Preparation and application of polyrotaxane nanoparticles capable of enhancing sonodynamic performance of tetraphenylporphyrin

Abstract

The invention relates to a preparation method and application of polyrotaxane nanoparticles for enhancing the sonodynamic performance of tetraphenylporphyrin, and can effectively solve the problems of low active oxygen yield and poor sonodynamic treatment effect caused by aggregation-induced quenching due to poor water solubility and easy pi-pi stacking of tetraphenylporphyrin. The material is tetraphenylporphyrin, polyethylene glycol and alpha-cyclodextrin. According to the technical scheme, tetraphenylporphyrin is modified through alpha-cyclodextrin, water solubility and biocompatibility are improved, and meanwhile, polyrotaxane is constructed through supramolecular host-guest combination of alpha-cyclodextrin and polyethylene glycol and folic acid end capping. Secondly, polyrotaxane, glucose oxidase, manganese dioxide and other nano-enzymes jointly construct polyrotaxane nano-particles; the nanoparticles have the functions of enhancing the sonodynamic performance of tetraphenylporphyrin and remarkable anti-tumor performance, the raw materials are safe, non-toxic, low in cost and easy to obtain, and the nano delivery system has the advantages of being easy to prepare, good in stability and the like and has good application prospects in the sonodynamic anti-tumor aspect.

Description

Technical Field

[0001] The invention belongs to the field of advanced nanocomposite materials and biomedicine, and specifically relates to the preparation and application of polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic properties. Background Art

[0002] Cancer treatment has always been a problem that needs to be solved urgently around the world. Sonodynamic therapy (SDT), as an emerging non-invasive tumor treatment mode, mainly activates sonosensitizers by ultrasound to produce reactive oxygen species (ROS), induce tumor cell apoptosis and autophagy, and thus efficiently kill tumor cells, and has made remarkable achievements in tumor treatment. Sonosensitizers are one of the core factors affecting sonosensitizers, and tetraphenylporphyrin, as a new generation of photo / sonosensitizers, has shown potential application value in sonosensitizers. However, due to its poor water solubility and the easy π-π stacking effect of its planar structure, it leads to the aggregation induced quenching (AIQ) effect, which affects the production of ROS and seriously limits its application in the biomedical field. At present, physical encapsulation or polymerization of amphiphilic compounds is commonly used to inhibit the AIQ effect of tetraphenylporphyrin sonosensitizers. However, physical encapsulation methods often have disadvantages such as poor water solubility and insufficient biosafety; the compounds in the polymerization reaction are usually polymer materials, which have certain cytotoxicity and poor biocompatibility problems, and the polymerization reaction involves a relatively complex reaction process.

[0003] Cyclodextrin polyrotaxane is a typical flexible mechanical interlocking molecular form, which is characterized by the free sliding of the ring molecule (cyclodextrin) on the axis chain and the large steric group end-capping, resulting in a high degree of conformational freedom and permanent spatial association. With the in-depth understanding of the supramolecular induced self-assembly process of cyclodextrin polyrotaxane, the application of cyclodextrin polyrotaxane in elastomers / gels, battery electrodes and drug delivery is becoming more and more extensive. Especially in drug delivery, cyclodextrin polyrotaxane has been widely studied in tumor treatment, rheumatoid arthritis, wound healing and other fields due to its advantages such as controllable structure, non-toxicity and good biocompatibility. Therefore, the use of non-toxic natural biomaterials to construct cyclodextrin polyrotaxane improves the water solubility of the sonosensitizer tetraphenylporphyrin, inhibits the π-π stacking effect and aggregation-induced effect, and enhances the targeting, providing potential drugs for anti-tumor, which is of great significance for expanding the anti-tumor treatment strategy based on sonodynamics. Summary of the invention

[0004] The primary purpose of the present invention is to overcome the deficiencies of the prior art and provide a method for preparing polyrotaxane nanoparticles with enhanced sonodynamic properties of tetraphenylporphyrin. The method first selects natural non-toxic cyclodextrin to improve the poor water solubility and insufficient biocompatibility of the sonosensitizer tetraphenylporphyrin. Secondly, polyethylene glycol and cyclodextrin-based monomers are self-assembled, and folic acid is capped to obtain polyrotaxane, which inhibits the aggregation induction effect caused by the π-π stacking effect of tetraphenylporphyrin through steric effects, while giving the polyrotaxane the ability of targeted recognition. Finally, polyrotaxane nanoparticles are prepared by electrostatic adsorption loading of glucose oxidase and manganese dioxide nanozymes prepared by oleic acid method to improve the hypoxic microenvironment of tumors.

