CN104383611A - Method for preparing medicine loading coating by assembling poly-electrolytes - Google Patents

Abstract

The invention discloses a method for preparing a medicine loading coating by assembling poly-electrolytes. The method mainly comprises the following steps: (1) preparing a coating by using a layered assembling method for the poly-electrolytes; (2) performing acid treatment on the coating to generate a micro-pore structure; (3) freezing and drying the coating to fix the micro-pore structure; (4) soaking the coating of the micro-pore structure into a medicine solution for medicine loading; (5) performing steam treatment on the medicine loading coating, and closing micro-pores in the coating to realize embedded loading of medicines. A poly-electrolyte coating prepared by virtue of the method is of the micro-pore structure, various types of medicines can be simultaneously loaded, the loading process is simple and convenient, and the loading capacity is controllable.

Description

A method for preparing drug-loaded coatings using polyelectrolyte assembly

technical field

The invention relates to the field of biomedical materials, in particular to a method for preparing a drug-loaded coating by using polyelectrolyte assembly.

Background technique

Drug-loaded supercoatings are constructed on the surface of medical devices and implants, and the functions of disease treatment, prevention, antibacterial, and interface biocompatibility can be improved through drug release.

The layered assembly method constructs supercoatings through the alternate adsorption of two interacting substances on the substrate, which has the advantages of simple operation, mild conditions, and no selectivity to the substrate. As a technology for constructing functional coatings, this method has been widely and deeply researched and applied in many frontier fields such as biomedicine, optoelectronic components, and artificial intelligence.

Polyelectrolytes, also known as polymer electrolytes, are a class of linear or branched synthetic and natural water-soluble polymers whose structural units contain ionizable groups. Generally, polyelectrolytes have a large number of charged groups, which can be classified into polymer acids, polymer bases, and amphoteric polymers according to the different charged groups. Upon dissolution, the protons of the polymer acid leave to form a polymer anion, and the polymer base can accept protons to form a polymer cation.

The patent application with the authorized notification number CN102319662B discloses a method for preparing a self-healing polyelectrolyte coating based on layer-by-layer assembly technology. into a solution with a certain concentration and a certain pH value; B. Immerse the treated substrate in the polycation solution for 1 to 30 minutes, then take the substrate out of the polycation solution, wash with water to remove the physically adsorbed substances on the surface of the substrate, and pass through N ₂ Drying; C. Immerse the substrate in step B into the polyanion solution for 1 to 30 minutes, then take the substrate out of the polyanion solution, wash with water to remove the physically adsorbed substances on the substrate surface, and then blow dry with N2 to complete a deposition Preparation of periodic coating; D. Repeat steps B and C to prepare self-healing polyelectrolyte coatings with multiple deposition cycles on the substrate. Among them, the polycation is polyallylamine hydrochloride, polyvinylimine, gelatin, polytetravinylpyridine, polydiallyldimethylamine hydrochloride, chitosan, poly-L-lysine One or more of acid and polyaniline; the polyanion is one or more of polyacrylic acid, sodium polystyrene sulfonate, sodium alginate, hyaluronic acid, sulfonated polyether ether ketone, and sulfonated dextran Several; solvents are water, ethanol, acetone, N,N-dimethylformamide, dimethyl sulfoxide or chloroform.

EP20028471A1 discloses a polymer layer with at least two opposite charges, and drugs are bound to one layer or two layers of the polymer in the form of covalent bonds. EP20169571A1 discloses a layer-by-layer assembly coating technology, in which the drug coating contains at least two layers of polyelectrolytes with opposite charges, and the drug is bound to it through covalent bonds.

In the field of biomedicine, the construction of drug-loaded coatings is one of the important applications of layered assembly methods. However, the loading of different types of drugs often takes different approaches: hydrophilic drugs usually have a strong interaction with the coating, so it is easier to generate enrichment in the pre-assembled coating; while the loading of hydrophobic drugs is more difficult , usually requires pretreatment of drugs or coatings, resulting in cumbersome steps, including multi-step synthesis, purification and other steps, and the types and quantities of applicable drugs are limited.

