Preparation method and application of surface active coating

Abstract

The invention discloses a preparation method and application of a surface active coating, and belongs to the technical field of artificial joint coatings. A magnesium-doped porous titanium dioxide nanotube array layer and a zinc-strontium co-doped mesoporous bioactive glass release layer are formed on the surface of a substrate; the single performance such as the bonding strength, the osteogenic activity and the antibacterial ability of the coating is exceeded, and more importantly, the unification and the continuous high efficiency of the performance are realized through the gradient and synergistic system design of the coating; a new solution is provided for solving the two major problems of artificial joint replacement, namely sterile looseness and antibacterial ability, and the method has extremely high clinical transformation value and market prospect.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of artificial joint coating, in particular to a preparation method of a surface active coating and application thereof. BACKGROUND

[0002] Artificial joints are implantable prostheses made of metal materials with good biocompatibility and physiological joints by engineering methods to achieve the purpose of removing lesions, eliminating pain and restoring joint activity and function. At present, artificial hip, knee, shoulder and elbow joint replacement surgery has become one of the most successful surgical methods for treating end-stage joint diseases, which significantly improves the quality of life of patients. The long-term stability of artificial joints, artificial implants, bone pins and spinal fusion devices depends on the formation of firm and durable bone integration between the implants and the host bone. However, the surface of a simple metal prosthesis is usually biologically inert, which is difficult to form effective biological bonding with bone tissue. Initially, the fixation of the prosthesis mainly depends on the mechanical embedding of bone cement. Although bone cement can provide good immediate fixation, it has the risk of aging and fragmentation in the long term, and may cause complications such as allergic reactions. In order to achieve more ideal biological fixation, surface modification technology has developed rapidly. By constructing a specific coating on the surface of the prosthesis, the surface properties can be significantly improved to guide bone tissue to grow in and integrate with it, while maintaining the mechanical properties of the prosthesis matrix.

[0003] Although the coating technology has developed greatly, there are still defects: 1. The interface bonding force of traditional macrostructure coating is weak, and the coating is prone to peeling and falling off from the interface during long-term service. The degradation rate does not match the new bone formation rate, resulting in premature loss of coating function. It lacks active antibacterial ability. 2. High-dose antibiotics or silver ions in a single functional coating can effectively resist bacteria, but may be toxic to osteoblasts, inhibit bone integration, and have a functional conflict between "antibacterial" and "osteogenic". Such coatings often face the problems of initial burst release and subsequent failure, and cannot achieve long-term and controllable release. Simple physical mixing or single-layer structure cannot coordinate the contradiction between the spatial distribution and release kinetics of multiple functional factors.

[0004] To solve the bonding problem, patent CN112545711A discloses a femoral stem prosthesis and a preparation method thereof. The bioactive coating includes a porous titanium coating and a hydroxyapatite coating. Although the bonding strength is improved, the double-layer coating attempts to balance the firm bonding of the titanium coating and the bioactivity of HA, but the problem of fibrous tissue growth at the distal end still exists, which may cause postoperative discomfort, and the antibacterial and osteogenic functions are still not coordinated and unified. SUMMARY

[0005] The application aims to provide a preparation method of a surface active coating and application thereof, so as to solve the technical problems of single function, poor bone integration effect, and easy failure of the coating during long time use of the artificial joint, resulting in loosening.

[0006] To achieve the above-mentioned purpose, the technical scheme adopted by the application is as follows: a preparation method of a surface active coating is provided, which specifically comprises the following steps: (1) substrate pretreatment: ultrasonic cleaning and blow drying of titanium alloy to obtain a pretreated substrate; (2) inner layer preparation: S1, first anodic oxidation: placing the pretreated substrate in an electrolyte for oxidation reaction to form TiO2 nanotube array on the surface of the pretreated substrate, thereby obtaining a first substrate; S2, nanotube array stripping: ultrasonic treatment of the first substrate to strip the formed TiO2 nanotube array layer from the first substrate to form sites on the surface, thereby obtaining a second substrate; S3, second anodic oxidation: placing the second substrate in an electrolyte containing magnesium for oxidation reaction, and then performing crystallization treatment after the reaction to dope magnesium on the sites to form an inner layer, thereby obtaining a third substrate; (3) outer layer preparation: S1, preparation of precursor solution: mixing and stirring cetyltrimethylammonium bromide, zinc nitrate, strontium nitrate, and auxiliary components to obtain a precursor solution; S2, coating and self-assembly: immersing the third substrate in the precursor solution, taking it out after aging and heating; then immersing it in an aqueous solution containing BMP-2 mimetic peptide and polyacrylic acid after the temperature is lowered to room temperature, and drying to obtain a surface active coating.

