Antibacterial and anticoagulant coating composition, antibacterial and anticoagulant coating, preparation method of antibacterial and anticoagulant coating and medical material

Abstract

The invention provides an antibacterial and anticoagulant coating composition, an antibacterial and anticoagulant coating, a preparation method of the antibacterial and anticoagulant coating and a medical material. The antibacterial and anticoagulant coating composition comprises a catechol compound, a polyamine compound, an anticoagulant substance and a metal salt. The preparation method of the antibacterial and anticoagulant coating based on the coating composition is simpler and more convenient. The preparation method comprises the following steps: firstly, carrying out a Schiff base reaction between a catechol compound and a polyamine compound to construct a phenol amine bottom layer coating; the activated anticoagulant substance and amino of the phenol amine bottom layer coating are subjected to a chemical reaction, and fixation of the anticoagulant substance is achieved; and finally, forming a complex coating of the anticoagulant substance and the metal ions on the surface through a coordination reaction of the anticoagulant substance and the metal ions. According to the invention, the complex coating of the anticoagulant substance and the metal ions can activate the anticoagulant ability of the anticoagulant substance and improve the self-cleaning ability and photocatalytic antibacterial ability of the surface of the coating.

Description

Technical Field

[0001] The invention belongs to the field of medical materials, and specifically relates to an antibacterial and anticoagulant coating composition, an antibacterial and anticoagulant coating and a preparation method thereof, and a medical material. Background Art

[0002] Cardiovascular diseases seriously threaten human health. At present, implanting blood-contact devices, such as catheters, heart valves, artificial blood vessels, etc., has become a common method for treating cardiovascular diseases in clinical practice. At present, polyurethane has been widely used in the preparation of medical devices due to its good chemical stability and mechanical properties. However, polyurethane materials have poor blood compatibility and are prone to thrombus formation during use, which limits their application in blood-contact devices. In addition, blood-contact devices usually do not have hydrophilic lubrication and sterilization capabilities, which will lead to the adhesion of bacteria and proteins during use, increasing the risk of infection. Therefore, improving the anticoagulant and antibacterial effects of blood-contact devices is of great practical significance for their practical application.

[0003] Regarding anticoagulation, studies have shown that fixing anticoagulants on the surface of blood-contacting devices can improve their anticoagulant effect. In general, the anticoagulant structure and anticoagulant mechanism of anticoagulants are quite different. For example, heparin, an indirect inhibitor of thrombin, has a special pentasaccharide sequence that can specifically bind to antithrombin III. Due to the charge effect of heparin, it can further bind to thrombin to form a ternary complex to inhibit the activity of thrombin and play an anticoagulant role. In addition, heparin also has the function of inhibiting smooth muscle cell proliferation and promoting endothelial cell proliferation, which can effectively reduce the occurrence of restenosis caused by common blood-contacting devices. At present, there are two strategies for surface modification of heparin: (1) forming a drug-releasing coating through physical adsorption means such as electrostatic interaction, but due to the instability of physical adsorption, it is easy to cause heparin to fall off; (2) obtaining a heparin functionalized surface formed by covalent bonds through chemical reactions such as amide reactions. This method has stable binding and high safety. However, although heparin can directly interact with coagulation factors, its loading capacity is limited. If it is used for too long, the anticoagulant effect will be significantly reduced.

[0004] For antibacterial purposes, the surface of blood-contact devices is usually formed by fixing antibacterial polymers or loading antibacterial agents. However, fixing antibacterial polymers usually weakens the anticoagulant ability of the surface, while loading antibacterial agents will induce drug resistance problems.

[0005] It can be seen that the anticoagulant and antibacterial effects of the surface of blood-contact devices still need to be further improved. Summary of the invention

[0006] In view of this, the purpose of the present invention is to provide an antibacterial and anticoagulant coating composition, an antibacterial and anticoagulant coating and a preparation method thereof, and a medical material. The antibacterial and anticoagulant composite coating has excellent antibacterial and anticoagulant effects.

[0007] To achieve this object, the present invention adopts the following technical solutions:

[0008] In a first aspect, the present invention provides an antibacterial and anticoagulant coating composition, comprising a catechol compound, a polyamine compound, an anticoagulant substance and a metal salt.

[0009] Preferably, the ratio of the catechol compound, the polyamine compound, the anticoagulant substance and the metal salt is (35-45):(35-45):(5-12):(10-20).

[0010] Preferably, the catechol compound includes any one or more of catechol, catechol acid, catecholamine or flavonoid compound.

[0011] Preferably, the polyamine compound includes any one or more of polyimine, polyamide or polyurethane.

[0012] Preferably, the anticoagulant substance comprises heparin and / or tissue plasminogen.

[0013] Preferably, the metal salt includes any one or more of titanium metal salt, zinc metal salt or tin metal salt.

[0014] More preferably, the catechol compound is selected from any one or more of levodopa, norepinephrine, dopamine, dopamine hydrochloride or phenylalanine.

[0015] More preferably, the polyamine compound is selected from polyethyleneimine and / or polyacrylamide.

[0016] More preferably, the titanium metal salt is selected from TiBALDH and / or tetrabutyl titanate, the zinc metal salt is selected from zinc acetate dihydrate and / or zinc nitrate, and the tin metal salt is selected from tin chloride dihydrate and / or tin acetate.

[0017] In a second aspect, the present invention provides an antibacterial and anticoagulant coating, which is formed from the above-mentioned antibacterial and anticoagulant coating composition.

[0018] In a third aspect, the present invention provides a method for preparing an antibacterial and anticoagulant coating, comprising the following steps:

[0019] S1: soaking the medical material in a solution containing a catechol compound and a polyamine compound, and after reaction, obtaining a medical material coated with a phenolamine base coating;

[0020] S2: soaking the medical material coated with the phenolamine base coating with an anticoagulant solution, and obtaining the medical material modified with the anticoagulant after reaction;

[0021] S3: soaking the medical material modified by the anticoagulant substance in a metal salt solution, and obtaining the medical material with an antibacterial and anticoagulant coating after the reaction.