[0005] Another object of the present invention is to obtain polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic performance through the above preparation method. The nanoparticles have good biocompatibility, can effectively enhance the generation of active oxygen, and show important application prospects in sonodynamic treatment of tumors.

[0006] Another object of the present invention is to provide the application of the above-mentioned polyrotaxane nanoparticles with enhanced sonodynamic properties of tetraphenylporphyrin, especially in the sonodynamic treatment of deep cancers such as breast cancer, liver cancer, lung cancer, pancreatic cancer, etc.

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

[0008] A preparation method of polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic performance comprises the following steps:

[0009] (1) Preparation of cyclodextrin-modified tetraphenylporphyrin: tetraphenylporphyrin, mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin, and potassium carbonate are dissolved in N,N-dimethylformamide and heated for reaction; then, the mixture is dialyzed with distilled water, filtered to remove the solvent and unreacted monomers, and dried to obtain cyclodextrin-modified tetraphenylporphyrin (CD-TPP);

[0010] (2) Preparation of charge reversal monomer: Mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin, 3-dimethylaminopropylamine and sodium carbonate were dissolved in distilled water, heated to react, and reaction system B was obtained. After dialysis with distilled water, suction filtration was performed to remove the solvent and unreacted monomers, and the monomer (CD-N) was obtained. Then, the monomer (CD-N) and 4-(bromomethyl)phenylboronic acid were dissolved in N,N-dimethylformamide, heated to react, ether precipitated, centrifuged, and dried to obtain a charge reversal monomer (CD-NB);

[0011] (3) Synthesis of polyrotaxane: Dissolve the cyclodextrin-modified tetraphenylporphyrin (CD-TPP) and charge reversal monomer (CD-NB) obtained in steps (1) and (2) in distilled water, then add double-terminal amino functionalized polyethylene glycol PEG-(NH2)2, react, separate the solid and liquid, and dry the obtained solid to obtain a pseudorotaxane. Dissolve folic acid, N-hydroxysuccinimide and 1-ethyl-3[3-dimethylaminopropyl]carbodiimide hydrochloride in N,N-dimethylformamide, react, add the obtained pseudorotaxane, dialyze with distilled water, and dry to obtain a polyrotaxane (PR);

[0012] (4) Synthesis of polyrotaxane nanoparticles: The polyrotaxane PR obtained in step (3) was dissolved in dimethyl sulfoxide and added dropwise to distilled water. Then, glucose oxidase and manganese dioxide nanozyme prepared by oleic acid method were added dropwise to the solution. The reaction was carried out to prepare polyrotaxane nanoparticles.

[0013] The tetraphenylporphyrin in step (1) is preferably tetraphenylporphyrin substituted with 1 to 4 hydroxyl groups at the para position.

[0014] The tetraphenylporphyrin, mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin and potassium carbonate in step (1) are used in a molar ratio of 1:0.25-4:2.5-40;

[0015] The amount of N,N-dimethylformamide used in step (1) is calculated based on dissolving 300-1200 mg of tetraphenylporphyrin per 10 mL;

[0016] As described in step (1), the heating reaction conditions are 90-120° C. constant temperature magnetic stirring reaction for 1-3 days; more preferably, N2 protection is used to react at 90° C. constant temperature magnetic stirring reaction for 1-3 days.

[0017] In step (1), the distilled water dialysis condition is to use a dialysis bag of 500-1000Da for 1-5 days; more preferably, to use a dialysis bag of 500Da for 1-5 days.

[0018] The mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin in steps (1) and (2) is preferably a mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin containing 6 to 8 D-pyranose glucose units; more preferably a mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin containing 6 to 7 D-pyranose glucose units.

