Preparation method of implant with snowflake-shaped calcium phosphate bone induction coating

Abstract

The invention relates to the technical field of medical materials, and discloses a preparation method of an implant with a snowflake-shaped calcium phosphate bone induction coating, which comprises the following steps: (1) pre-treating a titanium or titanium alloy implant, the pre-treatment comprising sand blasting and acid etching, to obtain a pre-treated implant; (2) soaking the pretreated implant in a zinc oxide solution containing polyacrylamide, gradually cooling to 1-5 DEG C, soaking for 4-6 hours, then taking out, and carrying out first drying treatment; and (3) soaking the implant in a calcium phosphate solution at 1-5 DEG C for 4-6 hours, heating to 35-40 DEG C, continuously soaking for 3-4 hours, taking out, and drying for the second time. According to the invention, nano zinc oxide and polyacrylamide are deposited on the surface of the implant firstly, and then calcium phosphate is deposited, so that a snowflake-shaped bone induction coating can be formed on the surface of the implant, and the implant has higher bonding firmness, strength and biocompatibility and good bone induction characteristics.

Description

Technical Field

[0001] The invention relates to the technical field of medical materials, and in particular to a method for preparing an implant with a snowflake-shaped calcium phosphate bone-inducing coating. Background Art

[0002] Titanium and titanium alloys have become important raw materials for the manufacture of orthopedic implants due to their excellent mechanical properties, high specific strength, good corrosion resistance and biocompatibility, and have broad application prospects. However, clinical applications have found that titanium and titanium alloys are biologically inert materials, and their structure and properties are very different from bone tissue, lacking biological activity, and the bone bonding between them and human bones is a simple mechanical interlocking, lacking bone induction, and no strong chemical bonding with surrounding tissues.

[0003] This problem can be solved by modifying the surface of titanium and titanium alloys. Currently, the surface modification of titanium and titanium alloys mainly includes the following two methods: 1. Through physical or chemical surface modification methods, TiO2, dopamine and other coatings are deposited on the surface of medical titanium and titanium alloys to improve the activity of titanium alloy implants and promote the growth of bone tissue; 2. Through micro-arc oxidation, laser 3D printing and other processing techniques, micron-scale porous structures are manufactured on the surface of medical titanium and titanium alloys to promote the growth of bone tissue into the pores to enhance the bonding ability of titanium and titanium alloys with human bones. The first method improves the biological activity of titanium and titanium alloys to a certain extent, but titanium and titanium alloy implants must bear the compressive stress, shear force, torsion force and friction transmitted by natural bones during human use. However, the physical properties of the above coatings and titanium and titanium alloys are very different, which makes it difficult to significantly improve the membrane / substrate bonding strength, and the interactive cyclic stress between bones during normal human movement also affects the premature failure of the coating. The micron-scale pores formed by the second method provide a channel for bone tissue growth, but the inner wall of the pores is relatively smooth, and the induction ability of the bone tissue around the bone implant is insufficient.

[0004] The invention patent with publication number CN101791434A discloses a method for preparing a hydroxyapatite coating / surface activated titanium-based composite coating, the steps of which are: 1) using titanium or titanium alloy as the base material for surface bioactivation treatment to form a titanium gel layer, washing and drying. 2) keeping the titanium base material warm at high temperature to form a titanium oxide film. 3) synthesizing hydroxyapatite by wet method, and sintering, crushing and sieving it to obtain a powder. 4) preparing a thin layer of hydroxyapatite coating on the titanium oxide film by plasma spraying technology. 5) placing the hydroxyapatite coating product in a container, treating the product with water vapor and then drying it. This patent adopts the above method to form a titanium oxide coating on the surface of titanium and titanium alloy to improve the activity of titanium alloy implants, but due to the relatively smooth surface and the difference in physical properties between the coating and the titanium alloy, there are also problems of low film / base bonding firmness and insufficient bone induction ability. Summary of the invention

[0005] In order to solve the above technical problems, the present invention provides a method for preparing an implant with a snowflake-shaped calcium phosphate osteoinductive coating, wherein an implant with a concave-convex micro-nanoporous surface structure is obtained by pretreatment to increase the specific surface area, and then an implant with a snowflake-shaped coating is obtained by calcium-phosphate deposition, which is beneficial to the adhesion and proliferation of osteoblasts. The surface can induce the deposition of minerals, thereby increasing the mineralization area. The formed modified layer can accommodate the gradient climbing growth of bone tissue in the pores, has excellent osteoinductive properties, and has the advantages of high hardness, corrosion resistance, and friction resistance.

