CN108721704B

CN108721704B - Long-acting sustained-release cardiovascular coating material with dual functions of calcification resistance and hyperplasia resistance and preparation method thereof

Info

Publication number: CN108721704B
Application number: CN201810731158.4A
Authority: CN
Inventors: 王云兵; 陆奖; 罗日方; 庄伟华; 李高参
Original assignee: Sichuan University
Current assignee: Sichuan University
Priority date: 2018-07-05
Filing date: 2018-07-05
Publication date: 2020-03-10
Anticipated expiration: 2038-07-05
Also published as: CN108721704A

Abstract

The invention discloses a long-acting slow-release cardiovascular coating material with double functions of calcification resistance and hyperplasia resistance and a preparation method thereof; the preparation method comprises the following steps: firstly, polishing, cleaning and drying a base material to be modified and modified; then placing the treated substrate in a dopamine solution, and reacting for 0.5-2 h at 10-30 ℃; then the reacted substrate is sequentially placed into a biological macromolecule solution, a heparin solution and a biological macromolecule solution, and is respectively soaked for 10min, and then the substrate is transferred into a drug-loaded micelle to be soaked for 20 min; and finally, repeating the soaking step for 1-30 times to obtain a final finished product. By adopting the method, the cardiovascular coating material with excellent mechanical properties and capable of realizing drug slow release can be prepared, and the anti-calcification and anti-hyperplasia properties of the cardiovascular stent coating material can be effectively supported by respectively loading the anti-vascular-calcification drug and the anti-vascular smooth muscle proliferation drug in the drug-loaded micelle.

Description

Long-acting sustained-release cardiovascular coating material with dual functions of calcification resistance and hyperplasia resistance and preparation method thereof

Technical Field

The invention belongs to the technical field of medical materials, and particularly relates to a long-acting slow-release cardiovascular coating material with dual functions of calcification resistance and hyperplasia resistance and a preparation method thereof.

Background

After the cardiovascular implantable material enters a human body, the contact with tissues can generate some unfavorable interactions between the tissues and the material, when the material is implanted, endothelial injury is inevitably caused, the growth speed of smooth muscle cells is higher than that of endothelial cells, the endothelial cells cannot be repaired in time, the smooth muscle cells are proliferated, blood fat deposition, calcium deposition and the like, and restenosis of a stent implantation part is easily caused. In the traditional preparation method of the drug-loaded modified coating, because the drug loading is small and burst release is easy to occur, smooth muscle cells can be continuously increased due to the fact that the inner cortex is not repaired enough for the rapid release of the drug, so that the problem cannot be solved only by a simple surface modification method, and the drug needs to be administered in multiple time sequences for a long time so as to achieve the purpose of treatment, such as long-acting slow-release anticoagulant heparin, anti-calcified atorvastatin calcium drugs, anti-proliferative rapamycin and the like.

However, most of the two drugs have very low solubility (hydrophobic drugs) in aqueous solution, and cannot carry a large amount of drugs in a self-assembly manner under the condition of a single self-assembly solvent (such as an aqueous solvent), and if the drug-carrying coating is soaked in an organic solvent, the assembled coating is easy to damage, so that the traditional method cannot easily realize a plurality of large amounts of drugs, and moreover, the drug-carrying coating prepared by the traditional method is easy to burst, and cannot realize long-acting slow release to achieve the purpose of treatment.

Disclosure of Invention

Aiming at the prior art, the invention provides a preparation method of a long-acting slow-release cardiovascular coating material with dual functions of calcification resistance and hyperplasia resistance and the cardiovascular coating material prepared by the method, so as to solve the technical problems of low drug loading capacity and easy burst release of drugs of the hyperplasia resistance and calcification resistance of a hydrophobic coating.