[0019] The monomer (CD-N), sodium carbonate and 3-dimethylaminopropylamine in step (2) are used in a molar ratio of 1:5-30:0.5-3;

[0020] The mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin and 4-(bromomethyl)phenylboronic acid in step (2) are used in a molar ratio of 1:(0.5-2.5);

[0021] The amount of distilled water used in step (2) is calculated by adding 0.5 to 5 mL of 3-dimethylaminopropylamine per 10 mL;

[0022] The amount of N,N-dimethylformamide used in step (2) is preferably calculated by adding 100-600 mg of 4-(bromomethyl)phenylboronic acid per 10 mL; the amount of ether used and N,N-dimethylformamide are in a volume ratio of 0.01-0.1:10.

[0023] In step (2), the heating reaction conditions for preparing system B are 80-120°C constant temperature magnetic stirring reaction for 12-36 hours; more preferably 110°C constant temperature magnetic stirring reaction for 1-3 days.

[0024] As described in step (2), the heating reaction conditions for preparing the charge reversal monomer (CD-NB) are 25-70° C. constant temperature magnetic stirring reaction for 12-36 hours; more preferably 60° C. constant temperature magnetic stirring reaction for 12-36 hours.

[0025] In step (2), the conditions for collecting the charge-reversed monomers by centrifugation are 1000-4000 rpm for 5-10 min; more preferably, centrifugation is performed at 1000 rpm for 5-10 min.

[0026] The double-terminal amino-functionalized polyethylene glycol in step (3) is preferably a double-terminal amino-functionalized polyethylene glycol with a molecular weight distribution of 1000 to 4000.

[0027] The amount of distilled water used in step (3) is preferably calculated based on 1 to 10 g of double-terminal amino-functionalized polyethylene glycol per 10 mL.

[0028] The amounts of the components in step (3) are calculated according to the ratio of cyclodextrin-modified tetraphenylporphyrin (CD-TPP): charge reversal monomer (CD-NB): double-terminal amino-functionalized polyethylene glycol: folic acid: N-hydroxysuccinimide and 1-ethyl-3[3-dimethylaminopropyl]carbodiimide hydrochloride = 0.1-2:0.1-2:1:2-10:2-15:3-20.

[0029] As described in step (3), the reaction conditions for preparing the pseudorotaxane are constant temperature magnetic stirring at 15-35° C. for 4-12 hours.

[0030] The solid-liquid separation method is preferably centrifugal, and the centrifugal conditions are 1000-4000 rpm and the time is 5-10 min; more preferably, centrifugation is performed at 1000 rpm for 5-10 min.

[0031] The amount of N,N-dimethylformamide used in step (3) is preferably calculated based on 0.1 to 1 g of folic acid per 10 mL.

[0032] As described in step (3), the reaction conditions for preparing polyrotaxane (PR) are 15-35°C constant temperature magnetic stirring reaction for 4-12 hours; the preferred conditions are 25-37°C, magnetic stirring reaction for 12 hours.

[0033] As described in step (3), the dialysis conditions for preparing the rotaxane by reaction are to use a dialysis bag of 500 to 2000 Da for 1 to 5 days.

[0034] The amount of distilled water used in step (4) is calculated based on the addition of 0.1 to 5 mg of polyrotaxane (PR) per 10 mL.

[0035] The amount of dimethyl sulfoxide used in step (4) is calculated based on the addition of 0.1 to 5 mg of polyrotaxane (PR) per 1 mL.

[0036] The amount of each component in step (4) is calculated according to the ratio of polyrotaxane: glucose oxidase: manganese dioxide nanozyme prepared by oleic acid method = 1:0.1~1:0.1~1.

[0037] As described in step (4), the reaction conditions for preparing polyrotaxane nanoparticles are 15-35° C. and magnetic stirring for 12-24 hours; the preferred conditions are 25-35° C. and magnetic stirring for 24 hours.

[0038] The drying in step (1) preparing cyclodextrin-modified tetraphenylporphyrin (CD-TPP), step (2) preparing monomer (CD-N), and step (3) preparing polyrotaxane (PR) is preferably freeze-drying.

[0039] The drying in step (2) preparing the charge reversal monomer (CD-NB) and step (3) preparing the pseudorotaxane is preferably vacuum drying.

[0040] The reaction temperature in the present invention is room temperature unless otherwise specified, and the room temperature is preferably 15 to 35°C.

[0041] A polyrotaxane nanoparticle with enhanced tetraphenylporphyrin sonodynamic performance is obtained by the preparation method.

[0042] The above-mentioned polyrotaxane nanoparticles with enhanced sonodynamic properties of tetraphenylporphyrin are used in sonodynamic therapy of tumors.