[0006] The purpose of the present invention is achieved through the following technical solutions: The present invention provides a method for preparing an implant having a snowflake-shaped calcium phosphate osteoinductive coating, comprising the following steps: (1) pre-treating a titanium or titanium alloy implant, wherein the pre-treatment includes sandblasting and acid etching to obtain a pre-treated implant; (2) Soaking the pretreated implant in a zinc oxide solution containing polyacrylamide and gradually cooling it to 1 to 5° C. for 4 to 6 hours, then taking it out and performing a first drying treatment; (3) The implant is then placed in a calcium phosphate solution at 1 to 5°C and soaked for 4 to 6 hours, then heated to 35 to 40°C and soaked for another 3 to 4 hours, then taken out and dried for a second time.

[0007] Since titanium or titanium alloy implants are biologically inert, they need to be surface treated to improve biocompatibility. First, the titanium or titanium alloy implants are pretreated. Sandblasting is to use sand particles to impact the surface of the implant to form irregular rough concave surfaces on the surface. Acid etching is to ensure that the surface of the implant forms a smaller rough concave surface. Pretreatment can increase the contact area between the implant and the bone tissue to enhance the bonding strength and facilitate the deposition and adhesion of the phosphate layer. Next, nano zinc oxide is deposited on the surface of the implant with an undulating micro-nanoporous surface structure. Zinc, as a bioactive element, has the effects of stimulating osteoblast osteogenesis, promoting mineralization, and stimulating osteoclast apoptosis. The dispersibility of zinc oxide in aqueous solution can be improved by adding polyacrylamide, and polyacrylamide also has good biocompatibility. Moreover, polyacrylamide can form a better bonding effect with the pretreated implant surface, thereby improving the bonding firmness of zinc oxide. During the implant immersion process, the solubility is reduced by gradually lowering the temperature, and the viscosity of polyacrylamide is increased, so that the bonding firmness is further improved. At the same time, it can locally agglomerate on the implant surface to form a snowflake-shaped protrusion base, which provides a site for the subsequent calcium phosphate deposition and can make the finally formed snowflake-shaped calcium phosphate have higher strength and biocompatibility.

[0008] By regulating the concentration, immersion temperature and time of the calcium phosphate solution, the crystallinity, thickness and morphology of calcium phosphate formed on the surface of the implant can be regulated. Calcium and phosphorus are not only essential nutrients for the human body, but also the main components of bone tissue. They can also participate in the bone reconstruction process, thereby forming a snowflake-shaped calcium phosphate coating on the surface of the implant. The snowflake is micro-nano amorphous, and each snowflake is 20 to 200 nm long, which is conducive to the adhesion and proliferation of osteoblasts. Its surface can induce the deposition of minerals and increase the mineralization area. The modified layer formed has excellent bone induction properties, and the bonding between it and the titanium matrix is ​​more firmly, and it has the advantages of high hardness, corrosion resistance and friction resistance.

[0009] Preferably, in step (1), the pretreatment comprises sandblasting, ultrasonic cleaning, acid etching and drying performed in sequence; more preferably, the pretreatment comprises sandblasting, ultrasonic cleaning, acid etching, water washing and drying performed in sequence.

[0010] Sandblasting is the process of using sand particles (usually alumina sand particles) to impact the surface of the implant to form irregular rough concave surfaces on the surface; ultrasonic cleaning is to remove the sand particles adhering to the implant surface after sandblasting; acid etching is to ensure that a smaller rough concave surface is formed on the implant surface, while further removing the sand particles that still remain after ultrasonic cleaning.

[0011] Preferably, the sandblasting is performed using aluminum oxide with a particle size of 80 to 100 meshes, the sandblasting distance is 3 to 10 mm, and the sandblasting pressure is 0.45 to 0.6 MPa.

[0012] Preferably, the acid etching is first performed by etching with a hydrofluoric acid solution and then etching with a mixed aqueous solution of hydrochloric acid and sulfuric acid.