In order to achieve the purpose, the invention adopts the technical scheme that: the preparation method of the long-acting sustained-release cardiovascular coating material with the double functions of calcification resistance and hyperplasia resistance is provided, and comprises the following steps:

1) mixing a polyamino compound and molecules simultaneously carrying catechol groups and aldehyde groups in a molar ratio of 1-2: 1, reacting for 1-3 h, adding sodium borohydride, stirring until no bubbles are generated, dialyzing, freeze-drying, and preparing a dried product into a solution with the concentration of 0.1-10 mg/ml and the pH value of 6;

2) preparing the negatively charged heparin into a solution with the concentration of 0.1-10 mg/ml and the pH value of 6;

3) and preparing the mixed drug-loaded micelle: mixing a hydrophilic substance and a hydrophobic substance, adding o-hydroxyphenylboronic acid into the mixture, stirring and reacting for 22-26 h, adding DCC and DMAP, continuing to react for 10-14 h, dialyzing, freeze-drying, and preparing a dried substance into a solution with the concentration of 0.1-10 mg/ml; then respectively dissolving the anticalcification drug and the antiproliferative drug in the solution, preparing drug-loaded micelles by a direct dissolution method, mixing the two drug-loaded micelles in equal volume to obtain mixed drug-loaded micelles for later use; wherein the mass ratio of the hydrophilic substance to the hydrophobic substance to the o-hydroxyphenylboronic acid to the DCC to the DMAP is 2: 1-1.5: 0.5-1;

4) polishing, cleaning and drying the base material needing modification;

5) placing the substrate treated in the step 4) into a dopamine solution with the concentration of 1-3 mg/ml, and reacting for 0.5-2 h at 10-30 ℃; the dopamine solution is prepared in a buffer system with the pH value of 7-9;

6) soaking the base material treated in the step 5) in the solution obtained in the step 1) for 10min, taking out and washing with deionized water; then placing the mixture in the solution obtained in the step 2) for soaking for 10min, taking out and washing the mixture with deionized water; then placing the mixture in the solution obtained in the step 1) for soaking for 10min, taking out and washing the mixture with deionized water; finally, placing the mixture in the medicine-carrying micelle obtained in the step 3) for soaking for 20min, taking out the mixture and cleaning the mixture by using deionized water to finish an assembly period;

7) and repeating the assembly cycle for 1-30 times to obtain the final product.

On the basis of the technical scheme, the invention can be further improved as follows.

Further, the polyamino compound is polydiallyl polyhexylpurple nitrile, dimethyl ammonium chloride, poly-L-arginine hydrochloride, polyallylamine hydrochloride, polyethyleneimine or chitosan.

Further, the molecule having both a catechol group and an aldehyde group is 3, 4-dihydroxybenzaldehyde.

Further, the hydrophilic substance is polyacrylic acid, sodium polystyrene sulfonate, chondroitin sulfate, heparin, heparan sulfate, sodium alginate, keratan sulfate, collagen, hyaluronic acid or polyglutamic acid; the hydrophobic substance is aliphatic hydrocarbon, cholesterol, polylysine or polylactic acid.

Further, the anticalcification drug is atorvastatin calcium; the antiproliferative drug is rapamycin or adriamycin.

Further, the buffer system is PBS buffer, N-Tris (hydroxymethyl) methylglycine buffer, triethanolamine buffer or Tris-HCl buffer.

Further, the substrate is a metal-based biomaterial, a ceramic-based biomaterial or a polymer-based biomaterial.

Further, the metal-based biomaterial is stainless steel, cobalt-based alloy, titanium and its alloy, magnesium and its alloy or pure iron; the ceramic-based biological material is TiO₂Isotropic pyrolytic carbon, hydroxyapatite, diamond or diamond-like; the polymer-based biomaterial is dacron, polytetrafluoroethylene, polyurethane, polyformaldehyde, silicone rubber, polylactic acid, glycolide-lactide copolymer, polytrimethylene carbonate or polycaprolactone.

By adopting the method, the cardiovascular coating material which has excellent mechanical property and can realize the slow release of the drug can be prepared.