Therefore, how to improve the technology on the basis of the polyelectrolyte coating method to solve the above-mentioned problem of limited types and quantities of drugs loaded on the coating is an important research direction at present.

Contents of the invention

The invention provides a method for preparing a drug-loaded coating by using polyelectrolyte assembly. The polyelectrolyte coating prepared by the method has a microporous structure and can simultaneously load various types of drugs, and the loading process is simple and the loading capacity is large. And controllable.

Specific embodiments of the present invention are as follows:

A method for preparing a drug-loaded coating utilizing polyelectrolyte assembly, comprising the following steps:

(1) prepare alkaline polycation electrolyte aqueous solution and acidic polyanion electrolyte aqueous solution;

(2) After soaking the base material in the polycation electrolyte aqueous solution, rinse with the aqueous solution; then soak in the polyanion electrolyte aqueous solution, and rinse with the aqueous solution; repeat this step for 5 to 500 times to obtain the polyelectrolyte coating;

(3) Immerse the polyelectrolyte coating in an acidic aqueous solution, freeze-dry after taking it out, and obtain a polyelectrolyte coating with a microporous structure;

(4) Immersing the polyelectrolyte coating with a microporous structure in the drug solution to obtain a drug-loaded coating;

(5) Place the drug-loaded coating in a water vapor environment to obtain a drug-loaded coating with closed micropores.

The principle of coating acid solution treatment to form microporous structure is the phase separation of polyelectrolyte: in acidic environment, the charge density of polyelectrolyte will change, which will lead to the movement and recombination of polycation and polyanion chain segments in the coating, Microvoids are formed in the final coating.

The principle of drug loading on the polyelectrolyte coating with microporous structure: the drug solution is infiltrated and filled into the micropores through capillary action.

The principle of closing micropores by treating porous coatings with water vapor is that water molecules can promote the movement of polyelectrolyte segments. Under the water vapor environment, the coating tends to have the lowest surface energy, resulting in the closure of micropores in the coating. The degree of micropore closure can be achieved by water vapor concentration or treatment time.

The pH value of the polycation electrolyte aqueous solution is 8-11, and the pH value of the polyanion electrolyte aqueous solution is 2-6.

The pH of the acidic aqueous solution is 2-6, and the size of the micropores can be regulated by changing the pH value of the acid solution, thereby controlling the drug loading, preferably 2.5-5.0.

The polycationic electrolyte can be artificially synthesized or a naturally positively charged polyelectrolyte, preferably, the polycationic electrolyte is protamine sulfate, polylysine, chitosan, polyethyleneimine, polypropylene amine hydrochloride or polydiallyldimethylammonium hydrochloride. Polyethyleneimine is preferred, having a molar molecular weight of 1,000 to 100,000, more preferably 20,000 to 50,000.

The polyanionic electrolyte can be artificially synthesized or naturally negatively charged polyelectrolyte, preferably, the polyanionic electrolyte is polyacrylic acid, hyaluronic acid, heparin, sodium polybenzenesulfonate or sodium alginate. Polyacrylic acid is preferred, having a molar molecular weight of 50,000 to 500,000, more preferably 100,000 to 200,000.

Preferably, the concentration of the polycation electrolyte solution or polyanion electrolyte solution is 0.1-10 mg/ml.

The length of soaking time can regulate the size of the micropores, thereby controlling the drug loading. The soaking time in step (2) is 1-60 minutes, preferably 5-20 minutes. Preferably, the steps are repeated for 10 to 100 times.

In step (3), the polyelectrolyte coating is immersed in the acidic aqueous solution for 10-200 minutes, preferably 30-90 minutes. The freeze-drying temperature ranges from -20°C to -80°C, and the time ranges from 1 to 20 hours.

The time for the polyelectrolyte coating to be immersed in the drug solution in step (4) is 30 seconds to 100 minutes, preferably 1 to 10 minutes. The type of the drug is not limited, and it can be hydrophilic, hydrophobic or lipophilic, and the drug can be configured as a solution or a suspension.

In step (5), the coating is placed in a water vapor environment, the humidity is 70-100%, preferably 90-100%, and the treatment time is 1-48 hours, preferably 2-5 hours.