[0013] Preferably, the mass concentration of the hydrofluoric acid solution is 1-3%; the reaction time of the hydrofluoric acid solution etching is 30-60s; in the mixed aqueous solution of hydrochloric acid and sulfuric acid, the mass concentration of hydrochloric acid is 16-19%, and the mass concentration of sulfuric acid is 32-34%; the reaction temperature of the mixed aqueous solution of hydrochloric acid and sulfuric acid etching is 75-85°C, and the time is 30-60min.

[0014] Preferably, the drying temperature is 70-85° C. and the drying time is not less than 30 minutes.

[0015] Preferably, in step (2), the preparation of the zinc oxide solution containing polyacrylamide comprises the following steps: adding nano zinc oxide with a particle size of 10 to 50 nm to a polyacrylamide aqueous solution with a mass concentration of 1 to 2%, and obtaining the zinc oxide solution containing polyacrylamide after ultrasonic dispersion.

[0016] Preferably, the mass ratio of the nano zinc oxide to polyacrylamide is 3-4:1; and the average molecular weight of the polyacrylamide is 5-10 million.

[0017] Preferably, in step (2), the first drying is performed at -20 to -10°C for 5 to 8 hours.

[0018] Preferably, in step (3), the preparation of the calcium phosphate salt solution comprises the following steps: mixing a phosphate buffer solution and ice water, and then dropping a calcium salt solution into the resulting mixed solution while stirring, and after the dropwise addition is completed, a calcium phosphate salt solution having a molar ratio of calcium to phosphorus of 1.5-1.7:1 is obtained.

[0019] Preferably, the concentration of the phosphate buffer solution is 0.05-0.2 mol / L, and the pH is 7.5-8.5; the concentration of the calcium salt solution is 0.05-0.2 mol / L; the calcium salt is calcium nitrate, calcium chloride, calcium lactate, calcium acetate or calcium gluconate; the volume ratio of the phosphate buffer solution to ice water is 1:3.5-4.5; and the temperature of the ice water is 1-5°C.

[0020] Preferably, in step (3), the second drying is: first drying at -50 to -10°C for 12 to 24 hours, and then drying at 30 to 60°C for 20 to 50 minutes.

[0021] Compared with the prior art, the present invention has the following beneficial effects: (1) Pre-treating titanium or titanium alloy implants by forming irregular rough concave surfaces on the implant surface through sandblasting and acid etching to increase the contact area between the implant and bone tissue to enhance the bonding strength; (2) Nano-zinc oxide is deposited on the surface of the implant with a concave-convex micro-nanoporous surface structure. The zinc element has the functions of stimulating osteoblasts to form bones, promoting mineralization, and stimulating osteoclast apoptosis. At the same time, the added polyacrylamide also has good biocompatibility and can improve the binding firmness of zinc oxide, providing sites for subsequent calcium phosphate deposition, thereby making the final snowflake-shaped calcium phosphate have higher strength and biocompatibility; (3) A snowflake-shaped bone-inducing coating is formed on the surface of the implant by depositing calcium phosphate. The snowflakes are micro-nano amorphous, and each snowflake is 20 to 200 nm long, which is conducive to the adhesion and proliferation of osteoblasts. Its surface can induce the deposition of minerals and increase the mineralization area. The modified layer formed has excellent bone-inducing properties and has the advantages of high hardness, corrosion resistance, and friction resistance. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is a SEM image of the implant with snowflake-like calcium phosphate osteoinductive coating in Example 1.

[0023] Figure 2 This is a SEM image of the implant with snowflake-like calcium phosphate osteoinductive coating in Example 2.

[0024] Figure 3 This is a SEM image of the implant with snowflake-like calcium phosphate osteoinductive coating in Example 3.

[0025] Figure 4 This is a SEM image of the implant with calcium phosphate osteoinductive coating in Comparative Example 1.

[0026] Figure 5 This is the SEM image of the implant with calcium phosphate osteoinductive coating in Comparative Example 2.

[0027] Figure 6 This is the SEM image of the implant with calcium phosphate osteoinductive coating in Comparative Example 3.

[0028] Figure 7 This is a SEM image of the implant with calcium phosphate osteoinductive coating in Comparative Example 4.

[0029] Figure 8 This is the SEM image of the implant with calcium phosphate osteoinductive coating in Comparative Example 5.