The invention has the beneficial effects that:

1. by adopting the method, the modified cardiovascular stent coating carrying the double drugs is prepared by the cross-linking synergistic effect of the covalent bonds among the catechol group, the biomacromolecule amino group, the heparin molecule and the phenylboronic acid group, the interaction force among charges and the hydrogen bond interaction force, and the loading capacity of the anti-calcification drug and the anti-hyperplasia drug is controlled by carrying the drugs through the micelle, so that the method has high research and application values. The catechol compound is easy to be oxidized and dehydrogenated, phenol in the structure is easy to be converted into a quinone structure, and the structure can further perform Michael addition reaction and Schiff base reaction with a compound containing amino, so that polyamino biomacromolecules modified by catechol groups are mutually crosslinked, and a polymerized crosslinked drug-loaded coating can be formed on the surface of an implantable material. After the material modified by the method is implanted into a body, the anti-calcification and anti-proliferation drugs are coated by micelles, cross-linked with a coating, stacked by pi-pi and the like, so that the slow release of the drugs is realized, and the purposes of long-term anti-calcification and anti-proliferation are achieved.

2. Most of the traditional layer-by-layer self-assembly modified membranes only participate in non-covalent acting force, so that adverse conditions such as poor mechanical properties of the formed membrane are caused. The invention not only utilizes the non-covalent acting forces such as the electrostatic interaction acting force, the pi-pi accumulation action, the hydrogen bond action and the like among molecules, but also utilizes the covalent acting forces among catechol groups and between catechol groups and phenylboronic acid groups, and the mechanical property of the self-assembly modified coating is enhanced because the covalent bond and the non-covalent bond exist simultaneously.

3. The traditional coating drug loading method generally takes drugs as a coating, or loads the drugs on the coating in a soaking mode, and the mode has the advantages that the drug loading capacity is small, a large amount of hydrophobic drugs cannot be loaded, the loaded drugs are single in type, and burst release is easy to generate; the modified coating after burst release is easy to disintegrate, resulting in poor biocompatibility and the like, and long-term administration treatment cannot be realized. The method of the invention not only realizes the large loading of two hydrophobic drugs, namely the calcification drug and the antiproliferation drug, by introducing the drug-loaded micelle, but also realizes the slow release of the anticalcification drug and the antiproliferation drug by the restraint of the micelle on the membrane, covalent crosslinking, pi-pi accumulation and the like, and simultaneously can well maintain the structural stability of the self-assembled membrane along with the release of the drugs. The method can construct the modified coating of the cardiovascular material (such as a stent, an artificial small vessel and an artificial valve) with the calcification-resisting and anti-hyperplasia functions, realize the calcification-resisting and anti-smooth muscle hyperplasia functions and further improve the treatment performance of the implant material.

Drawings

FIG. 1 is a scanning electron microscope image of a cardiovascular coating material coated on the surface of a silicon wafer;

FIG. 2 is a drug release profile of cardiovascular coating material coated on the surface of a silicon wafer.

Detailed Description

Aiming at the problems that the restenosis of an implanted part is easily caused after the cardiovascular stent material is implanted into a human body, and the anti-calcification and anti-proliferative drugs of an organic coating material in the prior art are too low in amount and too high in drug release rate, the invention provides a brand-new preparation method of the coating material. The preparation method mainly comprises the following steps:

1) preparing a polycationic electrolyte solution

The polycation electrolyte in the invention preferentially adopts a catechol group modified polyamino compound, because the catechol compound is easy to be oxidized and dehydrogenated, phenol in the structure is easy to be converted into a quinone structure, and the structure can further generate Michael addition reaction and Schiff base reaction with an amino-containing compound, so that the catechol derivative modified polyamino biomacromolecules can be crosslinked with each other, and a polymerized crosslinked drug-loaded coating can be formed on the surface of an implantable material.