[0022] Preferably, the soaking temperatures in steps S1, S2 and S3 are each independently 20 to 40°C, and the soaking times are each independently 0.1 to 30 hours.

[0023] Preferably, the soaking in step S2 is performed at a pH of 4 to 6.

[0024] Preferably, the anticoagulant in the anticoagulant solution in step S2 is activated.

[0025] Preferably, the soaking in step S3 is performed at a pH of 8 to 10.

[0026] In a fourth aspect, the present invention provides a medical material, comprising a medical material substrate and a coating formed on the surface of the medical material substrate;

[0027] The coating is the above-mentioned antibacterial and anticoagulant coating.

[0028] Preferably, the medical material matrix comprises a blood-contact material matrix.

[0029] Preferably, the material of the medical material matrix is ​​selected from metal materials, inorganic non-metallic matrix materials or polymer materials.

[0030] Preferably, the thickness of the antibacterial and anticoagulant coating is 10 to 25 μm.

[0031] Compared with the prior art, the present invention has the following beneficial effects:

[0032] The present invention provides an antibacterial and anticoagulant coating composition, which includes a catechol compound, a polyamine compound, an anticoagulant substance and a metal salt. At the same time, based on the coating composition, the present invention provides an antibacterial and anticoagulant coating, and its preparation method is simpler and more convenient. The preparation method first constructs a phenolamine base coating by a Schiff base reaction between a catechol compound and a polyamine compound; then uses an activated anticoagulant substance to react chemically with the amino group of the phenolamine base coating to achieve the fixation of the anticoagulant substance; finally, through the coordination reaction between the anticoagulant substance and the metal ion, a complex coating of the anticoagulant substance and the metal ion is formed on the surface. In the present invention, the complex coating of the anticoagulant substance and the metal ion can activate the anticoagulant ability of the anticoagulant substance, and improve the self-cleaning ability and photocatalytic antibacterial ability of the coating surface. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1 This is a comparison chart of the hemolysis rates of the products obtained in Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 4;

[0034] Figure 2 This is a comparative diagram of the evaluation of the anti-protein adhesion amount of the products obtained in Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 4;

[0035] Figure 3 This is a comparison electron microscope picture of platelet adhesion of the products obtained in Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 4;

[0036] Figure 4 It is a comparison chart of the activated partial thromboplastin time (APTT) and the thrombin time (TT) of the products obtained in Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 4;

[0037] Figure 5 This is a comparison chart of the antibacterial rates of the products obtained in Example 1, Comparative Example 1, and Comparative Example 2;

[0038] Figure 6 This is a comparison diagram of the visible light absorption peaks of the products obtained in Example 1 and Comparative Example 4;

[0039] Figure 7 The cell viability data graphs of the products obtained in Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 4 are shown. DETAILED DESCRIPTION

[0040] The technical solution of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0041] The present invention provides an antibacterial and anticoagulant coating composition, which comprises a catechol compound, a polyamine compound, an anticoagulant substance and a metal salt; the mass ratio of the catechol compound, the polyamine compound, the anticoagulant substance and the metal salt is (35-45):(35-45):(5-12):(10-20), preferably (35-40):(40-45):(8-10):(10-15), and more preferably 38:42:8:12.

[0042] In the present invention, the catechol compounds include any one or more of catechol, catechol acid, catechol acid amine or flavonoid compounds; the polyamine compounds include any one or more of polyimine, polyamide or polyurethane; the anticoagulant includes heparin and / or tissue plasminogen; the metal salt includes any one or more of titanium metal salt, zinc metal salt or tin metal salt.

[0043] In some preferred embodiments of the present invention, the catechol compound is dopamine hydrochloride; and / or, the polyamine compound is polyethyleneimine; and / or, the anticoagulant drug is heparin; and / or, the metal salt solution is a titanate solution.

[0044] The present invention also provides an antibacterial and anticoagulant coating formed by the antibacterial and anticoagulant coating composition.

[0045] The present invention also provides a method for preparing the above-mentioned antibacterial and anticoagulant coating, comprising the following steps:

[0046] S1: soaking the medical material in a solution containing a catechol compound and a polyamine compound, and after reaction, obtaining a medical material coated with a phenolamine base coating;

[0047] S2: soaking the medical material coated with the phenolamine base coating with an anticoagulant solution, and obtaining the medical material modified with the anticoagulant after reaction;

[0048] S3: soaking the medical material modified by the anticoagulant substance in a metal salt solution, and obtaining the medical material with an antibacterial and anticoagulant coating after the reaction.

[0049] According to the present invention, the medical material is firstly immersed in a solution containing catechol compounds and polyamine compounds, and after the reaction, the medical material coated with a phenolamine base coating is obtained.

[0050] Wherein, the medical material may include a blood contact material matrix; the material of the medical material is selected from metal materials, inorganic non-metallic matrix materials or polymer materials. In some embodiments of the present invention, the medical material is a thermoplastic polyurethane material, a polypropylene material, etc. Before soaking, the medical material is preferably pretreated, and the pretreatment generally includes: preparing the medical material into a substrate, washing, drying, and setting aside. The steps of preparing the substrate, washing and drying are not particularly limited, and can be performed according to means familiar to those skilled in the art.

[0051] In the present invention, the catechol compound includes any one or more of catechol, catechol acid, catecholamine or flavonoid compounds, and can be specifically selected from any one or more of levodopa, norepinephrine, dopamine, dopamine hydrochloride or phenylalanine. In the present invention, the concentration of the solution containing the catechol compound is 10 to 40 mg / mL, preferably 10 to 30 mg / mL, and most preferably 15 to 25 mg / mL, and the solvent includes any one or more of water, acetone or dimethyl sulfoxide.

[0052] In the present invention, the polyamine compound includes any one or more of polyimine, polyamide or polyurethane, and is specifically selected from polyethyleneimine (abbreviated as: PEI) and / or polyacrylamide. In the present invention, the concentration of the solution containing the polyamine compound is 10 to 40 mg / mL, preferably 10 to 30 mg / mL, and most preferably 15 to 25 mg / mL, and the solvent includes any one or more of water, acetone or ethanol.