[0030] Fig. 9 This is a diagram showing the effect of combining the implant with the snowflake-shaped calcium phosphate osteoinductive coating in Example 1 (experimental group) with an animal experiment. DETAILED DESCRIPTION

[0031] The technical solution of the present invention is described below with specific embodiments, but the protection scope of the present invention is not limited thereto.

[0032] The preparation of an implant having a snowflake-shaped calcium phosphate osteoinductive coating comprises the following steps: (1) Pretreatment of titanium or titanium alloy implants, including sandblasting, ultrasonic cleaning, acid etching and drying in sequence, to obtain a pretreated implant with a concave-convex micro-nanopore surface structure, with a pore size of 10 to 20 μm and a pore depth of 2 to 10 μm; (2) Adding nano zinc oxide with a particle size of 10 to 50 nm to a polyacrylamide aqueous solution with a mass concentration of 1 to 2% (the average molecular weight of polyacrylamide is 5 million to 10 million), with the mass ratio of nano zinc oxide to polyacrylamide being 3 to 4:1, and ultrasonically dispersing for 20 to 40 minutes to obtain a zinc oxide solution containing polyacrylamide; Soaking the pretreated implant in the zinc oxide solution containing polyacrylamide, with an initial temperature of 25 to 30°C, and gradually cooling it to 1 to 5°C by placing it in an ice water bath, with a cooling rate of 1 to 3°C / min, and soaking it at this temperature for 4 to 6 hours; then taking it out and performing a first drying treatment, freeze-drying it at -20 to -10°C for 5 to 8 hours; (3) mixing a phosphate buffer solution with a concentration of 0.05 to 0.2 mol / L and a pH of 7.5 to 8.5 with ice water at 1 to 5°C, wherein the volume ratio of the phosphate buffer solution to the ice water is 1:3.5 to 4.5, and then dripping a calcium salt solution with a concentration of 0.05 to 0.2 mol / L into the resulting mixed solution while stirring, and controlling the dripping time to be 20 to 40 min. After the dripping is completed, a calcium phosphate solution with a molar ratio of calcium to phosphorus of 1.5 to 1.7:1 is obtained; The calcium phosphate solution is cooled to 1-5°C, and then the implant in step (2) is placed in the calcium phosphate solution and soaked for 4-6 hours, and the solution is heated to 35-40°C and soaked for 3-4 hours; then the implant is taken out and subjected to a second drying treatment, first freeze-dried at -50--10°C for 12-24 hours, and then dried at 30-60°C for 20-50 minutes, to obtain an implant with a snowflake-shaped calcium phosphate osteoinductive coating, wherein the snowflake is micro-nano amorphous, and each snowflake is 20-200 nm in length.

[0033] In a specific embodiment of the present invention, the pretreatment includes cleaning, sandblasting, rinsing, ultrasonic cleaning, drying, acid etching, water washing and drying performed in sequence.

[0034] In a specific embodiment of the present invention, the sand blasting is performed by using alumina with a particle size of 80-100 meshes through a sand blasting machine, the sand blasting distance is 3-10 mm, and the sand blasting pressure is 0.45-0.6 MPa.

[0035] In a specific embodiment of the present invention, the acid etching is as follows: firstly, the implant is placed in a polytetrafluoroethylene beaker, and is etched with a hydrofluoric acid solution having a mass concentration of 1 to 3% for 30 to 60 seconds, and then the implant is taken out and rinsed with purified water; then, the implant is placed in a glass beaker, and is etched with a mixed aqueous solution of hydrochloric acid and sulfuric acid (containing a mass concentration of 16 to 19% hydrochloric acid and a mass concentration of 32 to 34% sulfuric acid), heated in a water bath to 75 to 85°C, and reacted for 30 to 60 minutes, with stirring required during the process.

[0036] In a specific embodiment of the present invention, ultrasonic cleaning is as follows: first, a cleaning agent (MICRO-90) and purified water are respectively placed in tank 1 at a volume ratio of 1.2:100, at a temperature of 60-70°C for 20-30 min, and the implant is ultrasonically cleaned; then, purified water is placed in tank 2 at a temperature of 60-70°C for 20-30 min, and the implant is ultrasonically cleaned; then, purified water is placed in tank 3 at a temperature of 60-70°C for 10-20 min, and the implant is bubble cleaned.

[0037] In a specific embodiment of the present invention, the water washing is: washing with purified water for 15 to 20 minutes, and then rinsing with purified water for 15 to 45 seconds.