The preparation method of the catechol group modified polyamino compound comprises the following steps: dissolving molecules with both catechol groups and aldehyde groups in an organic solvent, slowly dripping the solution into a weakly acidic polyamino compound solution under the protection of nitrogen, reacting for 2-4 hours after complete dripping, wherein the molar ratio of the polyamino compound to the molecules with both catechol groups and aldehyde groups is 1-2: 1, slowly adding sufficient sodium borohydride until no bubbles are generated, dialyzing for 3 days in deionized water with the pH value of 5.0 by using a dialysis bag with the molecular weight of 10000, taking out, and freeze-drying to obtain the chitosan/chitosan composite material. Wherein the molecules having both catechol groups and aldehyde groups are selected from the following compounds: 3, 4-dihydroxybenzaldehyde, and the like; the polyamino compound is selected from the following compounds: polydiallylpolyhexylpurple nitrile, dimethyl ammonium chloride, poly-L-arginine hydrochloride, polyallylamine hydrochloride, polyethyleneimine, chitosan, and the like.

The surface of the polyamino compound modified by the catechol group is electropositive, and the polyamino compound is used as a layer-by-layer self-assembled polycation electrolyte. In the using process, the catechol group modified polyamino compound needs to exist in a solution form so that the reaction can be smoothly carried out, and therefore, the catechol group modified polyamino compound is prepared into a solution with the concentration of 0.1-10 mg/ml and the pH value of 6.

2) Preparing a polyinosinic electrolyte solution

Preparing the heparin with electronegative molecular surface into a solution with the concentration of 0.1-10 mg/ml and the pH value of 6 to obtain the polyionic electrolyte solution.

3) And preparation of mixed drug-loaded micelle

In order to increase the loading capacity of the anti-hyperplasia and anti-calcification drugs of the organic coating and achieve the purpose of delaying the release of the drugs, the invention changes the traditional mode of loading the drugs on the coating by soaking, the drugs are loaded on the micelles firstly, and then the micelles are firmly fixed on the organic coating through covalent crosslinking, pi-pi accumulation and other actions between the micelles and other molecules, thereby not only achieving the purpose of loading a large amount of drugs, but also realizing the slow release of the drugs. Therefore, the preparation of drug-loaded micelles is particularly important.

The preparation method of the drug-loaded micelle comprises the following steps: dissolving hydrophilic substances and hydrophobic substances in an organic solvent (DMSO and the like), adding adjacent hydroxyphenylboronic acid, stirring for reacting for 22-26 h, adding DCC and DMAP, continuing to react for 10-14 h, dialyzing for 2 days by using a dialysis bag with the molecular weight of 2000, and freeze-drying; wherein the mass ratio of the hydrophilic substance to the hydrophobic substance to the phenylboronic acid derivative to the DCC to DMAP is 2: 1-1.5: 0.5-1. Preparing a freeze-dried sample into a solution with the concentration of 0.1-10 mg/ml and the pH value of 6, then respectively dissolving an anti-calcification drug (atorvastatin calcium and the like) and an anti-proliferation drug (rapamycin or adriamycin and the like) into the solution, preparing an anti-calcification drug-loaded micelle and an anti-proliferation drug micelle with negative surfaces by a direct dissolution method, and adding the drug in an amount of 5-10% of the drug-loaded micelle; and mixing the two drug-loaded micelles in equal volume to obtain the mixed drug-loaded micelle. The hydrophilic substance is selected from the following compounds: polyacrylic acid, sodium polystyrene sulfonate, chondroitin sulfate, heparin, heparan sulfate, sodium alginate, keratan sulfate, collagen, hyaluronic acid or polyglutamic acid; the hydrophobic substance is selected from the following compounds: aliphatic hydrocarbons, cholesterol, polylysine, or polylactic acid, and the like.

4) Pretreatment of the base material

The method comprises the steps of preparing a dopamine solution with the concentration of 1-3 mg/ml in a buffer system with the pH value of 7-9, wherein the buffer system can be selected from a PBS buffer solution, an N-Tris (hydroxymethyl) methylglycine buffer solution, a triethanolamine buffer solution or a Tris-HCl buffer solution according to different reaction systems. And then placing the substrate material subjected to polishing, cleaning and drying treatment in a dopamine solution, reacting for 0.5-2 h at 10-30 ℃, then ultrasonically cleaning for 3-5 times, 5min each time, and drying under the nitrogen condition to obtain the substrate polydopamine modified layer of the layer-by-layer self-assembly coating.

