Preparation method of azo functionalized magnetic nanoparticle drug-loaded protein

Abstract

The invention belongs to the technical field of biological medicine, and particularly relates to a preparation method of azo functionalized magnetic nanoparticle drug-loaded protein. According to the azo functionalized magnetic nanoparticle drug-loaded protein, 4-((4-aminophenyl) diazenyl) benzoic acid serves as an intermediate and is connected to carboxyl modified magnetic nanoparticles through an amidation reaction, and the magnetic nanoparticles serve as a cross-linking agent and are combined with bovine serum albumin and drugs to prepare the drug-loaded protein. The drug-loaded protein provided by the invention has a novel structure, and has unique design in structural composition and formula.

Description

Technical Field

[0001] The invention belongs to the technical field of biomedicine, and in particular relates to a method for preparing azo-functionalized magnetic nanoparticles and magnetic nanoparticle drug-loaded protein. Background Art

[0002] Smart responsive drug delivery system, that is, nanocarriers loaded with therapeutic drugs release drugs in the tumor microenvironment or under certain external specific stimuli after entering the tumor tissue to enhance the therapeutic effect. Albumin has been widely studied in drug delivery systems due to its excellent biocompatibility, biodegradability, high nutritional value and super strong binding ability with a variety of drugs.

[0003] Carrier proteins play an important role in drug delivery due to their unique molecular structure and functionality. Many carrier proteins, such as albumin and transferrin, can achieve targeted drug delivery by binding to specific receptors. In addition, carrier proteins have controllable drug binding capacity and sustained release properties, and can serve as a natural platform for drug storage and release. Binding magnetic nanoparticles to carrier proteins not only retains the biospecificity of the protein, but also improves the targeting of the drug through magnetic field guidance and external stimulation, forming an efficient and multifunctional drug delivery system.

[0004] The combination of magnetic nanoparticles and carrier proteins provides a synergistic mechanism for drug delivery. Under the action of an external magnetic field, magnetic nanoparticles can guide carrier proteins to specific sites, such as tumors or diseased tissues, thereby increasing local drug concentrations. At the same time, carrier proteins further enhance the stability and functionality of nanoparticles in vivo through their natural targeting and biocompatibility. In addition, by modifying the surface properties of magnetic nanoparticles (such as coating polymers or ligand modification), their binding efficiency with carrier proteins can be optimized, and the carriers can be given more abundant biological functions to meet diverse drug delivery needs.

[0005] Although the combination technology of magnetic nanoparticles and carrier proteins has shown great application potential, its practical application still faces many challenges. First, the stability of carrier proteins in the complex environment of the body may be affected by protein degradation or immune response, affecting the drug delivery efficiency. Secondly, the synthesis and surface modification of magnetic nanoparticles need to be precisely controlled to ensure their efficient binding with carrier proteins and maintain good biocompatibility. In addition, the penetration and control ability of magnetic fields in deep tissues of the human body is still the main bottleneck limiting the efficiency of drug delivery. Future research directions should focus on developing new coupling technologies of magnetic nanoparticles and carrier proteins, optimizing their synergistic effects in vivo and in vitro, and realizing more efficient and safe drug delivery platforms through multifunctional design. Summary of the invention

[0006] The purpose of the present invention is to overcome the defects of the prior art and provide a method for preparing azo-functionalized magnetic nanoparticles and magnetic nanoparticle drug-loaded protein. Azo-modified magnetic nanoparticles are prepared by synthesizing 4-((4-aminophenyl)diazenyl)benzoic acid and utilizing an amidation reaction; and nanoparticles based on bovine serum albumin are prepared by a solvation-chemical crosslinking method.