[0053] In some embodiments of the present invention, step S1 is: preferably immersing the medical material in a solution containing catechol compounds and polyamine compounds at 20-40°C for 0.1-30 hours, preferably immersing it at 25-37°C for 10-24 hours, and constructing a phenolamine base coating through a Schiff base reaction between the catechol compounds and the polyamine compounds to obtain a medical material coated with the phenolamine base coating.

[0054] In some preferred embodiments of the present invention, the bottom coating is obtained by reacting dopamine with polyethyleneimine, and the preparation method of the bottom coating comprises the following steps:

[0055] Step 1: ultrasonically clean the substrate material in an ethanol aqueous solution and dry it for later use;

[0056] Step 2: Place the cleaned medical material in a dopamine / polyethyleneimine mixed solution and soak for 8 to 14 hours, then take it out and shake and wash it for 5 to 12 hours to remove the compounds physically adhering to the surface and then dry it;

[0057] Wherein, in step 2, the concentration ratio of dopamine to polyethyleneimine is (0.5-2):1, preferably (0.75-1.5):1, and / or the solvent in the dopamine / polyethyleneimine mixed solution is at least one of water, ethanol or methanol. In step 2, the method for preparing the bottom coating solution also includes placing the cleaned medical material in the dopamine solution for 10-15 hours, shaking and washing for 10-15 hours, and then drying, and then placing it in the polyethyleneimine solution for 10-15 hours, shaking and washing for 10-15 hours, and then drying for standby use.

[0058] In the present invention, the phenolamine deposition process utilizes the non-covalent interaction between the polyamine compound and the medical material, such as hydrogen bonding or electrostatic interaction, to decorate the polyamine compound onto the substrate surface, so that the polymer properties of the polyamine compound are retained. In addition, the polyamine compound has strong hydrophilicity, and a large number of amino groups on the polyamine compound can participate in the oxidative self-polymerization of the catechol compound, thereby promoting the reaction, and the good hydrophilicity of the polyamine compound can inhibit the agglomeration of particles, which can improve the stability of the anticoagulant and antibacterial coating finally prepared.

[0059] After step S1 is completed, according to the present invention, the medical material coated with the phenolamine base coating is soaked in an anticoagulant solution, and after reaction, a medical material having an anticoagulant coating and a base coating is obtained. In the present invention, the anticoagulant includes heparin and / or tissue plasminogen. The concentration of the anticoagulant solution is 1 to 3 mg / mL, preferably 1.5 to 2.5 mg / mL, and the solvent includes any one or more of water, dimethyl sulfoxide, and ethanol.

[0060] In some embodiments of the present invention, it is preferred to prepare a solution of an anticoagulant substance, and soak the medical material obtained in step S1 in a solution of an anticoagulant drug at 20 to 40°C, preferably 25 to 37°C, and adjust the pH to 4 to 6 to activate the anticoagulant drug. The solution is immersed for 0.1 to 30 hours, preferably 0.1 to 10 hours, and after washing, an anticoagulant-modified medical material is obtained.

[0061] In some preferred embodiments of the present invention, the anticoagulant in the anticoagulant solution is activated, so that the phenolamine base coating is grafted with the activated anticoagulant, making the loading of the anticoagulant more stable and firm, and the grafting of the anticoagulant provides active sites for subsequent grafting.

[0062] Exemplarily, taking heparin as an example, in some embodiments of the present invention, the activation process of heparin includes the following steps:

[0063] Step (1): dissolving heparin sodium in a solvent to obtain a heparin sodium solution;

[0064] Step (2): dissolving 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS) in a solvent to obtain a mixed solution of EDC and NHS;

[0065] Step (3): adding the mixed solution of EDC and NHS to the sodium heparin solution, reacting at room temperature for 10 to 60 minutes to fully activate the sodium heparin so as to facilitate further grafting onto the medical material coated with the base coating;

[0066] Step (4), adding the medical material grafted with the phenolamine base coating to the activated heparin sodium solution for 0.1 to 30 hours, taking it out, shaking and washing it for 10 to 15 hours, and then drying it.

[0067] In the above step (3), the mass ratio of EDC to NHS is (0.3-0.8):1; and / or, the mass ratio of heparin sodium to EDC is (0.3-0.8):1; and / or, the mass ratio of heparin sodium to NHS is (0.1-0.5):1; when the EDC / NHS mixed solution is mixed, the pH is adjusted to between 4 and 6.

[0068] The above-mentioned heparin activation process uses EDC and NHS to activate heparin, so that the carboxyl groups on the heparin have reactive activity, thereby forming covalent bonds with the phenolamine base coating through amide reaction, thereby promoting the coating of heparin, which is firm and stable, and the reaction conditions are simple.

[0069] After step S2 is completed, according to the present invention, a medical material modified by an anticoagulant drug is soaked in a metal salt solution, and after reaction, a medical material having an antibacterial and anticoagulant coating is obtained. The soaking temperature is 20 to 40°C, preferably 25 to 37°C; the time is 0.1 to 30 hours, preferably 0.1 to 10 hours. In this step, the soaking is preferably carried out under a pH of 8 to 10 to promote the hydrolysis of the metal salt solution into metal oxides.

[0070] In the present invention, the metal salt in the metal salt solution is any one or more of titanium metal salt, zinc metal salt or tin metal salt. The titanium metal salt is selected from TiBALDH and / or tetrabutyl titanate, the zinc metal salt is selected from zinc acetate dihydrate and / or zinc nitrate, and the tin metal salt is selected from tin chloride dihydrate and / or tin acetate, tin acetate. The corresponding volume percentage of the metal salt solution is 2-6% (v / v), preferably 3-5% (v / v).

[0071] It should be noted that a single metal compound is difficult to dissolve in a solvent during the coating process, and there is a certain degree of dispersion difficulty, and problems such as uneven dispersion and unstable binding often occur. Therefore, the present invention uses a salt solution of a metal compound, which can promote its dispersion and make it easier to combine with an anticoagulant substance, thereby achieving an antibacterial effect under the action of photocatalysis. The method is simple and a uniform and stable coating can be easily prepared.