5) Preparation of the coating Material

The cardiovascular coating material is prepared on the dopamine modification layer in a layer-by-layer self-assembly mode, and the layer-by-layer self-assembly can be carried out in a spin coating, dip coating or spraying mode. The invention preferentially adopts a dip-coating mode, and the specific method comprises the following steps: the method comprises the steps of soaking a substrate material coated with a dopamine modification layer in a polycation electrolyte solution for 10min, washing with deionized water to remove substances which are not firmly combined, soaking in a polyanion electrolyte solution for 10min, washing with deionized water to remove substances which are not firmly combined, continuously soaking in the polycation electrolyte solution for 10min, washing with deionized water to remove substances which are not firmly combined, soaking in hydrophilic and hydrophobic amphiphilic drug-loaded micelles for 20min, washing with deionized water to remove substances which are not firmly combined, preparing a sample, and circulating to form an assembly period, wherein the assembly period is one to prepare a coating layer. And repeating the assembly cycle for 1-30 times according to different use occasions to obtain the bifunctional cardiovascular coating material.

Since the effects obtained by the materials used in the present invention are substantially the same, no examples are given for each material individually. The specific embodiment of the invention selects representative materials, and details the preparation method of the long-acting sustained-release cardiovascular coating material with the dual functions of calcification resistance and hyperplasia resistance.

Example one

A preparation method of a long-acting slow-release cardiovascular coating material with double functions of calcification resistance and hyperplasia resistance is realized based on mussel bionics and phenylboronic acid modification, and comprises the following steps:

A. modifying dimethyl ammonium chloride by using 3, 4-dihydroxybenzaldehyde to obtain a catechol derivative modified positive charge biomacromolecule, wherein the molar ratio of the dimethyl ammonium chloride to the 3, 4-dihydroxybenzaldehyde is 2:1 during modification; preparing the modifier into a solution with the concentration of 10mg/ml and the pH value of 6 after finishing modification; the solution is used as a layer-by-layer self-assembled polycation electrolyte component;

B. preparing the negatively charged heparin into a solution with the concentration of 10mg/ml and the pH value of 6, wherein the solution is used as a layer-by-layer self-assembled polyanion electrolyte component;

C. weighing 2mg of hyaluronic acid, dissolving the hyaluronic acid in DMSO, adding 1.5mg of aliphatic hydrocarbon and 1mg of ortho-hydroxyphenylboronic acid, stirring to react for 26 hours, adding 1mg of DCC and 1mg of DMAP to continue to react for 14 hours, dialyzing, drying, preparing a dried product into a solution with the concentration of 10mg/ml and the pH value of 6, respectively dissolving fat-soluble atorvastatin calcium and rapamycin in the solution to prepare two different types of drug-loaded micelles, and mixing the two types of drug-loaded micelles according to the volume ratio of 1:1 to obtain a mixed drug-loaded micelle;

D. polishing, cleaning and drying the stainless steel;

E. preparing a dopamine solution with the final concentration of 2mg/ml in Tris-HCl buffer solution with the pH value of 8; putting the sample obtained in the step D into the reaction kettle, reacting for 0.5h at the temperature of 30 ℃, then carrying out ultrasonic cleaning, and carrying out N₂Drying to obtain a substrate modification layer of the layer-by-layer self-assembly coating, wherein the layer carries negative charges;

F. putting the sample obtained in the step E into the solution obtained in the step A for soaking for 10min, and then washing away the biological macromolecules (positive charge layer) which are not firmly combined by deionized water; then placing the sample into the heparin solution obtained in the step B for soaking for 10min, and then washing off the heparin molecules (negative charge layer) which are not firmly combined by deionized water; then, placing the sample into the solution obtained in the step A again for soaking for 10min, and then cleaning with deionized water (positive charge layer); finally, placing the sample into the mixed drug-loaded micelle obtained in the step C, soaking for 20min, and then cleaning with deionized water (negative charge layer); completing a cycle to form an assembly layer;