[0007] To achieve the above purpose, the technical solution adopted by the present invention is as follows:

[0008] An azo functionalized magnetic nanoparticle, the preparation method of which comprises the following steps:

[0009] 1) Synthesis of 4-((4-aminophenyl)diazenyl)benzoic acid: dissolve p-aminobenzoic acid in deionized water, add concentrated hydrochloric acid and stir for 20 minutes, and cool in an ice bath after stirring; weigh sodium nitrite and dissolve in deionized water, cool the dissolved sodium nitrite solution in an ice bath, and keep the temperature at 0-5°C; slowly drop the cooled sodium nitrite solution into the p-aminobenzoic acid solution in the ice bath, and react for 15 minutes; filter the solution after the reaction, retain the filtrate and store it in an ice bath; take another large beaker, dissolve sodium acetate and aniline sulfite in deionized water, keep the solution temperature at 0-5°C, then drop the filtrate into the large beaker and stir for 1 hour, adjust the pH to keep the solution acidic, filter and dry; after drying, react with sodium hydroxide solution at 90°C for 3 hours, finally adjust the solution pH to 6 in an ice bath, filter and dry, and obtain 4-((4-aminophenyl)diazenyl)benzoic acid;

[0010] 2) Synthesis of oleic acid-coated magnetic nanoparticles: Dissolve ferric chloride hexahydrate and ferrous chloride tetrahydrate in deionized water in a beaker, heat to 40°C and stir until dissolved, cool to room temperature and set aside; add the two solutions to a flask and set the temperature to 70°C, while nitrogen is introduced and concentrated ammonia is slowly added dropwise under mechanical stirring, and anhydrous ethanol mixed with oleic acid is quickly added, and the reaction is continued for 1 hour; after the reaction is completed, wash the magnetic particles three times with anhydrous ethanol and deionized water and then with a magnet, and dry them in a vacuum drying oven at 50°C to obtain oleic acid-coated magnetic nanoparticles;

[0011] 3) Synthesis of carboxyl-modified magnetic nanoparticles: oleic acid-coated magnetic nanoparticles and 4-vinylbenzoic acid were added to deionized water, ultrasonically dispersed, and placed in a water bath for 3 hours; potassium persulfate was dissolved in deionized water, and slowly added to the magnetic nanoparticle solution along the wall of the beaker, and the polymerization reaction was continued to proceed. The reaction was stopped after 5 hours, and the product separated by a magnet was washed alternately with anhydrous ethanol and deionized water, and the washed magnetic nanoparticles were placed in a vacuum drying oven at 60°C for 2 hours to obtain carboxyl-modified magnetic nanoparticles;

[0012] 4) Synthesis of azo functionalized magnetic nanoparticles: Carboxyl modified magnetic nanoparticles were placed in a three-necked flask, and DMF was added as a reaction solvent. The 4-((4-aminophenyl)diazenyl)benzoic acid prepared in step 1) was mechanically stirred in the three-necked flask until it was completely dissolved. After complete dissolution, catalysts 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 4-dimethylaminopyridine were added to the reaction solution. The reaction system was stirred at 36° C. for 48 hours to obtain azo functionalized magnetic nanoparticles.

[0013] Furthermore, in the above-mentioned method for preparing azo functionalized magnetic nanoparticles, in step 1), the amount of p-aminobenzoic acid is 13.7 g, the amount of concentrated hydrochloric acid is 33.6 mL, the amount of sodium nitrite is 6.9 g, the amount of aniline sulfite is 21.1 g, the amount of sodium acetate is 60 g, and the amount of sodium hydroxide is 15 g.

[0014] Furthermore, in the above-mentioned method for preparing azo-functionalized magnetic nanoparticles, in step 2), 5.00 g of ferric chloride hexahydrate and 2.00 g of ferrous chloride tetrahydrate are dissolved in 10 mL of deionized water respectively.

[0015] Furthermore, in the above-mentioned method for preparing azo-functionalized magnetic nanoparticles, in step 2), the amount of concentrated ammonia water used is 10 mL; the amount of oleic acid used is 1 mL.