In the present invention, on the basis of using polyelectrolyte to construct the coating, the polyelectrolyte coating is modified by an acidic aqueous solution to obtain a polyelectrolyte coating with a microporous structure, which can load various types of drugs, and the loading process Simple, large loading capacity, no need for chemical covalent grafting, and no special equipment; the micropores of the coating can be closed after the drug-loaded coating is placed in a water vapor environment, which can make the gap between the drug and the coating The combination is tighter, thereby increasing the loading capacity and stability of the hydrophobic drug; it solves the problem in the prior art that the types and quantities of the drug loaded on the coating are limited and cannot be precisely regulated.

Description of drawings

Fig. 1 is the scanning electron micrograph after the acid treatment of polyethyleneimine/polyacrylic acid coating in embodiment 1;

FIG. 2 is a scanning electron micrograph of the polyethyleneimine/polyacrylic acid-loaded triclosan drug coating in Example 1 after water vapor treatment.

Detailed ways

Example 1

A method for preparing a drug-loaded coating utilizing polyelectrolyte assembly, comprising the following steps:

(1) preparation of pH 9, 1mg/ml polyethyleneimine aqueous solution and pH 3, 3mg/ml polyacrylic acid aqueous solution;

(2) Soak the stainless steel support in the polyethyleneimine aqueous solution for 15 minutes, then rinse with the aqueous solution; put it into the polyacrylic acid aqueous solution and soak for 15 minutes, then rinse with the aqueous solution; after repeating this step 20 times, obtain the polyelectrolyte coating;

(3) Immerse the polyelectrolyte coating in an aqueous hydrochloric acid solution with a pH of 2.9 for 30 minutes, freeze-dry and dehydrate at -80°C to obtain a polyelectrolyte coating with a microporous structure;

(4) Immerse the polyelectrolyte coating with a microporous structure in a triclosan solution (20mg/ml, ethanol) for 1 minute, and remove the ethanol in a vacuum to obtain a coating loaded with triclosan, with a drug loading of 1mg/ml cm ² ;

(5) After placing the triclosan-loaded coating in a water vapor environment with a humidity of 95% for 5 hours, a drug-loaded coating with closed micropores was obtained.

The above-mentioned polyelectrolyte coating with a microporous structure is shown in Figure 1; the drug-loaded coating after loading triclosan is shown in Figure 2; the drug-loaded coating realizes the rapid loading of triclosan, and the triclosan drug passes through Free molecular diffusion is released slowly; the in vitro antibacterial experiment of the drug-loaded coating stent shows that it has killing effect on both Escherichia coli and Staphylococcus aureus during the 30-day experiment period.

Example 2

A method for preparing a drug-loaded coating utilizing polyelectrolyte assembly, comprising the following steps:

(1) Prepare pH 10.0, 1mg/ml polylysine aqueous solution and pH 3, 2mg/ml hyaluronic acid aqueous solution;

(2) Soak the stainless steel cardiovascular stent in the polylysine aqueous solution for 20 minutes, then rinse with the aqueous solution; put it into the hyaluronic acid aqueous solution for 20 minutes, and then rinse with the aqueous solution; after repeating this step 40 times, the polyelectrolyte coated layer;

(3) Immerse the polyelectrolyte coating in an aqueous hydrochloric acid solution with a pH of 3.5 for 60 minutes, freeze-dry and dehydrate at -25°C to obtain a polyelectrolyte coating with a microporous structure;

(4) Immerse the polyelectrolyte coating with a microporous structure in the fisetin solution (20mg/ml, ethanol) for 5 minutes, remove the ethanol in a vacuum, and obtain a coating loaded with fisetin, with a drug loading of 0.5mg/ml cm ² ;

(5) After the coating loaded with fisetin was placed in a 100% humidity water vapor environment for 1 hour, a drug-loaded coating with closed micropores was obtained.

The drug-loaded coating realizes rapid loading of fisetin, and the drug-loaded fisetin is slowly released through free molecular diffusion; the in vitro anticoagulant experiment of the drug-loaded stent showed that during the 7-day experimental period, there was no coagulation and no coagulation on the surface of the stent. thrombosis.