[0043] The present invention has the following advantages and effects over the prior art:

[0044] (1) The present invention uses cyclodextrin-modified tetraphenylporphyrin and double-terminal amino-functionalized polyethylene glycol as the core, and the cyclodextrin-modified porphyrin can improve the water solubility of the sonosensitizer tetraphenylporphyrin.

[0045] (2) The present invention provides a method for utilizing the steric effect of polyrotaxane and cyclodextrin to inhibit the π-π stacking caused by the planar stacking of the sonosensitizer tetraphenylporphyrin, thereby weakening the aggregation induction effect, thereby greatly improving the active oxygen production and sonodynamic performance of tetraphenylporphyrin.

[0046] (3) The polyrotaxane prepared by the present invention with enhanced sonodynamic performance of tetraphenylporphyrin can be used to prepare polyrotaxane nanoparticles by electrostatically adsorbing negatively charged nanozymes through enhanced positive charges, thereby achieving enhanced sonodynamic performance and having a wide range of applications in the field of sonodynamic tumor treatment.

[0047] (4) The polyrotaxane nanoparticles prepared by the present invention with enhanced sonodynamic properties of tetraphenylporphyrin have good biocompatibility and anti-tumor effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1 The following is a flow chart for preparing polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic properties in Example 1:

[0049] Figure 2 For the implementation of α-cyclodextrin modified tetraphenylporphyrin (α-CD-TPP) in Example 1 1 H-NMR nuclear magnetic characterization spectrum;

[0050] Figure 3 For the charge reversal monomer (α-CD-NB) in Example 1 1 H-NMR nuclear magnetic characterization spectrum;

[0051] Figure 4 For the implementation of polyrotaxane (PR) in Example 1 1 H-NMR nuclear magnetic characterization spectrum;

[0052] Figure 5 FT-IR characterization diagram of α-cyclodextrin modified tetraphenylporphyrin (α-CD-TPP) in implementation case 1;

[0053] Figure 6 is a graph showing the nanoparticle size of polyrotaxane nanoparticles in implementation example 1;

[0054] Figure 7 is a graph showing the change in dissolved oxygen content (OD) of the polyrotaxane nanoparticle solution in Example 1;

[0055] Figure 8 Zeta potential diagram of different materials in implementation case 1;

[0056] Fig. 9 This is a comparison diagram of singlet oxygen generation of polyrotaxane nanoparticles and tetraphenylporphyrin under ultrasound conditions by ultraviolet monitoring in Example 1;

[0057] FIG10( a ) shows the inhibition rate of polyrotaxane nanoparticles on 4T1 cells before ultrasound in Example 1;

[0058] FIG10( b ) shows the inhibition rate of polyrotaxane nanoparticles on 4T1 cells after ultrasound in Example 1;

[0059] Fig.11 The tumor inhibition rate of polyrotaxane nanoparticles on 4T tumor-bearing mice in implementation case 1;

[0060] Fig.12 This is the effect of polyrotaxane nanoparticles on the body weight of 4T tumor-bearing mice in implementation case 1. DETAILED DESCRIPTION

[0061] The present invention is further described in detail below in conjunction with the accompanying drawings and embodiments, but the accompanying drawings and embodiments are not intended to limit the technical solutions of the present invention, and all changes or equivalent substitutions made based on the teachings of the present invention fall within the protection scope of the present invention.

[0062] Example 1

[0063] The method for preparing polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic performance in this embodiment comprises the following steps:

[0064] (1) Preparation of α-cyclodextrin-modified tetraphenylporphyrin: 5-(4-hydroxyphenyl-10,15,20-triphenylporphyrin (757 mg, 1.2 mmol), mono-(6-deoxy-6-toluenesulfonyl)-α-cyclodextrin (1.15 g, 1 mmol), and potassium carbonate (1.382 g, 10 mmol) were dissolved in 15 mL of N,N-dimethylformamide, and heated at 90° C. for 3 days under a nitrogen atmosphere; then, the mixture was dialyzed with distilled water using a dialysis bag with a molecular weight cutoff of 500 Da for 3 days, filtered to remove the solvent and unreacted monomers, and dried to obtain α-cyclodextrin-modified tetraphenylporphyrin (α-CD-TPP);

