and the implementation modes, and they can be fully applied to various fields suitable for the present invention. For those familiar with the art, additional modifications can be easily implemented. Therefore, without departing from the general concept defined by the claims and the scope of equivalents, the present invention is not limited to the specific details and the examples shown and described herein.

Claims

1. A method for preparing magnetic cottonseed protein nanospheres with high lipase loading, characterized in that: include: Using the anti-solvent method, nano-Fe3O4 particles were coated with cottonseed protein modified with plant polyphenols to obtain magnetic cottonseed protein nanospheres. The magnetic nanoprotein spheres were added to a lipase solution, and the lipase was loaded on the magnetic cottonseed protein nanospheres by utilizing the covalent and secondary effects between plant polyphenols and cottonseed protein. The particles were separated, washed, and dried to obtain magnetic cottonseed protein nanospheres with a high load of lipase.

2. The method for preparing the magnetic cottonseed protein nanospheres with high lipase loading according to claim 1, characterized in that: The following steps are involved: Step 1: dispersing nano Fe3O4 in deionized water to obtain a nano Fe3O4 particle suspension; dissolving plant polyphenols and cottonseed protein powder in deionized water to obtain plant polyphenol solutions and cottonseed protein solutions respectively; Step 2: using a plant polyphenol solution and a cottonseed protein solution to prepare cottonseed protein modified with plant polyphenols; adding a nano-Fe3O4 particle suspension to the cottonseed protein modified with plant polyphenols, and uniformly dispersing the suspension by ultrasonication to obtain a mixed system; Step 3, stirring the mixed system, and simultaneously adding an anti-solvent to the mixed system, with the addition of the anti-solvent, the solubility of the cottonseed protein modified by plant polyphenols decreases, and the cottonseed protein gradually precipitates on the surface of the nano-Fe3O4 particles, and the surface of the nano-Fe3O4 particles is coated, and after standing for a period of time at a certain temperature, the solid-liquid separation is performed, and the magnetic cottonseed protein nanospheres are obtained by washing and drying; Step 4: Add the magnetic cottonseed protein nanospheres to the lipase solution, utilize the covalent and secondary effects between plant polyphenols and cottonseed protein to load the lipase on the magnetic cottonseed protein nanospheres, perform solid-liquid separation, wash, and dry to obtain magnetic cottonseed protein nanospheres with high lipase loading.

3. The method for preparing the magnetic cottonseed protein nanospheres with high lipase loading according to claim 2, characterized in that: In the step 1, the mass volume ratio of nano-Fe3O4 to deionized water is 1-10 g: 20-50 mL; the mass volume ratio of plant polyphenols to deionized water is 1-2 g: 20 mL; and the mass volume ratio of cottonseed protein powder to deionized water is 3-5 g: 50-100 mL.

4. The method for preparing the magnetic cottonseed protein nanospheres with high lipase loading according to claim 2, characterized in that: In the step 2, the specific method for preparing cottonseed protein modified with plant polyphenols includes: stirring the cottonseed protein powder solution at a rotation speed of 200 to 800 rpm, then dripping glutaraldehyde drop by drop, stirring continuously and then standing to obtain a cross-linked system; stirring the cross-linked system at a rotation speed of 600 to 1200 rpm, dripping the plant polyphenol solution into the cross-linked system drop by drop, stirring continuously, heating to 60 to 80° C. and keeping warm for 1 to 2 hours after the dropwise addition is completed, stopping stirring, and cooling to obtain cottonseed protein modified with plant polyphenols; wherein the volume ratio of the plant polyphenol solution to the cottonseed protein solution is 1:1, and the mass ratio of glutaraldehyde to the plant polyphenols is 1 to 3:5 to 10.

5. The method for preparing the magnetic cottonseed protein nanospheres with high lipase loading according to claim 2, characterized in that: In the step 1, the plant polyphenol is one of tea polyphenol, apple polyphenol, grape polyphenol, resveratrol and bayberry tannin.

6. The method for preparing the magnetic cottonseed protein nanospheres with high lipase loading according to claim 2, characterized in that: In the step 2, the volume ratio of the plant polyphenol solution, the cottonseed protein solution and the nano-Fe3O4 particle suspension is 1:1:2-4; the ultrasonic dispersion frequency is 40-60 kHz, and the dispersion time is 1-3 hours.

7. The method for preparing the magnetic cottonseed protein nanospheres with high lipase loading according to claim 2, characterized in that: In the step 3, the anti-solvent is one of petroleum ether, chloroform or benzene; the volume ratio of the nano-Fe3O4 particle suspension to the anti-solvent is 2-4:5-8.

8. The method for preparing the magnetic cottonseed protein nanospheres with high lipase loading according to claim 2, characterized in that: In the step 3, the standing temperature is 30 to 60° C. and the standing time is 12 to 24 hours.

9. The method for preparing the magnetic cottonseed protein nanospheres with high lipase loading according to claim 2, characterized in that: In the step 4, the concentration of the lipase solution is 1-1.6 g / mL, and the mass volume ratio of the magnetic cottonseed protein nanospheres to the lipase solution is 5-10 mg:1 mL.

10. The method for preparing magnetic cottonseed protein nanospheres with high lipase loading according to claim 2, characterized in that: The nano-Fe3O4 in step 1 is replaced with modified nano-Fe3O4. The preparation method of modified nano-Fe3O4 includes: selecting nano-Fe3O4 with a particle size distribution of 20 to 500 nm, ultrasonically dispersing it in anhydrous ethanol, with an ultrasonic frequency of 30 to 60 kHz and an ultrasonic time of 20 to 40 minutes to obtain a nano-Fe3O4 suspension; adding polyvinyl pyrrolidone and γ-aminopropyltriethoxysilane to the nano-Fe3O4 suspension, magnetically stirring at a speed of 1200 to 2400 rpm for 1 to 2 hours, heating it to 80 to 90° C. within 5 to 10 minutes, and keeping it warm for 5 minutes. 15 to 60 minutes, cool to room temperature; add sodium dodecylbenzene sulfonate, ultrasonic dispersion, the ultrasonic frequency is 40 to 80kHz, the ultrasonic time is 20 to 60 minutes, the temperature is raised to 60 to 80°C and then kept warm for 1 to 2 hours, solid-liquid separation, washing and drying to obtain modified nano-Fe3O4, wherein the mass volume ratio of nano-Fe3O4, anhydrous ethanol, polyvinyl pyrrolidone, sodium dodecylbenzene sulfonate and γ-aminopropyltriethoxysilane is 0.5 to 1g:10 to 30mL:0.2 to 0.5g:0.05 to 0.2g:0.1 to 0.2g.