Example 3

A method for preparing a drug-loaded coating utilizing polyelectrolyte assembly, comprising the following steps:

(1) Protamine sulfate aqueous solution and pH 4,5mg/ml sodium polybenzenesulfonate aqueous solution of preparation pH 9.5,5mg/ml;

(2) Soak the urinary catheter in the protamine sulfate aqueous solution for 10 minutes, and rinse it with an aqueous solution; then soak it in an aqueous solution of sodium polybenzenesulfonate for 10 minutes, and rinse it with an aqueous solution; after repeating this step 100 times, obtain the polyelectrolyte coating;

(3) Immerse the polyelectrolyte coating in an aqueous hydrochloric acid solution with a pH of 4.0 for 120 minutes, freeze-dry and dehydrate at -80°C to obtain a polyelectrolyte coating with a microporous structure;

(4) Immerse the polyelectrolyte coating with a microporous structure in a mixed ethanol solution of triclosan and indomethacin (each 10 mg/mL) for 10 minutes, remove the ethanol in a vacuum, and obtain loaded triclosan and indomethacin coatings, the drug loads are 0.8mg/cm ² ;

(5) After the urinary catheter loaded with triclosan and indomethacin coating was placed in a 95% humidity water vapor environment for 10 hours, a drug-loaded coating with micropore closure was obtained.

The drug-loaded coating realizes the loading of triclosan and indomethacin, and the drug is slowly released through free molecular diffusion; the in vitro antibacterial test of the drug-loaded catheter shows that during the 90-day experimental period, the drug-loaded coating has no effect on Escherichia coli and Staphylococcus aureus has a killing effect.

Example 4

A method for preparing a drug-loaded coating utilizing polyelectrolyte assembly, comprising the following steps:

(1) prepare pH 8, 2mg/ml polylysine aqueous solution and pH 6, 2mg/ml heparin aqueous solution;

(2) After immersing the stainless steel cardiovascular stent in polylysine aqueous solution for 10 minutes, rinse with aqueous solution; then soak in heparin aqueous solution for 10 minutes, then rinse with aqueous solution; after repeating this step 80 times, obtain polyelectrolyte coating;

(3) Immerse the polyelectrolyte coating in an aqueous hydrochloric acid solution with a pH of 3.0 for 120 minutes, freeze-dry and dehydrate at -80°C to obtain a polyelectrolyte coating with a microporous structure;

(4) Immerse the polyelectrolyte coating with a microporous structure in an aqueous solution of hepatocyte growth factor (1 mg/mL) for 10 minutes, remove the water in a vacuum, and obtain a coating loaded with hepatocyte growth factor, and the growth factor drug load is 5 μg /cm ² ;

(5) After the stent loaded with the hepatocyte growth factor coating was placed in a 95% humidity water vapor environment for 10 hours, a drug-loaded coating with micropores closed was obtained.

The drug-loaded coating realizes the loading of hepatocyte growth factor, and the growth factor is slowly released through free molecular diffusion; the in vitro endothelial cell culture experiment of the drug-loaded stent shows that in the 7-day experimental period, the proliferation and migration of endothelial cells have a significant promoting effect.

Example 5

A method for preparing a drug-loaded coating utilizing polyelectrolyte assembly, comprising the following steps:

(1) prepare pH 8,3mg/ml alkali-soluble chitosan aqueous solution and pH 6,2mg/ml hyaluronic acid aqueous solution;

(2) After soaking the polydimethylsiloxane scaffold treated with plasma hydrophilicity in the alkali-soluble chitosan aqueous solution for 10 minutes, the aqueous solution was rinsed; after being put into the hyaluronic acid aqueous solution and soaked for 10 minutes, the aqueous solution was rinsed; the After the steps are repeated 50 times, a polyelectrolyte coating is obtained;

(3) Immerse the polyelectrolyte coating in an aqueous hydrochloric acid solution with a pH of 3.0 for 120 minutes, freeze-dry and dehydrate at -80°C to obtain a polyelectrolyte coating with a microporous structure;

(4) Immerse the polyelectrolyte coating with a microporous structure in an aqueous solution of vascular endothelial growth factor (1 mg/mL) for 10 minutes, remove the water in a vacuum, and obtain a coating loaded with vascular endothelial growth factor, and the growth factor drug loading is 3 μg /cm ² ;

(5) After the stent loaded with the vascular endothelial growth factor coating was placed in a 100% humidity water vapor environment for 3 hours, a drug-loaded coating with micropores closed was obtained.