[0016] Furthermore, in the above-mentioned method for preparing azo-functionalized magnetic nanoparticles, in step 3), the amount of the oleic acid-coated magnetic nanoparticles is 10 mg, the amount of 4-vinylbenzoic acid is 40 mg, and the amount of potassium persulfate is 8 mg.

[0017] Furthermore, in the above-mentioned method for preparing azo-functionalized magnetic nanoparticles, in step 4), the amount of the carboxyl-modified magnetic nanoparticles is 30 mg, and the amount of 4-((4-aminophenyl)diazenyl)benzoic acid is 0.12 g.

[0018] The invention discloses a method for preparing azo functionalized magnetic nanoparticle bovine serum albumin, comprising the following steps: taking the above-mentioned azo functionalized magnetic nanoparticles into a flask, adding anhydrous dimethyl sulfoxide thereto, adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide under mechanical stirring until they are completely dissolved, reacting at 37°C for 2 hours under light-proof conditions to obtain a black active ester solution for standby use; taking bovine serum albumin powder and dissolving it in deionized water, dripping 0.1 mol / L sodium hydroxide aqueous solution, adjusting the pH value to 9.0, continuing stirring for 1 hour, and dripping anhydrous ethanol at a speed of 4 mL / min; after desolvation is completed, slowly adding 0.8 mL of the black active ester solution for cross-linking and curing, continuing to react overnight under light-proof conditions at 37°C to obtain a stable albumin magnetic nanoparticle suspension; rotary evaporating the obtained albumin magnetic nanoparticle suspension at 37°C to remove ethanol, centrifuging at 10000 rpm for 15 minutes, and freeze-drying to obtain bovine serum albumin magnetic nanoparticles.

[0019] Furthermore, in the above-mentioned method for preparing azo-functionalized magnetic nanoparticles bovine serum albumin, the amount of the azo-functionalized magnetic nanoparticles is 30 mg, the amount of anhydrous dimethyl sulfoxide is 2 mL, 900 mg of bovine serum albumin powder is dissolved in 30 mL of deionized water, and the amount of anhydrous ethanol is 60 mL.

[0020] The invention discloses a method for preparing azo-functionalized magnetic nanoparticle drug-loaded protein, comprising the following steps: taking the above-mentioned azo-functionalized magnetic nanoparticles into a flask, adding anhydrous dimethyl sulfoxide thereto, adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide under mechanical stirring until they are completely dissolved, reacting at 37°C for 2 hours under light-proof conditions to obtain a black active ester solution for standby use; simultaneously taking bovine serum albumin powder and doxorubicin hydrochloride and dissolving them in deionized water, dripping 0.1 mol / L sodium hydroxide aqueous solution, adjusting the pH value to 9.0, continuing stirring for 1 hour, and dripping anhydrous ethanol at a speed of 4 mL / min; after desolvation is completed, slowly adding 0.8 mL of the black active ester solution for cross-linking and curing, continuing to react overnight under light-proof conditions at 37°C to obtain a stable drug-loaded protein magnetic nanoparticle suspension; rotary evaporating the obtained drug-loaded protein magnetic nanoparticle suspension at 37°C to remove ethanol, centrifuging at 10000 rpm for 15 minutes, and freeze-drying to obtain drug-loaded bovine serum albumin magnetic nanoparticles.

[0021] Furthermore, in the above-mentioned method for preparing the azo-functionalized magnetic nanoparticle drug-loaded protein, the amount of the azo-functionalized magnetic nanoparticles is 30 mg, the amount of anhydrous dimethyl sulfoxide is 2 mL, 900 mg of bovine serum albumin powder and 30 mg of doxorubicin hydrochloride are dissolved in 30 mL of deionized water, and the amount of anhydrous ethanol is 60 mL.

[0022] Compared with the prior art, the present invention has the following beneficial effects:

[0023] 1. The carboxyl-modified magnetic nanoparticles of the present invention have modifiable functional groups, have wider applications, and are uniform in size.