[0072] Taking the metal salt solution as a titanate solution as an example, in some embodiments of the present invention, the reaction process of the metal salt solution includes the following steps:

[0073] Step 1: dissolving titanium diammonium di(2-hydroxypropionic acid) dihydroxide (TiBALDH) in aqueous ammonia to form a titanium dioxide sol-gel;

[0074] Step 2: Add the anticoagulant drug-modified medical material into the titanium dioxide sol-gel for 18 to 30 hours, take it out, shake and wash it for 10 to 15 hours, and then dry it.

[0075] Wherein, the volume ratio of TiBALDH to ammonia water is (0.02-0.06):1, (0.02-0.04):1, and more preferably 0.02:1; and / or the concentration of ammonia water is 0.1-2 mol / L, preferably 0.1-1 mol / L, and more preferably 0.1 mol / L; and / or the mass concentration of TiBALDH is 20-60 wt% (w / v), preferably 50 wt% (w / v).

[0076] In other embodiments of the present invention, the reaction process of the metal salt solution comprises the following steps:

[0077] Metatitanic acid is dissolved in a hydrochloric acid solution to form a titanium dioxide gel. Then, the medical material modified with the anticoagulant drug is added to the titanium dioxide sol gel for 18 to 30 hours, taken out and washed by shaking for 10 to 15 hours, and then dried.

[0078] The dissolution temperature is 50-100°C, preferably 80°C; the dissolution time is 0.5-2h, preferably 1h; the hydrochloric acid concentration is 0.01-0.1mol / L, preferably 0.05-0.1mol / L, more preferably 1mol / L.

[0079] In the above-mentioned titanium dioxide coating process, titanate (such as TiBALDH) is hydrolyzed into titanium dioxide sol-gel, and then metal-carboxyl coordination bonds are generated with activated anticoagulants (such as heparin carboxyl) to achieve titanium dioxide grafting. At the same time, the titanium dioxide coating can regulate the concentration of anticoagulants (such as heparin), thereby promoting the proliferation of endothelial cells, promoting the endothelialization of coating materials, improving biocompatibility, and meeting the surface modification required during the use of blood-contact materials.

[0080] It should be noted that when titanium dioxide is exposed to ultraviolet light, ROS can be produced under the irradiation of ultraviolet light, which has a significant killing ability for both Gram-negative and Gram-positive bacteria. Titanium dioxide has good hydrophilic lubrication ability and can effectively prevent bacterial adhesion.

[0081] In summary, in some embodiments of the present invention, the method for preparing the antibacterial and anticoagulant coating comprises the following steps:

[0082] Step 1: preparing a solution containing catechol compounds and polyamine compounds, immersing the medical material in the solution containing catechol compounds and polyamine compounds to form a base coating, and obtaining the medical material containing the base coating after washing;

[0083] Step 2: preparing a solution of an anticoagulant substance, soaking the medical material obtained in step 1 in the solution of an anticoagulant drug, adjusting the pH to 4 to 6, soaking for 0.1 to 10 hours, and obtaining the anticoagulant-modified medical material after washing;

[0084] Step 3: Soak the medical material obtained in step 2 in a metal salt solution, adjust the pH to 8-10, soak for 0.1-10 hours, take out, wash and dry, and obtain a medical material with an antibacterial and anticoagulant coating.

[0085] In some embodiments of the present invention, the catechol compound is dopamine hydrochloride; the polyamine compound is polyethyleneimine; the anticoagulant drug is heparin; and the metal salt solution is titanate solution.

[0086] Based on the technical scheme, compared with the prior art, in the preparation method of the multifunctional coating with antibacterial and anticoagulant capabilities provided by the present invention, the base material (i.e., medical material) is subjected to a Schiff base reaction under weak alkaline conditions using polyethyleneimine and dopamine to accelerate the co-deposition of dopamine and polyethyleneimine to form a layer of polyphenol-amine bottom coating, and then the heparin after the activated carboxyl group is amidated and interacted with the residual amino group by charge, thereby fixing the heparin, and finally titanium dioxide is grafted on the surface through the coordination of the carboxyl group and the metal ion, and then a multifunctional coating composition is obtained. Through a simple and efficient reaction process, a stable and firm coating composition is prepared, which provides a new idea for the preparation of a multifunctional coating.

[0087] The present invention also provides a medical material, which includes a medical material substrate and a coating formed on the surface of the medical material substrate, wherein the coating is the above-mentioned antibacterial and anticoagulant coating.

[0088] In some embodiments of the present invention, the medical material matrix includes a blood contact material matrix; the material of the medical material matrix is ​​selected from metal materials, inorganic non-metallic matrix materials or polymer materials (such as thermoplastic polyurethane, polypropylene, etc.). The thickness of the antibacterial and anticoagulant coating is 20 μm.

[0089] To further illustrate the present invention, the following examples are used to provide a detailed description. The thermoplastic polyurethane used in the following examples of the present invention was purchased from Covestro Bayer, 285A, and the polypropylene was purchased from Formosa Plastics Industry (Ningbo) Co., Ltd., 5600XT.

[0090] Example 1

[0091] (1) Preparation of substrate: Preheat the flat plate vulcanizer to 130°C for the upper plate and 140°C for the lower plate. Weigh 25g of thermoplastic polyurethane pellets and put them into the mold. Put the mold into the flat plate vulcanizer and compress it at a pressure of 7MPa for 7min. Then adjust the flat plate vulcanizer to repeatedly squeeze the mold to remove the air in the thermoplastic polyurethane. Then compress it at a pressure of 10MPa for 5min. Then take it out and put it into the cold press. Cool it down at room temperature and shape it at a pressure of 10MPa. After 10min, take out the mold to obtain the thermoplastic polyurethane substrate.