The drug-loaded coating realizes the loading of vascular endothelial cell growth factor, and the growth factor is slowly released through free molecular diffusion; the in vitro endothelial cell culture experiment of the drug-loaded coating stent shows that during the 7-day experimental period, the adhesion to endothelial cells, Proliferation and migration are significantly promoted.

Example 6

A method for preparing a drug-loaded coating utilizing polyelectrolyte assembly, comprising the following steps:

(1) preparation of pH 9, 3mg/ml polyethyleneimine aqueous solution and pH 5, 3mg/ml hyaluronic acid aqueous solution;

(2) After soaking the titanium alloy artificial bone joint in the polyethyleneimine aqueous solution for 10 minutes, rinse with the aqueous solution; then soak in the aqueous hyaluronic acid solution for 10 minutes, and rinse with the aqueous solution; after repeating this step 50 times, the polyelectrolyte coating;

(3) Immerse the polyelectrolyte coating in an aqueous hydrochloric acid solution with a pH of 2.5 for 60 minutes, freeze-dry and dehydrate at -80°C to obtain a polyelectrolyte coating with a microporous structure;

(4) Immerse the polyelectrolyte coating with a microporous structure in an aqueous solution of lysozyme (1 mg/mL) for 10 minutes, remove the water in a vacuum to obtain a coating loaded with lysozyme, and the drug loading is 5 μg/cm ² ;

(5) After the artificial bone joint loaded with the lysozyme coating was placed in a 100% humidity water vapor environment for 1 hour, a drug-loaded coating with closed micropores was obtained.

The drug-loaded coating realizes the loading of lysozyme, and the enzyme is slowly released through free molecular diffusion; the in vitro bacterial culture experiment of the artificial bone joint with the drug-loaded coating shows that in the 14-day experimental period, the killing effect on Escherichia coli and Staphylococcus aureus Extermination and inhibition have a significant effect.

Example 7

A method for preparing a drug-loaded coating utilizing polyelectrolyte assembly, comprising the following steps:

(1) polyallylamine hydrochloride aqueous solution and pH 3,3mg/ml sodium polybenzenesulfonate aqueous solution of preparation pH 9,1mg/ml;

(2) Soak the glass in an aqueous solution of polyallylamine hydrochloride for 8 minutes, and rinse it with an aqueous solution; then soak it in an aqueous solution of sodium polybenzenesulfonate for 8 minutes, and rinse it with an aqueous solution; after repeating this step 150 times, a polyelectrolyte coating;

(3) Immerse the polyelectrolyte coating in an aqueous hydrochloric acid solution with a pH of 3 for 60 minutes, freeze-dry and dehydrate at -80°C to obtain a polyelectrolyte coating with a microporous structure;

(4) Immerse the polyelectrolyte coating with a microporous structure in an aqueous solution of lysozyme (1 mg/mL) for 10 minutes, remove the water in a vacuum to obtain a coating loaded with lysozyme, and the drug loading is 2 μg/cm ² ;

(5) After the glass loaded with the lysozyme coating was placed in a 100% humidity water vapor environment for 1 hour, a drug-loaded coating with micropores closed was obtained.

The drug-loaded coating realizes the loading of lysozyme, and the enzyme is slowly released through free molecular diffusion; the in vitro bacterial culture experiment of the drug-loaded coating shows that in the 7-day experimental period, the killing and killing of Escherichia coli and Staphylococcus aureus Inhibition has a significant effect.