[0065] (2) Preparation of charge reversal monomer: Mono-(6-deoxy-6-toluenesulfonyl)-α-cyclodextrin (1.15 g, 1.0 mmol), 3-dimethylaminopropylamine (124 μL, 1.5 mmol) and sodium carbonate (212 mg, 2.0 mmol) were dissolved in distilled water, and the mixture was refluxed at 110°C for 24 h to obtain reaction system B. The reaction system was dialyzed with distilled water for 3 days using a dialysis bag with a molecular weight cutoff of 500 Da. The solvent and unreacted monomers were removed by suction, and the mixture was freeze-dried to obtain a monomer (α-CD-N). Next, the monomer (α-CD-N) (2.144 g, 2.0 mmol) and 4-(bromomethyl)phenylboronic acid (644.46 mg, 3.0 mmol) were dissolved in 20 mL of N,N-dimethylformamide, heated at 50 °C for 24 h under a nitrogen atmosphere, precipitated with ether, centrifuged at 3000 rpm for 5 min, and dried in vacuum to obtain a charge-reversed monomer (α-CD-NB);

[0066] (3) Synthesis of polyrotaxane: α-cyclodextrin-modified tetraphenylporphyrin (α-CD-TPP) (481 mg, 0.3 mmol) and charge reversal monomer (α-CD-NB) (226 mg, 0.2 mmol) obtained in steps (1) and (2) were dissolved in 5 mL of water, and then double-terminal amino-functionalized polyethylene glycol (molecular weight Mw=2000) (508 mg, 0.22 mmol) was added thereto. The reaction was carried out at room temperature for 24 h, and the reaction was centrifuged at 5000 rpm for 5 min. The obtained solid was vacuum dried to obtain a pseudorotaxane. (441 mg, 1.0 mmol) of folic acid, (576 mg, 5.0 mmol) of N-hydroxysuccinimide and (986 mg, 5.0 mmol) of 1-ethyl-3[3-dimethylaminopropyl]carbodiimide hydrochloride were dissolved in N,N-dimethylformamide for reaction, and the obtained pseudorotaxane (100 mg) was added thereto, and the mixture was dialyzed with distilled water for 3 days using a dialysis bag with a molecular weight cutoff of 1000 Da, and freeze-dried to obtain a polyrotaxane (PR);

[0067] (4) Synthesis of polyrotaxane nanoparticles: 5 mg of polyrotaxane (PR) obtained in step (3) was dissolved in dimethyl sulfoxide and added dropwise to 5 mL of distilled water. Then, 200 μL of glucose oxidase solution (5 mg / mL) and 200 μL of manganese dioxide nanozyme prepared by oleic acid method were added dropwise to the solution. The reaction was stirred under magnetic stirring at room temperature for 24 h to prepare polyrotaxane nanoparticles.

[0068] Example 2

[0069] The method for preparing polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic performance in this embodiment comprises the following steps:

[0070] (1) Preparation of α-cyclodextrin-modified tetraphenylporphyrin: 5,15-(4-hydroxyphenyl-10,20-triphenylporphyrin (746 mg, 1.0 mmol), mono-(6-deoxy-6-toluenesulfonyl)-α-cyclodextrin (3.45 g, 3.0 mmol), and potassium carbonate (2.77 g, 20 mmol) were dissolved in 20 mL of N,N-dimethylformamide, and heated at 90° C. for 3 days under a nitrogen atmosphere; then, the mixture was dialyzed with distilled water for 3 days using a dialysis bag with a molecular weight cutoff of 1000 Da, and the solvent and unreacted monomers were removed by suction filtration, and the mixture was dried to obtain α-cyclodextrin-modified tetraphenylporphyrin (α-CD-TPP);

[0071] Steps (2), (3), and (4) are the same as those in implementation case 1.