[0092] (2) Substrate cleaning: The thermoplastic polyurethane substrate was placed in a mixed solution of ethanol: water = 1:1 (volume ratio), ultrasonically cleaned for 5 min, and then taken out and dried at room temperature for later use;

[0093] (3) Preparation of Tris buffer solution: Dissolve 6.057 g of tris(hydroxymethyl)aminomethane in 1 L of deionized water, and adjust the pH to 8.5 using 1 mol / L hydrochloric acid solution and 1 mol / L sodium hydroxide solution;

[0094] (4) Preparation of phenolamine base coating: 0.11g PEI (Mw=600) was dissolved in 50mL Tris buffer. After PEI was completely dissolved, 0.1g DA was added to obtain a mixed deposition solution of DA and PEI. The thermoplastic polyurethane substrate was immersed in the mixed deposition solution and reacted at 37°C for 12h to complete co-deposition. After being placed in ultrapure water for 12h for shaking and washing, it was dried in a vacuum oven at 25°C to constant weight to obtain a thermoplastic polyurethane substrate coated with a phenolamine base coating;

[0095] (5) Grafting of heparin: 0.1917 g of EDC and 0.2877 g of NHS were dissolved in 50 mL of 0.1 mol / L MES buffer solution, and 0.05 g of sodium heparin was added. The mixture was reacted at room temperature for 0.5 h to fully activate the carboxyl groups of heparin. Then, a thermoplastic polyurethane substrate coated with a phenolamine primer was added, and the mixture was shaken in a constant temperature shaker at 37°C for 24 h. The mixture was taken out and washed with ultrapure water for 12 h, and dried in a vacuum oven at 25°C to constant weight to obtain a heparinized thermoplastic polyurethane substrate.

[0096] (6) Grafting of titanium dioxide: 48 μL of TiBALDH was dissolved in 1 mL of 0.01 mol / L ammonia solution and added to a heparinized thermoplastic polyurethane substrate. The mixture was oscillated at 37°C for 24 h, then taken out and placed in ultrapure water for 12 h for washing. The mixture was dried in a vacuum oven at 25°C to constant weight to obtain a thermoplastic polyurethane substrate with a dopamine / polyethyleneimine-heparin-titanium dioxide coating.

[0097] The dopamine / polyethyleneimine-heparin-titanium dioxide coating utilizes polyethyleneimine to participate in the oxidative self-polymerization of dopamine, thereby promoting the deposition of dopamine and constructing a phenolamine base coating, and then covalently grafts heparin through an amide reaction, and finally coordinates titanium dioxide through carboxyl groups to construct a multifunctional coating with both antibacterial and anticoagulant properties on the surface of a thermoplastic polyurethane substrate, which can be applied to blood-contact devices.

[0098] Compared with the existing heparin coating, this multifunctional coating has a more obvious anticoagulant effect due to the improvement of hydrophilicity by titanium dioxide. It can significantly reduce platelet adhesion, activation and aggregation, and can also reduce the risk of local thrombosis after being used to prepare blood-contact devices. In addition, since heparin is fixed on the phenolamine bottom coating by covalent bonding, it will not enter the body during use, and thus will not affect the patient's systemic coagulation function, and has high safety performance. At the same time, because titanium dioxide modifies the coating, photocatalysis occurs under ultraviolet light, generating ROS to destroy bacterial membranes and kill bacteria, which is safe and efficient and has potential for clinical application.

[0099] Example 2

[0100] (1) Preparation of substrate: Preheat the flat plate vulcanizer to 130°C for the upper plate and 140°C for the lower plate. Weigh 25g of polypropylene pellets and put them into the mold. Put the mold into the flat plate vulcanizer and compress it at a pressure of 7MPa for 7min. Then adjust the flat plate vulcanizer to repeatedly squeeze the mold to remove the air in the thermoplastic polypropylene. Then compress it at a pressure of 10MPa for 5min, take it out and put it into the cold press. Cool it down at 25°C and shape it at a pressure of 10MPa. After 10min, take out the mold to obtain a polypropylene substrate.

[0101] (2) Substrate cleaning: Place the polypropylene substrate in an acetone solution and ultrasonically clean it for 5 minutes. Take it out and dry it at room temperature for later use;

[0102] (3) The preparation of Tris buffer solution is the same as step (3) of Example 1;

[0103] (4) Preparation of phenolamine base coating: 0.11g PEI (Mw=600) was dissolved in 50mL Tris buffer. After PEI was completely dissolved, 0.1g norepinephrine was added to obtain a mixed deposition solution of norepinephrine and PEI. The polypropylene substrate was immersed in the mixed deposition solution and reacted at 37°C for 12h to complete co-deposition. The substrate was then placed in ultrapure water for 12h for shaking and washing, and then dried in a vacuum oven at 25°C to constant weight to obtain a polypropylene substrate coated with a phenolamine base coating;

[0104] (5) Grafting of heparin is the same as step (5) of Example 1;

[0105] (6) Grafting of zinc oxide: 0.5% low molecular weight polyethylene glycol (Mw=300) was added to 20 mL of 0.025 mol / L NaOH at room temperature, and then 15 mL of 1 mg / mL zinc acetate dihydrate solution was added dropwise, and the heparinized thermoplastic polypropylene substrate was added and stirred for 3 h. Then, the substrate was taken out and placed in ultrapure water for 12 h for shaking and washing, and dried in a vacuum oven at 25°C to constant weight.

[0106] Example 3

[0107] (1) The substrate is prepared in the same manner as step (1) of Example 1;

[0108] (2) Cleaning the substrate is the same as step (2) of Example 1;

[0109] (3) The preparation of Tris buffer solution is the same as step (3) of Example 1;

[0110] (4) Preparation of phenolamine base coating: 0.11g PEI (Mw=600) was dissolved in 50mL Tris buffer. After PEI was completely dissolved, 0.1g phenylalanine was added to obtain a mixed deposition solution of phenylalanine and PEI. The thermoplastic polyurethane substrate was immersed in the mixed deposition solution and reacted at 37°C for 12h to complete co-deposition. The substrate was then placed in ultrapure water for 12h for shaking and washing, and then dried in a vacuum oven at 25°C to constant weight to obtain a thermoplastic polyurethane substrate coated with a phenolamine base coating;

[0111] (5) Grafting of heparin is the same as step (5) of Example 1;

[0112] (6) Grafting of tin dioxide: 0.407 g of tin chloride dihydrate was dissolved in 5 mL of 2 mol / L HCl solution to obtain a precursor sol. Then, a heparinized thermoplastic polyurethane substrate was added and the reaction was carried out at 75°C for 18 h. The sol was then taken out and placed in ultrapure water for 12 h for washing and dried in a vacuum oven at 25°C to constant weight.