[0007] In one embodiment, The specific steps of step (1) ultrasonic cleaning are as follows: sequentially ultrasonic cleaning in acetone, anhydrous ethanol, 10 vol% dilute hydrochloric acid, and deionized water for 15 min each time.

[0008] In one embodiment, In step (2) S1, the electrolyte is an ethylene glycol electrolyte containing 0.5 wt% NH4F and 2 vol% deionized water; and the voltage of the oxidation reaction is 60 V.

[0009] In one embodiment, In step (2) S2, the specific steps of ultrasonic treatment are as follows: immersing in a 15% H2O2 solution, and ultrasonic treatment at 150 W for 10 min.

[0010] In one embodiment, In step (two) S3, the electrolyte is a glycol electrolyte containing 0.5 wt% NH4F and 0.1 M magnesium acetate; the voltage of the oxidation reaction is 30-80 V.

[0011] In one embodiment, In step (two) S3, the specific steps of the crystallization treatment are as follows: the dried sample after the reaction is placed in a muffle furnace, heated to 450 °C at a heating rate of 5 °C / min in air, and then cooled with the furnace.

[0012] In one embodiment, In step (three) S1, the preparation of the precursor solution is as follows: cetyltrimethylammonium bromide is dissolved in a mixed solution of deionized water and anhydrous ethanol, and then tetraethyl orthosilicate, calcium nitrate tetrahydrate, zinc nitrate, strontium nitrate and triethyl phosphate are added in sequence, and stirred and mixed to prepare the precursor solution. The mass ratio of zinc nitrate to strontium nitrate is 1:1.0-2.0, and the pH of the precursor solution is 2.8-3.2.

[0013] In one embodiment, In step (three) S2, the specific heating steps are as follows: the aged sample is placed in a tube furnace, and heated to 200 °C at a rate of 1 °C / min under nitrogen protection, and then kept for 2 h.

[0014] In one embodiment, In step (three) S2, the mass ratio of the BMP-2 mimetic peptide to the polyacrylic acid is 1:2.

[0015] The application also provides an application of the surface-active coating, and the surface-active coating in any of the above embodiments is applied to an artificial joint, an artificial implant, a bone nail or a spinal fusion cage.

[0016] The application provides a preparation method of a surface-active coating and an application thereof, and has the following beneficial effects compared with the prior art: 1. In the two-layer structure, the magnesium ions in the inner layer focus on firm combination with the matrix and long-term osteogenesis; the outer layer serves as a buffer and control center, and cooperatively releases the osteogenic ion strontium and the mild antibacterial ion zinc; the gradient structure simulates the transition interface of the natural bone tissue in physical properties and chemical composition, avoids performance mutation, is more conducive to host cell recognition, adhesion and tissue ingrowth, is a biological fixation, significantly improves the stability, and solves the mutual inhibition problem of antibacterial and osteogenic functions; 2. Under normal physiological pH conditions, the outer layer's nanovalve (polyacrylic acid) is in a contracted state, releasing strontium ions and BMP-2 mimic peptides at a slow, constant rate, continuously nourishing bone tissue and promoting bone integration. Under infected conditions, when bacterial growth leads to acidification of the local microenvironment, the polyacrylic acid segments expand, opening mesoporous channels and accelerating the release of zinc ions, providing precise antibacterial action on the infected area and extending the coating's functional lifespan. The synergistic effect of multiple ions changes the strategy of relying on a single ion or drug, constructing a Mg-Zn-Sr ion synergistic network. Magnesium ions promote early osteoblast adhesion and proliferation, strontium ions strongly stimulate bone formation and inhibit osteoclast activity, and zinc ions provide broad-spectrum, long-lasting, and low-cytotoxic antibacterial capabilities, while also aiding in osteogenic processes. The release of these three ions matches different stages of the bone healing process, resulting in osteogenic efficiency far exceeding that of single-ion coatings. The outer layer's pH-responsive antibacterial action ensures that even if a small number of bacteria invade, they are confined by the nanotube structure and cannot penetrate deeply. 3. The inner TiO2 nanotube array of this application is grown in situ on the substrate by electrochemical method, forming a strong chemical bond and mechanical interlock with the substrate. Its bonding strength is better than that of traditional spray coatings, reducing the risk of aseptic loosening caused by coating peeling and providing mechanical protection for the service life of the coating. Attached Figure Description

[0017] To more clearly illustrate the technical solutions in the embodiments of this application, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0018] Figure 1 This is a partial SEM image of a TiO2 nanotube array; Figure 2 This is a BMP-2 mimic peptide release curve; Figure 3 This is a graph showing the number of bacterial colonies on a plate, as shown in Comparative Example 1. Figure 4 This is a graph showing the number of bacterial colonies on a plate in Example 1. Detailed Implementation

[0019] To make the technical problems, technical solutions, and beneficial effects to be solved by this application clearer, this application will be further described in detail. It should be understood that the specific embodiments described herein are only for explaining this application and are not intended to limit this application.