[0072] Example 3

[0073] The method for preparing polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic performance in this embodiment comprises the following steps:

[0074] (1) Preparation of β-cyclodextrin-modified tetraphenylporphyrin: 5-(4-hydroxyphenyl-10,15,20-triphenylporphyrin (757 mg, 1.2 mmol), mono-(6-deoxy-6-toluenesulfonyl)-β-cyclodextrin (1.29 g, 1 mmol), and potassium carbonate (1.382 g, 10 mmol) were dissolved in 15 mL of N,N-dimethylformamide, and heated at 90°C for 3 days under a nitrogen atmosphere; then, the mixture was dialyzed with distilled water for 3 days using a dialysis bag with a molecular weight cutoff of 1000 Da, and the solvent and unreacted monomers were removed by suction filtration, and then dried to obtain β-cyclodextrin-modified tetraphenylporphyrin (β-CD-TPP);

[0075] (2) Preparation of charge reversal monomer: Mono-(6-deoxy-6-toluenesulfonyl)-β-cyclodextrin (1.29 g, 1.0 mmol), 3-dimethylaminopropylamine (124 μL, 1.5 mmol) and sodium carbonate (212 mg, 2.0 mmol) were dissolved in distilled water and refluxed at 110°C for 24 h to obtain reaction system B. The system was dialyzed with distilled water for 3 days using a dialysis bag with a molecular weight cutoff of 500 Da. The solvent and unreacted monomers were removed by suction and the system was freeze-dried to obtain a monomer (β-CD-N). Next, the monomer (β-CD-N) (2.28 g, 2.0 mmol) and 4-(bromomethyl)phenylboronic acid (644.46 mg, 3.0 mmol) were dissolved in 20 mL of N,N-dimethylformamide, heated at 60 °C for 24 h under a nitrogen atmosphere, precipitated with ether, centrifuged at 3000 rpm for 5 min, and dried in vacuum to obtain a charge-reversed monomer (β-CD-NB);

[0076] Steps (3) and (4) are the same as those in implementation case 1.

[0077] Example 4

[0078] The method for preparing polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic performance in this embodiment comprises the following steps:

[0079] Steps (1) and (2) are the same as those in implementation case 1.

[0080] (3) Synthesis of polyrotaxane: α-cyclodextrin-modified tetraphenylporphyrin (α-CD-TPP) (481 mg, 0.3 mmol) and charge reversal monomer (α-CD-NB) (226 mg, 0.2 mmol) obtained in steps (1) and (2) were dissolved in 6 mL of water, and then double-terminal amino-functionalized polyethylene glycol (molecular weight Mw=1000) (508 mg, 0.22 mmol) was added thereto. The reaction was carried out at room temperature for 24 h, and the reaction was centrifuged at 5000 rpm for 5 min. The obtained solid was dried to obtain a pseudorotaxane. (441 mg, 1.0 mmol) of folic acid, (576 mg, 5.0 mmol) of N-hydroxysuccinimide and (986 mg, 5.0 mmol) of 1-ethyl-3[3-dimethylaminopropyl]carbodiimide hydrochloride were dissolved in N,N-dimethylformamide for reaction, and the obtained pseudorotaxane (100 mg) was added thereto, and the mixture was dialyzed with distilled water for 3 days using a dialysis bag with a molecular weight cutoff of 1000 Da, and freeze-dried to obtain a polyrotaxane (PR);

[0081] Step (4) is the same as in implementation case 1.

[0082] Example 5

[0083] The method for preparing polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic performance in this embodiment comprises the following steps:

[0084] Steps (1) and (2) are the same as those in implementation case 1.

[0085] (3) Synthesis of polyrotaxane: α-cyclodextrin-modified tetraphenylporphyrin (α-CD-TPP) (321 mg, 0.1 mmol) and charge reversal monomer (α-CD-NB) (113 mg, 0.1 mmol) obtained in steps (1) and (2) were dissolved in 5 mL of water, and then double-terminal amino-functionalized polyethylene glycol (molecular weight Mw=2000) (508 mg, 0.22 mmol) was added thereto. The reaction was carried out at room temperature for 24 h, and the reaction was centrifuged at 5000 rpm for 5 min. The obtained solid was dried to obtain a pseudorotaxane. (441 mg, 1.0 mmol) of folic acid, (576 mg, 5.0 mmol) of N-hydroxysuccinimide and (986 mg, 5.0 mmol) of 1-ethyl-3[3-dimethylaminopropyl]carbodiimide hydrochloride were dissolved in N,N-dimethylformamide for reaction, and the obtained pseudorotaxane (100 mg) was added thereto, and the mixture was dialyzed with distilled water for 3 days using a dialysis bag with a molecular weight cutoff of 1000 Da, and freeze-dried to obtain a polyrotaxane (PR);

[0086] Step (4) is the same as in implementation case 1.