[0038] In a specific embodiment of the present invention, the drying temperature is 70-85° C. and the drying time is not less than 30 minutes.

[0039] In a specific embodiment of the present invention, in step (3), the calcium salt is calcium nitrate, calcium chloride, calcium lactate, calcium acetate or calcium gluconate.

[0040] Example 1 (1) pre-treating the titanium or titanium alloy implant, the pre-treating comprising sequentially cleaning, sandblasting, rinsing, ultrasonic cleaning, drying, acid etching, water washing and drying to obtain a pre-treated implant; (2) Adding nano zinc oxide with an average particle size of 30 nm to a polyacrylamide aqueous solution with a mass concentration of 1% (the average molecular weight of polyacrylamide is 8 million), with the mass ratio of nano zinc oxide to polyacrylamide being 3:1, and ultrasonically dispersing for 30 minutes to obtain a zinc oxide solution containing polyacrylamide; soaking the pretreated implant in the zinc oxide solution containing polyacrylamide, with the initial temperature being room temperature 28°C, and gradually cooling to 4°C by placing it in an ice water bath, with an average cooling rate of 1°C / min, and soaking at this temperature for 5 hours; then taking it out and performing the first drying treatment, freeze-drying it at -15°C for 6 hours; (3) Mixing a phosphate buffer solution with a concentration of 0.1 mol / L and a pH of 7.5-8.5 with 4°C ice water, wherein the volume ratio of the phosphate buffer solution to the ice water is 1:4, and then dripping a calcium salt solution with a concentration of 0.1 mol / L into the resulting mixed solution while stirring. The dripping time is controlled to be 30 min. After the dripping is completed, a calcium phosphate solution with a molar ratio of calcium to phosphorus of 1.6:1 is obtained; cooling the calcium phosphate solution to 4°C, and then placing the implant in step (2) in the calcium phosphate solution and soaking it for 4 h, heating the solution to 35°C and continuing to soak it for 4 h; then taking it out and performing a second drying treatment, first freeze-drying it at -50°C for 12 h, and then drying it at 40°C for 30 min, to obtain an implant with a snowflake-like calcium phosphate osteoinductive coating.

[0041] Example 2 (1) pre-treating the titanium or titanium alloy implant, the pre-treating comprising sequentially cleaning, sandblasting, rinsing, ultrasonic cleaning, drying, acid etching, water washing and drying to obtain a pre-treated implant; (2) Adding nano zinc oxide with an average particle size of 30 nm to a polyacrylamide aqueous solution with a mass concentration of 1% (the average molecular weight of polyacrylamide is 8 million), with the mass ratio of nano zinc oxide to polyacrylamide being 4:1, and ultrasonically dispersing for 30 minutes to obtain a zinc oxide solution containing polyacrylamide; placing the pretreated implant in the zinc oxide solution containing polyacrylamide, with the initial temperature being room temperature 25°C, and gradually cooling it to 4°C by placing it in an ice water bath, with an average cooling rate of 1°C / min, and soaking it at this temperature for 6 hours; then taking it out and performing the first drying treatment, freeze-drying it at -20°C for 5 hours; (3) Mixing a phosphate buffer solution with a concentration of 0.1 mol / L and a pH of 7.5-8.5 with 4°C ice water, wherein the volume ratio of the phosphate buffer solution to the ice water is 1:4, and then dripping a calcium salt solution with a concentration of 0.1 mol / L into the resulting mixed solution while stirring. The dripping time is controlled to be 30 min. After the dripping is completed, a calcium phosphate solution with a molar ratio of calcium to phosphorus of 1.6:1 is obtained; cooling the calcium phosphate solution to 4°C, and then placing the implant in step (2) in the calcium phosphate solution and soaking it for 4 h, heating the solution to 35°C and continuing to soak it for 4 h; then taking it out and performing a second drying treatment, first freeze-drying it at -50°C for 12 h, and then drying it at 40°C for 30 min, to obtain an implant with a snowflake-like calcium phosphate osteoinductive coating.