[0113] Example 4

[0114] (1) The substrate is prepared in the same manner as step (1) of Example 1;

[0115] (2) Cleaning the substrate is the same as step (2) of Example 1;

[0116] (3) The preparation of Tris buffer solution is the same as step (3) of Example 1;

[0117] (4) Preparation of phenolamine base coating: 0.11g PEI (Mw=600) was dissolved in 50mL Tris buffer. After PEI was completely dissolved, 0.1g phenylalanine was added to obtain a mixed deposition solution of phenylalanine and PEI. The thermoplastic polyurethane substrate was immersed in the mixed deposition solution and reacted at 37°C for 12h to complete co-deposition. It was then placed in ultrapure water for 12h for shaking and washing, and then dried in a vacuum oven at 25°C to constant weight to obtain a thermoplastic polyurethane substrate coated with a phenolamine base coating;

[0118] (5) Grafting of tissue plasminogen: 0.1917 g EDC and 0.2877 g NHS were dissolved in 50 mL 0.1 mol / L MES buffer solution, and 0.05 g recombinant tissue plasminogen activator was added. The mixture was reacted at room temperature for 0.5 h to fully activate the carboxyl groups of heparin. A thermoplastic polyurethane substrate coated with an amide primer was then added. The mixture was shaken in a constant temperature shaker at 37 °C for 24 h. The mixture was then taken out and washed with ultrapure water for 12 h, and dried in a vacuum oven at 25 °C to constant weight.

[0119] (6) Grafting of titanium dioxide is the same as step (6) in Example 1.

[0120] Example 5

[0121] In this embodiment, steps (1)(2)(3)(4)(5) are the same as those in Embodiment 1, and step (6) is the same as that in Embodiment 2.

[0122] Example 6

[0123] In this embodiment, steps (1)(2)(3)(4)(5) are the same as those in Embodiment 1, and step (6) is the same as that in Embodiment 3.

[0124] Example 7

[0125] In this embodiment, steps (1), (2), (3), (5), and (6) are the same as those in Embodiment 1, and step (4) is the same as that in Embodiment 2.

[0126] Example 8

[0127] In this embodiment, steps (1)(2)(3)(5)(6) are the same as those in Example 1, and step (4) is the same as that in Example 3.

[0128] Comparative Example 1 (modification without heparin and titanium dioxide)

[0129] The difference between this comparative example and Example 1 is that the grafting of heparin and titanium dioxide is not performed, that is, steps (5) and (6) are not performed, and dopamine and polyethyleneimine are directly fixed on the surface of the thermoplastic polyurethane substrate by co-deposition.

[0130] Comparative Example 2 (modification without titanium dioxide)

[0131] The difference between this comparative example and Example 1 is that titanium dioxide grafting is not performed, that is, step (6) is not performed, and heparin is grafted onto the surface of the thermoplastic polyurethane substrate by utilizing the amide reaction of the phenolamine base coating and heparin.

[0132] Comparative Example 3 (Modification without heparin)

[0133] The difference between this comparative example and Example 1 is that no heparin modification is performed, and titanium dioxide is directly grafted onto the surface of the thermoplastic polyurethane substrate through the coordination of catechol of dopamine on the phenolamine base coating with titanium dioxide.

[0134] Comparative Example 4

[0135] Provided is a thermoplastic polyurethane substrate plate obtained in step (1) of Example 1.

[0136] Comparative Example 5 (direct use of nano titanium dioxide)

[0137] (1) The substrate is prepared in the same manner as step (1) of Example 1;

[0138] (2) Cleaning the substrate is the same as step (2) of Example 1;

[0139] (3) The preparation of Tris buffer solution is the same as step (3) of Example 1;

[0140] (4) Preparation of phenolamine base coating: 0.11g polyacrylamide was dissolved in 50mL Tris buffer, and after it was completely dissolved, 0.1g dopamine was added to obtain a mixed deposition solution of dopamine and PEI. The polyurethane substrate was immersed in the mixed deposition solution, reacted at 37°C for 12h to complete co-deposition, and then placed in ultrapure water for 12h for shaking and washing, and then dried in a vacuum oven at 25°C to constant weight to obtain a thermoplastic polyurethane substrate coated with a phenolamine base coating;

[0141] (5) Grafting of heparin is the same as step (5) of Example 1;

[0142] (6) Grafting of titanium dioxide: 0.5 g of nano-titanium dioxide was added to 10 mL of deionized water, ultrasonically dispersed for 10 min, added to a heparinized thermoplastic polyurethane substrate, and oscillated at 37 °C for 24 h. The substrate was then taken out and placed in ultrapure water for 12 h for oscillation and washing, and dried in a vacuum oven at 25 °C to constant weight.

[0143] Performance Testing

[0144] (1) Hemolysis rate test

[0145] The test method is:

[0146] Take fresh whole blood and centrifuge it at 4°C and 1500rpm for 15min, discard the supernatant, take the lower layer and centrifuge it at 1500rpm in normal saline for 3min, and wash the red blood cells until the supernatant is clear. The red blood cells are diluted with normal saline to a volume fraction of 4%. After the sample (the product obtained in Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 4) is cut into 1×1cm, it is placed in 500μL red blood cell suspension and an equal amount of normal saline and incubated at 37°C for 1h, and then the incubated red blood cells are collected and centrifuged at 2000rpm for 3min, the supernatant is extracted, and the absorbance is measured at 540nm. Set the negative control group to add 500μL red blood cell suspension and an equal amount of normal saline, and the positive control group to add 500μL red blood cell suspension and an equal amount of deionized water, and no sample is added.