G. and F, circulating the step F for 5 times to obtain a final finished product.

Example two

The preparation method of the long-acting slow-release cardiovascular coating material with the double functions of calcification resistance and hyperplasia resistance is realized based on mussel bionics and phenylboronic acid modification, and comprises the following steps:

A. modifying polydiallyl polyhexyl purple nitrile by using 3, 4-dihydroxybenzaldehyde to obtain a catechol derivative modified positive charge biomacromolecule, wherein the molar ratio of polydiallyl polyhexyl purple nitrile to 3, 4-dihydroxybenzaldehyde is 1:1 during modification; preparing the modifier into a solution with the concentration of 2mg/ml and the pH value of 6 after finishing modification; the solution is used as a layer-by-layer self-assembled polycation electrolyte component;

B. preparing the negatively charged heparin into a solution with the concentration of 2mg/ml and the pH value of 6, wherein the solution is used as a layer-by-layer self-assembled polyanion electrolyte component;

C. weighing 2mg of hyaluronic acid, dissolving the hyaluronic acid in DMSO, adding 1mg of aliphatic hydrocarbon and 0.5mg of orthotopic hydroxyphenylboronic acid, stirring to react for 24 hours, adding 0.5mg of DCC and 0.5mg of DMAP, continuing to react for 12 hours, dialyzing, drying, preparing a dried substance into a solution with the concentration of 2mg/ml and the pH value of 6, respectively dissolving fat-soluble atorvastatin calcium and adriamycin in the solution to prepare two different types of drug-loaded micelles, and mixing the two types of drug-loaded micelles according to the volume ratio of 1:1 to obtain a mixed drug-loaded micelle;

D. polishing, cleaning and drying the silicon wafer;

E. preparing a dopamine solution with the final concentration of 3mg/ml in PBS buffer solution with the pH value of 7, placing the sample obtained in the step D into the dopamine solution, carrying out ultrasonic cleaning after reacting for 1 hour at the temperature of 20 ℃, and carrying out N washing₂Drying to obtain a substrate modification layer of the layer-by-layer self-assembly coating, wherein the layer carries negative charges;

G. and F, circulating for 20 times to obtain a final finished product.

The Scanning Electron Microscope (SEM) picture of the prepared bifunctional cardiovascular coating material is shown in figure 1, and micelle particles of about 200nm can be seen from the figure 1 and are uniformly distributed on the coating, which shows that the medicine-carrying micelle is successfully loaded in the coating. And the coating material is divided into a plurality of layers, each layer has a cross-linked network structure, and a large number of spherical micelle particles exist on the network structure. The spherical micelle particles are loaded with a large amount of hydrophobic drugs, and the slow release of the drugs can be realized through the restraint of the micelle on a membrane, covalent crosslinking, pi-pi stacking effect and the like.

FIG. 2 is a graph of the release profile of rapamycin from the coating material. The sustained release of rapamycin can be seen in the figure for more than 2 months, which shows that the coating of the invention has good sustained release property of long-acting drugs.

EXAMPLE III

A. modifying poly-L-arginine hydrochloride by using 3, 4-dihydroxybenzaldehyde to obtain catechol derivative modified positive charge biomacromolecule, wherein the molar ratio of the poly-L-arginine hydrochloride to the 3, 4-dihydroxybenzaldehyde is 1:1 during modification, and after the modification is finished, preparing a modifier into a solution with the concentration of 0.1mg/ml and the pH value of 6; the solution is used as a layer-by-layer self-assembled polycation electrolyte component;

B. preparing the negatively charged heparin into a solution with the concentration of 0.1mg/ml and the pH value of 6, wherein the solution is used as a layer-by-layer self-assembled polyanion electrolyte component;

C. weighing 2mg of hyaluronic acid, dissolving the hyaluronic acid in DMSO, adding 1.5mg of aliphatic hydrocarbon and 0.8mg of orthotopic hydroxyphenylboronic acid, stirring to react for 22 hours, adding 0.5mg of DCC and 0.5mg of DMAP, continuing to react for 10 hours, dialyzing, drying, preparing a dried substance into a solution with the concentration of 5mg/ml and the pH value of 6, respectively dissolving fat-soluble atorvastatin calcium and rapamycin in the solution to prepare two different types of drug-loaded micelles, and mixing the two types of drug-loaded micelles according to the volume ratio of 1:1 to obtain a mixed drug-loaded micelle;