Example 8

A method for preparing a drug-loaded coating utilizing polyelectrolyte assembly, comprising the following steps:

(1) polydiallyldimethylammonium hydrochloride aqueous solution and pH 4, 3mg/ml sodium polybenzenesulfonate aqueous solution of preparation pH 9,1mg/ml;

(2) Soak the polyethylene terephthalate artificial blood vessel in the polydiallyldimethylammonium hydrochloride aqueous solution for 8 minutes, and rinse it with the aqueous solution; put it into the sodium polybenzenesulfonate aqueous solution and soak it for 8 minutes Minutes later, the aqueous solution was rinsed; after this step was repeated 150 times, a polyelectrolyte coating was obtained;

(3) Immerse the polyelectrolyte coating in an aqueous hydrochloric acid solution with a pH of 4.0 for 120 minutes, freeze-dry and dehydrate at -80°C to obtain a polyelectrolyte coating with a microporous structure;

(4) Immerse the polyelectrolyte coating with a microporous structure in a mixed ethanol solution of triclosan and indomethacin (each 10 mg/mL) for 10 minutes, remove the ethanol in a vacuum, and obtain loaded triclosan and indomethacin The coating, the drug load is 0.3mg/cm ² respectively;

(5) After the artificial blood vessel loaded with triclosan and indomethacin coating was placed in a 100% humidity water vapor environment for 5 hours, a drug-loaded coating with micropore closure was obtained.

The drug-loaded coating realizes the loading of triclosan and indomethacin, and the drug is slowly released through free molecular diffusion; the in vitro antibacterial experiment of the drug-loaded coating artificial blood vessel shows that in the 90-day experimental period, it is resistant to Escherichia coli and golden Staphylococcus has a killing effect.

Claims (8)

1. A method utilizing polyelectrolyte assembly to prepare drug-loaded coating, is characterized in that, comprises the following steps: (1) preparing alkaline polycation electrolyte aqueous solution and acidic polyanion electrolyte aqueous solution; (2) After soaking the base material in the polycation electrolyte aqueous solution, rinse with the aqueous solution; then soak in the polyanion electrolyte aqueous solution, and rinse with the aqueous solution; repeat this step for 5 to 500 times to obtain the polyelectrolyte coating; (3) immerse the polyelectrolyte coating in an acidic aqueous solution, freeze-dry after taking it out, and obtain a polyelectrolyte coating with a microporous structure; (4) immersing the polyelectrolyte coating with a microporous structure in the drug solution to obtain a drug-loaded coating; (5) placing the drug-loaded coating in a water vapor environment to obtain a drug-loaded coating with micropores closed. 2. the method utilizing polyelectrolyte assembly to prepare drug-loaded coating as claimed in claim 1, is characterized in that, the pH value of described polycation electrolyte aqueous solution is 8～11, the pH value of polyanion electrolyte aqueous solution is 2～ 6. 3 . The method for preparing a drug-loaded coating by assembling polyelectrolytes according to claim 1 , wherein the pH value of the acidic aqueous solution is 2-6. 4 . 4 . The method for preparing a drug-loaded coating by assembling polyelectrolyte as claimed in claim 1 , wherein the polyelectrolyte coating is immersed in an acidic aqueous solution for 10 minutes to 200 minutes. 5 . The method for preparing a drug-loaded coating by assembling polyelectrolytes according to claim 1 , wherein the freeze-drying temperature is -20° C. to -80° C. and the time is 1 to 20 hours. 6 . The method for preparing a drug-loaded coating by assembling polyelectrolytes as claimed in claim 1 , wherein the humidity of the water vapor environment is 70-100%, and the treatment time is 1-48 hours. 7. utilize polyelectrolyte assembly as claimed in claim 1 to prepare the method for drug-loaded coating, it is characterized in that, described polycation electrolyte is protamine sulfate, polylysine, chitosan, polyethyleneimine , polyallylamine hydrochloride or polydiallyldimethylammonium hydrochloride. 8. the method utilizing polyelectrolyte assembly to prepare drug-loaded coating as claimed in claim 1, is characterized in that, described polyanionic electrolyte is polyacrylic acid, hyaluronic acid, heparin, sodium polybenzenesulfonate or sodium alginate .