[0147] Finally, the following formula is used for calculation:

[0148] Hemolysis rate (%) = (OD 实验组 -OD 阴性对照组 ) / (OD 阳性对照组 -OD 阴性对照组 )×100%.

[0149] The comparison of the hemolysis rates of the products obtained in Example 1, Comparative Example 1, Comparative Example 2 and Comparative Example 4 is shown in the figure below. Figure 1 As shown, Figure 1 In the figure, from left to right, the six tubes correspond to Example 1, Comparative Example 1, Comparative Example 2, Comparative Example 4, positive control group, and negative control group, respectively.

[0150] (2) Anti-protein adhesion ability

[0151] When blood-contact materials come into contact with vascular endothelial cells, activation of the coagulation cascade and complement system often occurs. In this process, the adhesion of various proteins is considered to be the first step of the coagulation cascade reaction. The anti-protein adhesion ability is evaluated by measuring the content of fibrinogen (Fib) and bovine serum albumin (BSA) adsorbed on the sample surface using the BCA method.

[0152] The test results are shown in Table 1 and Figure 2 (Comparative diagram of the evaluation of the anti-protein adhesion of the products obtained in Example 1, Comparative Example 1, Comparative Example 2 and Comparative Example 4) is shown.

[0153] (3) Antiplatelet adhesion ability

[0154] Platelet adhesion, activation and diffusion are the main causes of thrombus formation. The platelet adhesion ability of blood-contact materials is an important indicator for evaluating their anticoagulant ability. In the present invention, PRP (platelet-rich plasma) is incubated on samples (products obtained in Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 4) for 2 hours, then fixed with paraformaldehyde, and then dehydrated with ethanol gradient to observe the adhesion amount of platelets on the sample surface using a scanning electron microscope.

[0155] The platelet adhesion electron microscopic comparison diagrams of the products obtained in Example 1, Comparative Example 1, Comparative Example 2 and Comparative Example 4 are shown in Figure 3 shown.

[0156] Depend on Figure 3 It can be seen that the product obtained in Example 1 has excellent anti-platelet adhesion ability. In Example 1, the hydrophilicity of the surface of the substrate material (i.e., medical material) is enhanced due to the grafting of titanium dioxide. Since the surface has a high affinity for water, when in contact with water, the coating surface will be directly connected to the water molecules through hydrogen bonds, so that the water molecules are dispersed on the surface to form a hydration layer. When BSA and Fib contact the coating surface, due to the presence of the hydration layer, platelets cannot adhere, thereby effectively inhibiting the further progress of the coagulation cascade reaction and playing an anticoagulant role. In addition, heparin can participate in endogenous coagulation, inhibit coagulation factor Xa and thrombin, thereby inhibiting the activation of platelets, and further improving the anticoagulant ability.

[0157] (4) Thrombin time test

[0158] The activated partial thromboplastin time (APTT) and thrombin time (TT) tests were performed on the obtained product, specifically measured by APTT kit.

[0159] The test results are shown in Table 1 and Figure 4 (Comparison diagram of Example 1, Comparative Example 1, Comparative Example 2, Comparative Example 4) is shown.

[0160] (5) Antibacterial rate test

[0161] Infection is the main complication of all implantable materials during use. The present invention conducted 12 groups of control experiments, and the materials used were the products obtained in Examples 1 to 9, Comparative Example 1, Comparative Example 2, and Comparative Example 4, and a blank group (i.e., only bacterial solution) was set. The antibacterial rate was obtained by the plate coating method, specifically:

[0162] 8×10 7 CFU / mL of bacterial solution (Staphylococcus aureus (S. aureus) or Escherichia coli (E. coli)) was co-cultured with materials from different groups, and then incubated in agar medium at 37°C for 24 hours, the number of colonies was observed, and the difference in the number of colonies between the blank group and the experimental group was used to calculate the inhibition rate;

[0163] The formula for calculating the inhibition rate is: (the number of colonies in the blank group - the number of colonies in the experimental group) / the number of colonies in the blank group);

[0164] Test results such as Figure 5 As shown in (Comparison of antibacterial rates of Example 1, Comparative Example 1 and Comparative Example 2) and Table 1, it can be seen that the antibacterial ability of the product obtained in Example 1 is better than that of other control groups and has good application potential.

[0165] (6) Stability test

[0166] After the products obtained in Examples 1 to 9, Comparative Example 1, Comparative Example 2 and Comparative Example 4 were immersed in PBS for 7 days, the constituent elements of the coatings were analyzed to evaluate their stability.

[0167] The specific evaluation criteria are: observe whether the element content decreases. For example, for titanium dioxide coating, if the titanium content decreases, it means that the coating is damaged during the immersion process, which proves that it is not stable enough. On the contrary, if the content does not decrease, it means that the coating is stable.

[0168] The test results are shown in Table 1.

[0169] Depend on Figures 1 to 6 It can be seen that Example 1 has good absorption capacity in the ultraviolet light region, can absorb ultraviolet light to show photocatalytic properties, and has less protein adhesion, a higher antibacterial rate, and a longer coagulation time, showing good antibacterial and anticoagulant capabilities.

[0170] (7) Visible light absorption peak test

[0171] The products obtained in Example 1 and Comparative Example 4 were tested for visible light absorption peaks. The results are as follows: Figure 6 shown.

[0172] (8) Cytotoxicity

[0173] Natural endothelial cells are considered to be the most ideal surface for blood-contact materials. Blood-contact devices with rapid endothelialization have unlimited potential in clinical applications. Human umbilical vein endothelial cells (HUVEC) were used to evaluate the rapid endothelialization ability of the sample surface. After observing the growth state of HUVEC under a microscope, the culture medium in the culture dish was discarded, washed twice with PBS, and 1 mL of pancreatic enzyme solution containing EDTA was added. Then, it was placed in an incubator for digestion for 3 minutes. After being taken out, an appropriate amount of culture medium was added to terminate the digestion. After centrifugation at 1000 rpm for 5 minutes, the supernatant was discarded, and the culture medium was added to mix the cells by blowing. Finally, the culture medium containing the cells was added to a 6-well plate so that each well plate contained about 5×10 3The samples were cut into 1×1 cm pieces and placed in the well plates for 24 h and 48 h. Cell viability was detected by CCK-8 method.