D. polishing, cleaning and drying the hydroxyapatite;

E. preparing a dopamine solution with the final concentration of 1mg/ml in triethanolamine buffer solution with the pH value of 9, placing the sample obtained in the step D into the dopamine solution, reacting for 2 hours at the temperature of 30 ℃, ultrasonically cleaning, and carrying out N-washing₂Drying to obtain a substrate modification layer of the layer-by-layer self-assembly coating, wherein the layer carries negative charges;

G. and F, circulating for 30 times to obtain a final finished product.

Example four

A. modifying chitosan by using 3, 4-dihydroxy benzaldehyde to obtain catechol derivative modified positive charge biomacromolecule, wherein the molar ratio of chitosan to 3, 4-dihydroxy benzaldehyde is 1:1 during modification, and after modification is finished, preparing a modifier into a solution with the concentration of 5mg/ml and the pH value of 7; the solution is used as a layer-by-layer self-assembled polycation electrolyte component;

B. preparing the negatively charged heparin into a solution with the concentration of 5mg/ml and the pH value of 7, wherein the solution is used as a layer-by-layer self-assembled polyanion electrolyte component;

C. weighing 2mg of hyaluronic acid, dissolving the hyaluronic acid in DMSO, adding 1mg of aliphatic hydrocarbon and 0.5mg of ortho-hydroxyphenylboronic acid, stirring to react for 24 hours, adding 1mg of DCC and 1mg of DMAP, continuing to react for 12 hours, dialyzing, drying, preparing a dried product into a solution with the concentration of 10mg/ml and the pH value of 6, respectively dissolving fat-soluble atorvastatin calcium and adriamycin in the solution to prepare two different drug-loaded micelles, and mixing the two drug-loaded micelles according to the volume ratio of 1:1.5 to obtain a mixed drug-loaded micelle;

D. polishing, cleaning and drying the polyurethane;

E. preparing a dopamine solution with the final concentration of 2mg/ml in an N-tris (hydroxymethyl) methylglycine buffer solution with the pH value of 8, placing the sample obtained in the step D in the dopamine solution, reacting for 2 hours at the temperature of 10 ℃, ultrasonically cleaning, and carrying out N-type ultrasonic cleaning₂Drying to obtain a substrate modification layer of the layer-by-layer self-assembly coating, wherein the layer carries negative charges;

G. and F, circulating for 10 times to obtain a final finished product.

The invention changes the traditional preparation method of the drug-loaded coating, realizes the large loading of two hydrophobic drugs of anti-inflammation and NO release catalysis by introducing the drug-loaded micelle, realizes the slow release of the drugs by the restraint of the micelle on the membrane, covalent crosslinking, pi-pi accumulation and the like, and simultaneously can well maintain the structural stability of the self-assembled membrane along with the release of the drugs.

While the present invention has been described in detail with reference to the embodiments, it should not be construed as limited to the scope of the patent. Various modifications and changes may be made by those skilled in the art without inventive step within the scope of the appended claims.

Claims

1. A preparation method of a long-acting sustained-release cardiovascular coating material with dual functions of calcification resistance and hyperplasia resistance is characterized by comprising the following steps:

1) mixing a polyamino compound and molecules simultaneously carrying catechol groups and aldehyde groups according to a molar ratio of 1-2: 1, reacting for 1-3 hours, adding sodium borohydride, stirring until no bubbles are generated, dialyzing, freeze-drying, and preparing a dried product into a solution with the concentration of 0.1-10 mg/ml and the pH value of 6;

2) preparing heparin into a solution with the concentration of 0.1-10 mg/ml and the pH value of 6;