[0020] Example 1 (i) Pretreatment of the substrate: A medical Ti-6Al-4VELI titanium alloy was selected as the substrate material and cut into a test piece with a size of 10 mm x 10 mm x 1 mm; the test piece was sequentially ultrasonically cleaned in acetone, anhydrous ethanol, 10 vol% dilute hydrochloric acid, and deionized water for 15 min each to remove surface oil stains and natural oxidation films, and then dried with nitrogen to obtain a pretreated substrate; the surface roughness Ra of the pretreated substrate was 0.36 μm, and the contact angle was <30°. ° ; (ii) Preparation of the inner layer: (magnesium-doped porous titanium dioxide nanotube array) S1, first anodic oxidation: the pretreated substrate was used as the anode, a platinum electrode was used as the cathode, magnetic stirring was started, and the substrate was oxidized in an ethylene glycol electrolyte containing 0.5 wt% NH4F and 2 vol% deionized water at a room temperature and under a direct current voltage of 60 V for 2 h; after the reaction, the substrate was rinsed with deionized water and air-dried, and a layer of TiO2 nanotube array was formed on the surface of the pretreated substrate to obtain a first substrate; S2, peeling of the nanotube array: the first substrate was immersed in a 15% H2O2 solution and ultrasonically treated at 150 W for 10 min, the formed nanotube array layer was peeled off from the first substrate, and regular and ordered pits were left on the surface as the sites for the second oxidation; after the peeling, the substrate was ultrasonically cleaned with deionized water for 3 times (3 min each time) to remove residual H2O2, and a second substrate was obtained; S3, second anodic oxidation: the second substrate was used as the anode, and the substrate was oxidized in an ethylene glycol electrolyte containing 0.5 wt% NH4F and 0.1 M magnesium acetate at a room temperature and under a direct current voltage of 50 V for 1 h; after the reaction, the substrate was thoroughly rinsed with deionized water and vacuum dried at 80 °C for 2 h; crystallization treatment: the dried sample was placed in a muffle furnace, heated to 450 °C at a heating rate of 5 °C / min in air, and kept at the temperature for 2 h, and then cooled with the furnace; this process converted amorphous TiO2 into anatase and completed the doping of magnesium, forming the inner layer, and a third substrate was prepared; (iii) Preparation of the outer layer: (zinc / strontium co-doped mesoporous bioactive glass release layer) S1, preparation of the precursor solution: 2.0 g of cetyltrimethylammonium bromide (CTAB) was dissolved in a mixed solution of 100 mL of deionized water and 20 mL of anhydrous ethanol as a template agent, and then 10 mL of tetraethyl orthosilicate (TEOS), 0.4 g of calcium nitrate tetrahydrate, 0.12 g of zinc nitrate, 0.18 g of strontium nitrate, and 0.2 g of triethyl phosphate (TEP) were sequentially added, and the mixture was stirred vigorously for 4 h to allow complete hydrolysis, and the pH of the solution was controlled at 2.8-3.2 by 0.1 M hydrochloric acid to prepare a precursor solution; S2, coating and self-assembly: the third substrate was immersed in the precursor solution, and pulled out at a speed of 1 cm / min, so that the solution was uniformly attached to the inner and outer walls of the nanotube, and was placed in an environment of 50 °C for aging for 24 h; the aged sample was placed in a tube furnace, and was heated to 200 °C at a rate of 1 °C / min under nitrogen protection, and was kept for 2 h, so that the CTAB was removed, and the mesoporous structure was fixed; after being reduced to room temperature, the sample was immersed in an aqueous solution containing 50 mg of BMP-2 mimetic peptide (sequence: KIPKASSVPTELSAISTLYL, obtained by solid-phase synthesis) and 100 mg of polyacrylic acid (nanometer valve), and was vacuum-impregnated at 40 °C for 2 h, so that the mimetic peptide and the polyacrylic acid entered the mesoporous channels (the mesoporous pore size was tested by BET, and the average pore size was 8±0.7 nm); finally, the sample was vacuum-dried at 80 °C for 2 h, so that a surface-active coating coated with a two-layer structure was prepared.