[0174] The cell viability data of the products obtained in Example 1, Comparative Example 1, Comparative Example 2 and Comparative Example 4 are shown in the figure below. Figure 7 shown.

[0175] It can be seen that the cell viabilities of Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 4 were all greater than 100%, indicating that they were harmless to cells.

[0176] Table 1

[0177]

[0178]

[0179] It can be seen from the data in Table 1 that compared with Examples 5 to 7, Example 1 has less protein adhesion and a longer thrombin time, showing better anticoagulant ability. Although the antibacterial rate has decreased, it has shown a relatively good bactericidal effect as a whole. Compared with Example 9, all performances have been enhanced. After comprehensive evaluation, the antibacterial and anticoagulant ability of Example 1 is better.

[0180] The catechol compounds used in Examples 3 and 6 are phenylalanine (levodopa), which has an additional carboxyl group compared to dopamine hydrochloride, has greater steric hindrance, and has a poorer self-polymerization efficiency. Under the same time (4h), the stability of the obtained coating will be poor.

[0181] Comparative Example 4 is a blank thermoplastic polyurethane substrate without modification of anticoagulant and antibacterial substances. It can be seen that without modification of titanium dioxide and heparin, it exhibits poor performance.

[0182] Comparative Example 1 is based on Comparative Example 4 and has a phenolamine intermediate layer. Due to its improved hydrophilicity and the charge carried by a portion of polyethyleneimine, its ability to resist protein adhesion and antibacterial properties is enhanced, but it is still insufficient and does not have good anticoagulant ability (thrombin time). After further modification of heparin (Comparative Example 2), its anticoagulant ability is improved, but due to the charge balance with the charge carried by the phenolamine intermediate layer, the antibacterial rate is reduced.

[0183] Comparative Example 5 directly uses nano titanium dioxide powder instead of hydrolyzing it with a metal salt solution, which results in uneven dispersion and poor coating effect, thereby significantly reducing the antibacterial rate.

[0184] In summary, the antibacterial and anticoagulant coating provided by the present invention, in which heparin provides excellent anticoagulant ability, titanium dioxide provides excellent bactericidal performance, and further improves hydrophilicity, thereby improving the antibacterial and anticoagulant abilities as a whole. However, Comparative Example 2 does not have titanium dioxide, and has insufficient bactericidal performance. Comparative Example 1 does not have titanium dioxide and heparin, and has insufficient antibacterial and anticoagulant abilities. Comparative Example 4 does not have any modification, and has insufficient hydrophilicity, antibacterial and anticoagulant abilities.

[0185] The above description of the disclosed embodiments enables those skilled in the art to implement or use the present invention. Various modifications to these embodiments will be apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to the embodiments shown herein, but rather to the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. An antibacterial and anticoagulant coating composition, characterized in that: These include catechol compounds, polyamine compounds, anticoagulants and metal salts; The mass ratio of the catechol compound, the polyamine compound, the anticoagulant substance and the metal salt is (35-45):(35-45):(5-12):(10-20).

2. The antibacterial and anticoagulant coating composition according to claim 1, characterized in that: The catechol compounds include any one or more of catechol, catechol acid, catecholamine or flavonoid compounds; The polyamine compound includes any one or more of polyimine, polyamide or polyurethane; The anticoagulant substance includes heparin and / or tissue plasminogen; The metal salt includes any one or more of titanium metal salt, zinc metal salt or tin metal salt.

3. The antibacterial and anticoagulant coating composition according to claim 1 or 2, characterized in that: The catechol compound is selected from any one or more of levodopa, norepinephrine, dopamine, dopamine hydrochloride or phenylalanine; The polyamine compound is selected from polyethyleneimine and / or polyacrylamide; The titanium metal salt is selected from TiBALDH and / or tetrabutyl titanate, the zinc metal salt is selected from zinc acetate dihydrate and / or zinc nitrate, and the tin metal salt is selected from tin chloride dihydrate and / or tin acetate.

4. An antibacterial and anticoagulant coating, characterized in that: The coating is formed from the antibacterial and anticoagulant coating composition according to any one of claims 1 to 3.

5. A method for preparing an antibacterial and anticoagulant coating, characterized in that: The following steps are involved: S1: soaking the medical material in a solution containing a catechol compound and a polyamine compound, and after reaction, obtaining a medical material coated with a phenolamine base coating; S2: soaking the medical material coated with the phenolamine base coating with an anticoagulant solution, and obtaining the medical material modified with the anticoagulant after reaction; S3: soaking the medical material modified by the anticoagulant substance in a metal salt solution, and obtaining the medical material with an antibacterial and anticoagulant coating after the reaction.

6. The preparation method according to claim 5, characterized in that: The soaking temperatures in steps S1, S2 and S3 are independently 20 to 40°C, and the soaking times are independently 0.1 to 30 hours.

7. The preparation method according to claim 5 or 6, characterized in that: The soaking in step S2 is performed under a pH condition of 4 to 6; The anticoagulant in the anticoagulant solution in step S2 is activated; The soaking in step S3 is performed under the condition of pH 8-10.

8. A medical material, characterized in that: It comprises a medical material matrix and a coating formed on the surface of the medical material matrix; The coating is the antibacterial and anticoagulant coating according to claim 4 or the antibacterial and anticoagulant coating obtained by the preparation method according to any one of claims 5 to 7.

9. The medical material according to claim 9, characterized in that: The medical material matrix includes a blood contact material matrix; The material of the medical material matrix is ​​selected from metal materials, inorganic non-metallic matrix materials or polymer materials.

10. The medical material according to claim 8 or 9, characterized in that: The thickness of the antibacterial and anticoagulation coating is 10 to 25 μm.

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