3) and preparing the mixed drug-loaded micelle: mixing a hydrophilic substance and a hydrophobic substance, adding o-hydroxyphenylboronic acid into the mixture, stirring and reacting for 22-26 h, adding DCC and DMAP, continuing to react for 10-14 h, dialyzing, freeze-drying, and preparing a dried substance into a solution with the concentration of 0.1-10 mg/ml; then respectively dissolving the anticalcification drug and the antiproliferative drug in the solution, preparing a drug-loaded micelle by a direct dissolution method, and mixing the two drug-loaded micelles in equal volume to obtain a mixed drug-loaded micelle for later use; wherein the mass ratio of the hydrophilic substance to the hydrophobic substance to the o-hydroxyphenylboronic acid to the DCC to the DMAP is 2: 1-1.5: 0.5-1;

4) polishing, cleaning and drying the base material needing modification;

6) soaking the base material treated in the step 5) in the solution obtained in the step 1) for 10min, taking out and washing with deionized water; then placing the mixture in the solution obtained in the step 2) for soaking for 10min, taking out and washing the mixture with deionized water; then placing the mixture in the solution obtained in the step 1) for soaking for 10min, taking out and washing the mixture with deionized water; finally, placing the gel in the drug-loaded micelle obtained in the step 3) for soaking for 20min, taking out and washing the gel with deionized water to finish a self-assembly period;

7) repeating the self-assembly cycle for 1-30 times to obtain the final product.

2. The preparation method of the long-acting sustained-release cardiovascular coating material with the dual functions of calcification resistance and hyperplasia resistance according to claim 1, which is characterized in that: the polyamino compound is polydiallyl polyhexylpurple nitrile, dimethyl ammonium chloride, poly-L-arginine hydrochloride, polyallylamine hydrochloride, polyethyleneimine or chitosan.

3. The preparation method of the long-acting sustained-release cardiovascular coating material with the dual functions of calcification resistance and hyperplasia resistance according to claim 1, which is characterized in that: the molecule with both catechol group and aldehyde group is 3, 4-dihydroxy benzaldehyde.

4. The preparation method of the long-acting sustained-release cardiovascular coating material with the dual functions of calcification resistance and hyperplasia resistance according to claim 1, which is characterized in that: the hydrophilic substance is polyacrylic acid, sodium polystyrene sulfonate, chondroitin sulfate, heparin, heparan sulfate, sodium alginate, keratan sulfate, collagen, hyaluronic acid or polyglutamic acid; the hydrophobic substance is aliphatic hydrocarbon, cholesterol, polylysine or polylactic acid.

5. The preparation method of the long-acting sustained-release cardiovascular coating material with the dual functions of calcification resistance and hyperplasia resistance according to claim 1, which is characterized in that: the anti-calcification drug is atorvastatin calcium; the antiproliferative drug is rapamycin or adriamycin.

6. The preparation method of the long-acting sustained-release cardiovascular coating material with the dual functions of calcification resistance and hyperplasia resistance according to claim 1, which is characterized in that: the buffer system is PBS buffer solution, N-Tris (hydroxymethyl) methylglycine buffer solution, triethanolamine buffer solution or Tris-HCl buffer solution.

7. The preparation method of the long-acting sustained-release cardiovascular coating material with the dual functions of calcification resistance and hyperplasia resistance according to claim 1, which is characterized in that: the substrate is a metal-based biomaterial, a ceramic-based biomaterial or a polymer-based biomaterial.

8. The preparation method of the long-acting sustained-release cardiovascular coating material with the dual functions of calcification resistance and hyperplasia resistance according to claim 7, wherein the preparation method comprises the following steps: the metal-based biomaterial is stainless steel, cobalt-based alloy, titanium and alloy thereof, magnesium and alloy thereof or pure iron; the ceramic-based biological material is TiO₂A film, isotropic pyrolytic carbon, hydroxyapatite, diamond or diamond-like; the polymer-based biomaterial is terylene, polytetrafluoroethylene, polyurethane, polyformaldehyde, silicon rubber, polylactic acid, glycolide-lactide copolymer, polytrimethylene carbonate or polycaprolactone.

9. The cardiovascular coating material prepared by the preparation method of any one of claims 1 to 8.