[0024] 2. The magnetic particle protein of the present invention has sufficient drug loading, uniform size and good dispersibility. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 The infrared absorption spectra of (a) oleic acid-coated magnetic nanoparticles, (b) carboxyl-modified magnetic nanoparticles, and (c) azo-functionalized magnetic nanoparticles prepared in Examples 1 and 2 are shown.

[0026] Figure 2 This is a transmission electron microscope (TEM) image of the carboxyl-modified magnetic nanoparticles prepared in Example 1.

[0027] Figure 3 This is a transmission electron microscope (TEM) image of the azo-functionalized magnetic nanoparticle drug-loaded protein prepared in Example 4. DETAILED DESCRIPTION

[0028] In order to further understand the present invention, the preferred experimental scheme of the present invention is described below in conjunction with embodiments, but it should be understood that these descriptions are only for further illustrating the features and advantages of the present invention, rather than limiting the requirements of the present invention.

[0029] Example 1 Preparation of carboxyl-modified magnetic nanoparticles

[0030] 1) Synthesis of 4-((4-aminophenyl)diazenyl)benzoic acid: 13.7 g of p-aminobenzoic acid was dissolved in 30 mL of deionized water, and 33.6 mL of concentrated hydrochloric acid was added and stirred for 20 minutes. After stirring, the mixture was cooled in an ice bath. 6.9 g of sodium nitrite was weighed and dissolved in 10 mL of deionized water. The dissolved sodium nitrite solution was cooled in an ice bath and maintained at a temperature of 0-5°C. The cooled sodium nitrite solution was slowly added dropwise to the p-aminobenzoic acid solution in the ice bath and reacted for 15 minutes. The solution after the reaction was Filter with suction, retain the filtrate and store it in an ice bath; take another large beaker, dissolve 60g of sodium acetate and 21.1g of aniline sulfite salt in deionized water, maintain the solution temperature at 0-5°C, then add the filtrate dropwise into the large beaker and stir for 1 hour, adjust the pH to maintain acidity, filter with suction and dry; after drying, react with sodium hydroxide solution (15g of sodium hydroxide dissolved in 50mL of deionized water) at 90°C for 3 hours, finally adjust the solution pH to 6 in an ice bath, filter with suction and dry to obtain 4-((4-aminophenyl)diazenyl)benzoic acid.

[0031] 2) Synthesis of oleic acid-coated magnetic nanoparticles: Weigh 5.00 g of ferric chloride hexahydrate and 2.00 g of ferrous chloride tetrahydrate, dissolve them in 10 mL of deionized water in a 100 mL beaker, heat to 40° C. and stir until dissolved, cool to room temperature and set aside; add the above two solutions into a flask and set the temperature to 70° C., while nitrogen is introduced and 10 mL of concentrated ammonia water is slowly added dropwise under mechanical stirring, and anhydrous ethanol mixed with 1 mL of oleic acid is quickly added, and the reaction is continued for 1 hour; after the reaction is completed, wash with anhydrous ethanol and deionized water three times, separate the magnetic particles with a magnet, and dry in a vacuum drying oven at 50° C. to obtain oleic acid-coated magnetic nanoparticles.

[0032] 3) Synthesis of carboxyl-modified magnetic nanoparticles: 10 mg of oleic acid-coated magnetic nanoparticles and 40 mg of 4-vinylbenzoic acid were weighed and added to deionized water, and then ultrasonically dispersed and placed in a water bath for 3 hours; 8 mg of potassium persulfate was weighed and dissolved in 10 mL of deionized water, and slowly added to the magnetic nanoparticle solution along the wall of the beaker, and the polymerization reaction was continued to proceed. The reaction was stopped after 5 hours, and then the product separated by a magnet was alternately washed with anhydrous ethanol and deionized water, and the washed magnetic nanoparticles were placed in a vacuum drying oven at 60° C. and dried for 2 hours to obtain carboxyl-modified magnetic nanoparticles.