[0021] Example 2 In this example, different from Example 1, in step (two) S3, the direct current voltage of the oxidation reaction was 30 V, and the rest of the operations were the same, so that a surface-active coating was prepared.

[0022] Example 3 In this example, different from Example 1, in step (two) S3, the direct current voltage of the oxidation reaction was 80 V, and the rest of the operations were the same, so that a surface-active coating was prepared.

[0023] Example 4 In this example, different from Example 1, the mass ratio of zinc nitrate to strontium nitrate was 1:1.0, and the rest of the operations were the same, so that a surface-active coating was prepared.

[0024] Example 5 In this example, different from Example 1, the mass ratio of zinc nitrate to strontium nitrate was 1:2.0, and the rest of the operations were the same, so that a surface-active coating was prepared.

[0025] Comparative Example 1 Medical Ti-6Al-4VELI titanium alloy plasma sprayed hydroxyapatite coating.

[0026] Comparative Example 2 Only the inner layer was prepared, and the outer layer was not prepared.

[0027] Experimental Example 1 SEM measurement of TiO2 nanotube array tube diameter: the coating morphology of the third substrate prepared in Example 1 was characterized by SEM, as shown in FIG. 1, a highly ordered nanotube array with clear openings was formed on the surface of the sample; the average inner diameter of the randomly selected nanotubes was 85±8 nm. Figure 1 ​

[0028] TiO2nanotube array tube length measurement: using the "spin coating + pull combined" process, first fix the sample on the rotating table, the speed is 500 rpm, drop the precursor solution (1 mL / cm2) and rotate for 10 s, then pull at a speed of 1 cm / min, the average tube length is 1.8 ± 0.1 μm.

[0029] Experimental example 2 BMP-2 mimetic peptide release: the final product prepared in example 1 was placed in PBS buffer and incubated in a constant temperature shaker at 37 °C to simulate the internal environment of the human body; a certain amount of release medium was collected on the 1st, 3rd, 7th, 14th, 21st, and 28th day, and fresh buffer was added in time to maintain the constant volume; the peptide concentration was quantitatively analyzed by HPLC, and the release curve was drawn as shown in Figure 2 The release rate of the BMP-2 mimetic peptide coated layer was 73 ± 2.06 % on the 12th day, and then entered a sustained slow release phase, with a cumulative release rate of 86 ± 1.43 % within 28 days, meeting the slow release requirements of bone repair.

[0030] Experimental example 3 The bonding strength pull test was carried out according to ASTM C633 standard, and the results were as follows: Comparative example 1: 35 MPa; comparative example 2: 60 MPa; example 1: 55 MPa, the fracture occurred in the adhesive or the outer layer, and the entire coating system was firmly combined with the substrate, meeting the requirements of the implant.

[0031] Experimental example 4 The in vitro wear test was carried out according to ASTM F732 standard, simulating the joint friction condition; the pin-on-disc friction and wear testing machine was used to install the substrate with coating prepared in example 1 on the testing machine as the fixed disc, and the medical zirconia ceramic ball as the counter part; the test was carried out in 37 °C, 25 % fetal bovine serum lubricating liquid, with a vertical load of 20 N, a sliding frequency of 1 Hz, and a total sliding distance of 1000 m; after the test, the white light interference three-dimensional profilometer was used to measure the wear track on the disc surface, and the volume wear amount of the coating was calculated; the results showed that the specific wear rate of the coating of the present application was 1 × 10 -6 mm 3 / (N m), which had excellent wear resistance.

[0032] Experimental example 5 To evaluate the killing ability of the coating of the present application to the bacteria adhered to the surface, S. aureus (ATCC 25923) was cultured on the coating of Example 1 and Comparative Example 1, respectively, in a shaking bed at 37 ℃ for 24 h, and the in vitro antibacterial performance evaluation was carried out; Comparative Example 1 was used as a negative control group, and Example 1 was used as an experimental group; after 24 h, the bacteria co-cultured in the negative control group and the experimental group were diluted to 105 CFU / mL, and plated on LB agar plates, and then cultured at 37 ℃ for 24 h, and then observed, and the results are shown in Figures 3-4 As can be seen from the number of S. aureus on the plate, there is a significant difference between the two, and the coating prepared in Comparative Example 1 has a large number of live bacteria on the surface, while the coating prepared in Example 1 can effectively kill the bacteria adhered to the surface, and has excellent contact antibacterial ability.