[0087] Example 6

[0088] The method for preparing polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic performance in this embodiment comprises the following steps:

[0089] Steps (1), (2), and (3) are the same as those in implementation case 1.

[0090] (4) Synthesis of polyrotaxane nanoparticles: 5 mg of polyrotaxane (PR) obtained in step (3) was dissolved in dimethyl sulfoxide and added dropwise to 5 mL of distilled water. Then, 400 μL of glucose oxidase solution (5 mg / mL) and 400 μL of manganese dioxide nanozyme prepared by oleic acid method were added dropwise to the solution. The reaction was carried out under magnetic stirring at room temperature for 24 h to prepare polyrotaxane nanoparticles.

[0091] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be equivalent replacement methods and are included in the protection scope of the present invention.

Claims

1. A method for preparing polyrotaxane nanoparticles having enhanced sonodynamic properties of tetraphenylporphyrin, characterized in that: The following steps are involved: (1) Preparation of cyclodextrin-modified tetraphenylporphyrin: tetraphenylporphyrin, mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin, and potassium carbonate are dissolved in N,N-dimethylformamide and heated for reaction; then, the mixture is dialyzed with distilled water, filtered to remove the solvent and unreacted monomers, and dried to obtain cyclodextrin-modified tetraphenylporphyrin (CD-TPP); (2) Preparation of charge reversal monomer: Mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin, 3-dimethylaminopropylamine and sodium carbonate were dissolved in distilled water, heated to react, and reaction system B was obtained. After dialysis with distilled water, the solvent and unreacted monomers were removed by suction filtration, and the monomer (CD-N) was obtained. Then, the monomer (CD-N) and 4-(bromomethyl)phenylboronic acid were dissolved in N,N-dimethylformamide, heated to react, ether precipitated, centrifuged, and dried to obtain a charge reversal monomer (CD-NB). (3) Synthesis of polyrotaxane: dissolve the cyclodextrin-modified tetraphenylporphyrin (CD-TPP) and charge reversal monomer (CD-NB) obtained in steps (1) and (2) in distilled water, then add double-terminal amino-functionalized polyethylene glycol PEG-(NH2)2 thereto, react, separate the solid and the liquid, and dry the obtained solid to obtain a pseudorotaxane; dissolve folic acid, N-hydroxysuccinimide and 1-ethyl-3[3-dimethylaminopropyl]carbodiimide hydrochloride in N,N-dimethylformamide, react, add the obtained pseudorotaxane thereto, dialyze with distilled water, and dry to obtain a polyrotaxane (PR); (4) Synthesis of polyrotaxane nanoparticles: The polyrotaxane (PR) obtained in step (3) is dissolved in dimethyl sulfoxide and added dropwise to distilled water; glucose oxidase and manganese dioxide nanozyme prepared by the oleic acid method are then added dropwise to the solution; and the reaction is carried out to prepare polyrotaxane nanoparticles.

2. The method for preparing polyrotaxane nanoparticles having enhanced tetraphenylporphyrin sonodynamic properties as claimed in claim 1, characterized in that: The tetraphenylporphyrin, mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin and potassium carbonate in step (1) are used in a molar ratio of 1:0.25-4:2.5-40; The monomer (CD-N), sodium carbonate and 3-dimethylaminopropylamine in step (2) are used in a molar ratio of 1:5-30:0.5-3; The mono-(6-deoxy-6-toluenesulfonyl)-cyclodextrin and 4-(bromomethyl)phenylboronic acid in step (1) are used in a molar ratio of 1:(0.5-2.5); The amounts of the components in step (3) are in the following ratios: cyclodextrin-modified tetraphenylporphyrin (CD-TPP): charge reversal monomer (CD-NB): double-terminal amino-functionalized polyethylene glycol: folic acid: N-hydroxysuccinimide and 1-ethyl-3[3-dimethylaminopropyl]carbodiimide hydrochloride = 0.1-2: 0.1-2: 1: 2-10: 2-15: 3-20; The dosage of each component in step (4) is in the ratio of polyrotaxane: glucose oxidase: manganese dioxide nanozyme prepared by oleic acid method = 1:0.1~1:0.1~1.