[0033] Example 2 Preparation of azo-functionalized magnetic nanoparticles

[0034] Synthesis of azo-functionalized magnetic nanoparticles: 30 mg of carboxyl-modified magnetic nanoparticles were weighed into a three-necked flask, DMF was added as a reaction solvent, and 0.12 g of 4-((4-aminophenyl)diazenyl)benzoic acid prepared in step 1) of Example 1 was weighed into the three-necked flask and mechanically stirred until completely dissolved. After complete dissolution, catalysts 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 4-dimethylaminopyridine were added to the reaction solution, and the reaction system was stirred at 36° C. for 48 hours to obtain azo-functionalized magnetic nanoparticles.

[0035] The magnetic nanoparticles were placed in a vacuum drying oven at 60° C. and dried for 2 hours to obtain carboxyl-modified magnetic nanoparticles.

[0036] Figure 1 The infrared absorption spectra of (a) oleic acid-coated magnetic nanoparticles, (b) carboxyl-modified magnetic nanoparticles, and (c) azo-functionalized magnetic nanoparticles prepared in Examples 1 and 2 are shown. Figure 1 It can be seen that (b) curve shows that carboxyl groups are added to the magnetic nanoparticles, and (c) curve shows that the appearance of azo peak means that azo-modified magnetic particles are successfully synthesized.

[0037] Figure 2 This is a transmission electron microscope (TEM) image of the carboxyl-modified magnetic nanoparticles prepared in Example 1. Figure 2 It can be seen that the magnetic particles are uniform in size.

[0038] Example 3 Preparation of azo-functionalized magnetic nanoparticle carrier protein

[0039] 30 mg of the azo functionalized magnetic nanoparticles prepared in Example 2 were weighed into a flask, 2 mL of anhydrous dimethyl sulfoxide was added thereto, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide were added under mechanical stirring until they were completely dissolved, and the mixture was reacted at 37° C. for 2 hours under light-proof conditions to obtain a black active ester solution for standby use; at the same time, 900 mg of bovine serum albumin powder was weighed and dissolved in 30 mL of deionized water, 0.1 mol / L of sodium hydroxide aqueous solution was added dropwise, the pH value was adjusted to 9.0, stirring was continued for 1 hour, and 60 mL of anhydrous ethanol was added dropwise at a rate of 4 mL / min; after desolvation was completed, 0.8 mL of the black active ester solution was slowly added for cross-linking and curing, and the reaction was continued overnight at 37° C. under light-proof conditions to obtain a stable albumin magnetic nanoparticle suspension; the obtained albumin magnetic nanoparticle suspension was rotary evaporated at 37° C. to remove ethanol, centrifuged at 10000 rpm for 15 min, and freeze-dried to obtain bovine serum albumin magnetic nanoparticles.

[0040] Example 4 Preparation of azo-functionalized magnetic nanoparticles loaded with drug protein

[0041] Weigh 30 mg of the azo functionalized magnetic nanoparticles prepared in Example 2 into a flask, add 2 mL of anhydrous dimethyl sulfoxide thereto, add 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide under mechanical stirring until they are completely dissolved, react at 37° C. for 2 hours under light-proof conditions to obtain a black active ester solution for use; at the same time, weigh 900 mg of bovine serum albumin powder and 30 mg of doxorubicin hydrochloride and dissolve them in 30 mL of deionized water, add 0.1 mol / L sodium hydroxide aqueous solution dropwise, and adjust the pH value to 9.0, continue stirring for 1 hour, and add 60 mL of anhydrous ethanol at a rate of 4 mL / min; after desolvation is completed, slowly add 0.8 mL of black active ester solution for cross-linking and curing, and continue to react overnight at 37° C. in the dark to obtain a stable drug-loaded protein magnetic nanoparticle suspension; the obtained drug-loaded protein magnetic nanoparticle suspension is rotary evaporated at 37° C. to remove ethanol, centrifuged at 10000 rpm for 15 min, and freeze-dried to obtain drug-loaded bovine serum albumin magnetic nanoparticles, which have good drug encapsulation efficiency and an encapsulation rate of about 95%.