[0033] The present application provides a preparation method of a surface-active coating and its application. The surface-active coating prepared in the present application not only surpasses the prior art in terms of single performance such as coating bonding strength, osteogenic activity and antibacterial ability, but more importantly, through the gradient and synergistic system design, the unification and continuous high efficiency of these performances are achieved; a new solution is provided for solving the two major problems of artificial joint replacement: aseptic loosening and antibacterial ability, which has very high clinical conversion value and market prospect.

[0034] In addition, the terms "first", "second" are only for descriptive purposes, and cannot be understood as indicating or implying relative importance or implicitly indicating the number of indicated technical features. Therefore, the features defined with "first", "second" can explicitly or implicitly include one or more of the features. In the description of the present application, the meaning of "multiple" is two or more, unless otherwise specifically limited.

[0035] The above only describes the preferred embodiments of the present application and is not intended to limit the present application. Any modification, equivalent replacement and improvement made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A method for preparing a surface-active coating, characterized in that Specifically comprising the following steps: (I) base body pretreatment: ultrasonic cleaning and blow-drying of the titanium alloy to obtain a pretreated base body; (II) inner layer preparation: S1, first anodic oxidation: oxidizing the pretreated base body in an electrolyte to form a TiO2 nanotube array on the surface of the pretreated base body, thereby obtaining a first base body; S2, nanotube array stripping: ultrasonic treatment of the first base body to strip the TiO2 nanotube array layer formed thereon, thereby forming sites on the surface and obtaining a second base body; S3, second anodic oxidation: oxidizing the second base body in an electrolyte containing magnesium to form an inner layer by doping magnesium at the sites after crystallization treatment, thereby obtaining a third base body; (III) outer layer preparation: S1, preparation of precursor solution: mixing and stirring cetyltrimethylammonium bromide, zinc nitrate, strontium nitrate and auxiliary components to obtain a precursor solution; S2, coating and self-assembly: after the third base body is taken out after being immersed in the precursor solution and then aged and heated, it is immersed in an aqueous solution containing BMP-2 analog peptide and polyacrylic acid, and dried to obtain a surface active coating.

2. A method of preparing a surface active coating according to claim 1, characterised in that, The specific steps of the ultrasonic cleaning in step (I) are as follows: sequentially ultrasonic cleaning in acetone, anhydrous ethanol, 10 vol% dilute hydrochloric acid and deionized water for 15 min each.

3. The method of claim 1, wherein the surface active coating is prepared by the steps of: In step (II) S1, the electrolyte is an ethylene glycol electrolyte containing 0.5 wt% NH4F and 2 vol% deionized water; and the voltage of the oxidation reaction is 60 V.

4. The method of claim 1, wherein the surface active coating is prepared by the steps of: In step (II) S2, the specific steps of the ultrasonic treatment are as follows: immersing in a 15% H2O2 solution and ultrasonic treatment at 150 W for 10 min.

5. The method of claim 1, wherein the surface active coating is prepared by the steps of: In step (II) S3, the electrolyte is an ethylene glycol electrolyte containing 0.5 wt% NH4F and 0.1 M magnesium acetate; and the voltage of the oxidation reaction is 30-80 V.

6. The method of claim 1, wherein the surface active coating is prepared by the steps of: In step (II) S3, the specific steps of the crystallization treatment are as follows: placing the dried sample after the reaction in a muffle furnace, heating to 450 °C at a rate of 5 °C / min in air, maintaining the temperature for 2 h, and then cooling with the furnace.

7. The method of claim 1, wherein the surface active coating is prepared by the steps of: In step (III) S1, the preparation of the precursor solution is as follows: dissolving cetyltrimethylammonium bromide in a mixed solution of deionized water and anhydrous ethanol, sequentially adding tetraethyl orthosilicate, calcium nitrate tetrahydrate, zinc nitrate, strontium nitrate and triethyl phosphate, and stirring to obtain the precursor solution. The mass ratio of the zinc nitrate to the strontium nitrate is 1:1.0-2.0, and the pH of the precursor solution is 2.8-3.

2.

8. The method of claim 1, wherein the surface active coating is prepared by the steps of: In step (III) S2, the specific steps of the heating are as follows: placing the aged sample in a tube furnace, heating to 200 °C at a rate of 1 °C / min under nitrogen protection, and maintaining the temperature for 2 h.

9. The method of claim 1, wherein the surface active coating is prepared by the steps of: In step (III) S2, the mass ratio of the BMP-2 analog peptide to the polyacrylic acid is 1:

2.

10. Use of a surface active coating, characterized in that The surface active coating according to any one of claims 1-9 is applied to artificial joints, artificial implants, bone screws and spinal fusion cages.

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