3. A method for preparing polyrotaxane nanoparticles having enhanced tetraphenylporphyrin sonodynamic properties as claimed in claim 1 or 2, characterized in that: The amount of N,N-dimethylformamide used in step (1) is calculated based on dissolving 300-1200 mg of sonosensitizer tetraphenylporphyrin per 10 mL; The amount of distilled water used in step (2) is calculated by adding 0.5 to 5 mL of 3-dimethylaminopropylamine per 10 mL; The amount of N,N-dimethylformamide used in step (2) is calculated by adding 100-600 mg of 4-(bromomethyl)phenylboronic acid per 10 mL; the amount of ether used is 0.01-0.1:10 by volume to N,N-dimethylformamide; The amount of distilled water used in step (3) is calculated by adding 1 to 10 g of double-terminated amino-functionalized polyethylene glycol per 10 mL; The amount of N,N-dimethylformamide used in step (3) is calculated by adding 0.1-1 g of folic acid per 10 mL; The amount of dimethyl sulfoxide used in step (4) is calculated based on the addition of 0.1 to 5 mg of polyrotaxane (PR) per 1 mL; The amount of distilled water used in step (4) is calculated based on the addition of 0.1 to 5 mg of polyrotaxane (PR) per 10 mL.

4. A method for preparing polyrotaxane nanoparticles having enhanced tetraphenylporphyrin sonodynamic properties as claimed in claim 1 or 2, characterized in that: The heating reaction in step (1) is a constant temperature magnetic stirring reaction at 90-120° C. for 1-3 days under N2 protection; The distilled water dialysis condition in step (1) is to use a 500-1000Da dialysis bag for 1-5 days; The heating reaction conditions for preparing system B in step (2) are 80-120° C. constant temperature magnetic stirring reaction for 12-36 hours; The heating reaction in step (2) to prepare the charge reversal monomer (CD-NB) is carried out under constant temperature magnetic stirring at 25-70° C. for 12-36 hours; The conditions for centrifugation to collect the charge reversal monomers in step (2) are 1000-4000 rpm and the time is 5-10 min; The conditions for preparing the pseudorotaxane in step (3) are constant temperature magnetic stirring at 15-35° C. for 4-12 hours; The reaction conditions for preparing polyrotaxane (PR) in step (3) are 15-35° C. constant temperature magnetic stirring reaction for 4-12 hours; The dialysis conditions for preparing the rotaxane by the reaction in step (3) are to use a dialysis bag of 500 to 2000 Da for 1 to 5 days; The reaction conditions for preparing polyrotaxane nanoparticles in step (4) are 15-35° C. and magnetic stirring for 12-24 hours.

5. The method for preparing polyrotaxane nanoparticles having enhanced tetraphenylporphyrin sonodynamic properties as claimed in claim 3, characterized in that: The heating reaction in step (1) is a constant temperature magnetic stirring reaction at 90-120° C. for 1-3 days under N2 protection; The distilled water dialysis condition in step (1) is to use a 500-1000Da dialysis bag for 1-5 days; The heating reaction conditions for preparing system B in step (2) are 80-120° C. constant temperature magnetic stirring reaction for 12-36 hours; The heating reaction in step (2) to prepare the charge reversal monomer (CD-NB) is carried out under constant temperature magnetic stirring at 25-70° C. for 12-36 hours; The conditions for centrifugation to collect the charge reversal monomers in step (2) are 1000-4000 rpm and the time is 5-10 min; The conditions for preparing the pseudorotaxane in step (3) are constant temperature magnetic stirring at 15-35° C. for 4-12 hours; The reaction conditions for preparing polyrotaxane (PR) in step (3) are 15-35° C. constant temperature magnetic stirring reaction for 4-12 hours; The dialysis conditions for preparing the rotaxane by the reaction in step (3) are to use a dialysis bag of 500 to 2000 Da for 1 to 5 days; The reaction conditions for preparing polyrotaxane nanoparticles in step (4) are 15-35° C. and magnetic stirring for 12-24 hours.

6. A polyrotaxane nanoparticle having enhanced sonodynamic properties of tetraphenylporphyrin, characterized in that: The method is obtained by the preparation method according to any one of claims 1 to 5.

7. The polyrotaxane nanoparticles with enhanced tetraphenylporphyrin sonodynamic properties as claimed in claim 6, characterized in that: Application in the preparation of anti-tumor drugs.

Images