[0042] Figure 3 This is a transmission electron microscope (TEM) image of the azo-functionalized magnetic nanoparticle drug-loaded protein obtained in Example 4. Figure 3 It can be seen that the protein is evenly wrapped around the magnetic particles.

[0043] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some or all of the technical features thereof may be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. An azo-functionalized magnetic nanoparticle, characterized in that: The preparation method thereof comprises the following steps: 1) Synthesis of 4-((4-aminophenyl)diazenyl)benzoic acid: dissolve p-aminobenzoic acid in deionized water, add concentrated hydrochloric acid and stir for 20 minutes, and cool in an ice bath after stirring; weigh sodium nitrite and dissolve in deionized water, cool the dissolved sodium nitrite solution in an ice bath, and keep the temperature at 0-5°C; slowly drop the cooled sodium nitrite solution into the p-aminobenzoic acid solution in the ice bath, and react for 15 minutes; filter the solution after the reaction, retain the filtrate and store it in an ice bath; take another large beaker, dissolve sodium acetate and aniline sulfite in deionized water, keep the solution temperature at 0-5°C, then drop the filtrate into the large beaker and stir for 1 hour, adjust the pH to keep the solution acidic, filter and dry; after drying, react with sodium hydroxide solution at 90°C for 3 hours, finally adjust the solution pH to 6 in an ice bath, filter and dry, and obtain 4-((4-aminophenyl)diazenyl)benzoic acid; 2) Synthesis of oleic acid-coated magnetic nanoparticles: Dissolve ferric chloride hexahydrate and ferrous chloride tetrahydrate in deionized water in a beaker, heat to 40°C and stir until dissolved, cool to room temperature and set aside; add the two solutions to a flask and set the temperature to 70°C, while nitrogen is introduced and concentrated ammonia is slowly added dropwise under mechanical stirring, and anhydrous ethanol mixed with oleic acid is quickly added, and the reaction is continued for 1 hour; after the reaction is completed, wash the magnetic particles three times with anhydrous ethanol and deionized water and then with a magnet, and dry them in a vacuum drying oven at 50°C to obtain oleic acid-coated magnetic nanoparticles; 3) Synthesis of carboxyl-modified magnetic nanoparticles: oleic acid-coated magnetic nanoparticles and 4-vinylbenzoic acid were added to deionized water, ultrasonically dispersed, and placed in a water bath for 3 hours; potassium persulfate was dissolved in deionized water, and slowly added to the magnetic nanoparticle solution along the wall of the beaker, and the polymerization reaction was continued to proceed. The reaction was stopped after 5 hours, and the product separated by a magnet was washed alternately with anhydrous ethanol and deionized water, and the washed magnetic nanoparticles were placed in a vacuum drying oven at 60°C for 2 hours to obtain carboxyl-modified magnetic nanoparticles; 4) Synthesis of azo functionalized magnetic nanoparticles: Carboxyl modified magnetic nanoparticles were placed in a three-necked flask, and DMF was added as a reaction solvent. The 4-((4-aminophenyl)diazenyl)benzoic acid prepared in step 1) was mechanically stirred in the three-necked flask until it was completely dissolved. After complete dissolution, catalysts 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 4-dimethylaminopyridine were added to the reaction solution. The reaction system was stirred at 36° C. for 48 hours to obtain azo functionalized magnetic nanoparticles.

2. The azo-functionalized magnetic nanoparticle according to claim 1, characterized in that: In step 1), the amount of p-aminobenzoic acid is 13.7 g, the amount of concentrated hydrochloric acid is 33.6 mL, the amount of sodium nitrite is 6.9 g, the amount of aniline sulfite is 21.1 g, the amount of sodium acetate is 60 g, and the amount of sodium hydroxide is 15 g.

3. The azo-functionalized magnetic nanoparticle according to claim 1, characterized in that: In step 2), 5.00 g of ferric chloride hexahydrate and 2.00 g of ferrous chloride tetrahydrate were dissolved in 10 mL of deionized water respectively.

4. The azo-functionalized magnetic nanoparticle according to claim 1, characterized in that: In step 2), the amount of concentrated ammonia water is 10 mL; the amount of oleic acid is 1 mL.

5. The azo-functionalized magnetic nanoparticle according to claim 1, characterized in that: In step 3), the amount of oleic acid-coated magnetic nanoparticles is 10 mg, the amount of 4-vinylbenzoic acid is 40 mg, and the amount of potassium persulfate is 8 mg.

6. The azo-functionalized magnetic nanoparticle according to claim 1, characterized in that: In step 4), the amount of the carboxyl-modified magnetic nanoparticles used is 30 mg, and the amount of 4-((4-aminophenyl)diazenyl)benzoic acid used is 0.12 g.

7. A method for preparing azo-functionalized magnetic nanoparticle bovine serum albumin, characterized in that: The method comprises the following steps: taking the azo functionalized magnetic nanoparticles according to claim 1 into a flask, adding anhydrous dimethyl sulfoxide thereto, adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide under mechanical stirring until they are completely dissolved, reacting at 37°C for 2 hours under light-proof conditions to obtain a black active ester solution for standby use; taking bovine serum albumin powder and dissolving it in deionized water, dripping 0.1 mol / L sodium hydroxide aqueous solution, adjusting the pH value to 9.0, continuing stirring for 1 hour, and dripping anhydrous ethanol at a speed of 4 mL / min; after desolvation is completed, slowly adding 0.8 mL of the black active ester solution for cross-linking and curing, continuing to react overnight under light-proof conditions at 37°C to obtain a stable albumin magnetic nanoparticle suspension; removing ethanol from the obtained albumin magnetic nanoparticle suspension by rotary evaporation at 37°C, centrifuging at 10000 rpm for 15 minutes, and freeze-drying to obtain bovine serum albumin magnetic nanoparticles.

8. The preparation method according to claim 7, characterized in that: The amount of the azo functionalized magnetic nanoparticles used is 30 mg, the amount of anhydrous dimethyl sulfoxide used is 2 mL, 900 mg of bovine serum albumin powder is dissolved in 30 mL of deionized water, and the amount of anhydrous ethanol used is 60 mL.

9. A method for preparing azo-functionalized magnetic nanoparticles loaded with drug protein, characterized in that: The method comprises the following steps: taking the azo functionalized magnetic nanoparticles according to claim 1 into a flask, adding anhydrous dimethyl sulfoxide thereto, adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide under mechanical stirring until they are completely dissolved, reacting at 37°C for 2 hours under light-proof conditions to obtain a black active ester solution for standby use; simultaneously taking bovine serum albumin powder and doxorubicin hydrochloride and dissolving them in deionized water, dripping 0.1 mol / L sodium hydroxide aqueous solution, adjusting the pH value to 9.0, continuing stirring for 1 hour, and dripping anhydrous ethanol at a speed of 4 mL / min; after desolvation is completed, slowly adding 0.8 mL of the black active ester solution for cross-linking and curing, continuing to react overnight under light-proof conditions at 37°C to obtain a stable drug-loaded protein magnetic nanoparticle suspension; rotary evaporating the obtained drug-loaded protein magnetic nanoparticle suspension at 37°C to remove ethanol, centrifuging at 10000 rpm for 15 minutes, and freeze-drying to obtain drug-loaded bovine serum albumin magnetic nanoparticles.

10. The preparation method according to claim 9, characterized in that: The dosage of the azo functionalized magnetic nanoparticles is 30 mg, the dosage of anhydrous dimethyl sulfoxide is 2 mL, 900 mg of bovine serum albumin powder and 30 mg of doxorubicin hydrochloride are dissolved in 30 mL of deionized water, and the dosage of anhydrous ethanol is 60 mL.

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