[0042] Example 3 (1) pre-treating the titanium or titanium alloy implant, the pre-treating comprising sequentially cleaning, sandblasting, rinsing, ultrasonic cleaning, drying, acid etching, water washing and drying to obtain a pre-treated implant; (2) Adding nano zinc oxide with an average particle size of 20 nm to a 2% polyacrylamide aqueous solution (the average molecular weight of polyacrylamide is 10 million), with the mass ratio of nano zinc oxide to polyacrylamide being 3:1, and ultrasonically dispersing for 40 minutes to obtain a zinc oxide solution containing polyacrylamide; placing the pretreated implant in the zinc oxide solution containing polyacrylamide, with the initial temperature being room temperature 30°C, and gradually cooling it to 4°C by placing it in an ice water bath, with an average cooling rate of 1°C / min, and soaking it at this temperature for 4 hours; then taking it out and performing the first drying treatment, freeze-drying it at -20°C for 6 hours; (3) Mixing a phosphate buffer solution with a concentration of 0.2 mol / L and a pH of 7.5-8.5 with 4°C ice water, wherein the volume ratio of the phosphate buffer solution to the ice water is 1:4, and then dripping a calcium salt solution with a concentration of 0.2 mol / L into the resulting mixed solution while stirring. The dripping time is controlled to be 40 min. After the dripping is completed, a calcium phosphate solution with a molar ratio of calcium element to phosphorus element of 1.5:1 is obtained; cooling the calcium phosphate solution to 4°C, and then placing the implant in step (2) in the calcium phosphate solution and soaking it for 3 h, heating the solution to 40°C and continuing to soak it for 3 h; then taking it out and performing a second drying treatment, first freeze-drying it at -30°C for 16 h, and then drying it at 50°C for 30 min, to obtain an implant with a snowflake-like calcium phosphate osteoinductive coating.

[0043] Comparative Example 1 The difference from Example 1 is that zinc oxide and polyacrylamide are not added.

[0044] (1) pre-treating the titanium or titanium alloy implant, the pre-treating comprising sequentially cleaning, sandblasting, rinsing, ultrasonic cleaning, drying, acid etching, water washing and drying to obtain a pre-treated implant; (2) A phosphate buffer solution with a concentration of 0.1 mol / L and a pH of 7.5-8.5 and 4°C ice water are mixed, and the volume ratio of the phosphate buffer solution to the ice water is 1:4. A calcium salt solution with a concentration of 0.1 mol / L is then dripped into the resulting mixture while stirring. The dripping time is controlled to be 30 min. After the dripping is completed, a calcium phosphate solution with a molar ratio of calcium to phosphorus of 1.6:1 is obtained. The calcium phosphate solution is cooled to 4°C, and the pretreated implant is immersed in the calcium phosphate solution for 4 h. The solution is heated to 35°C and immersed for another 4 h. The implant is then taken out and dried for a second time, first freeze-dried at -50°C for 12 h, and then dried at 40°C for 30 min, to obtain an implant with a calcium phosphate osteoinductive coating.

[0045] Comparative Example 2 The difference from Example 1 is that no polyacrylamide is added.

[0046] (1) pre-treating the titanium or titanium alloy implant, the pre-treating comprising sequentially cleaning, sandblasting, rinsing, ultrasonic cleaning, drying, acid etching, water washing and drying to obtain a pre-treated implant; (2) Adding nano zinc oxide with an average particle size of 30 nm to water in a mass ratio of 3:100, and ultrasonically dispersing for 30 minutes to obtain a zinc oxide solution; soaking the pretreated implant in the zinc oxide solution at an initial temperature of 28°C, and gradually cooling it to 4°C in an ice water bath at an average cooling rate of 1°C / min, and soaking it at this temperature for 5 hours; then taking it out and performing the first drying treatment, freeze-drying it at -15°C for 6 hours; (3) Mixing a phosphate buffer solution with a concentration of 0.1 mol / L and a pH of 7.5-8.5 with 4°C ice water, wherein the volume ratio of the phosphate buffer solution to the ice water is 1:4, and then dripping a calcium salt solution with a concentration of 0.1 mol / L into the resulting mixed solution while stirring. The dripping time is controlled to be 30 min. After the dripping is completed, a calcium phosphate solution with a molar ratio of calcium to phosphorus of 1.6:1 is obtained; cooling the calcium phosphate solution to 4°C, and then placing the implant in step (2) in the calcium phosphate solution and soaking it for 4 h, heating the solution to 35°C and continuing to soak it for 4 h; then taking it out and performing a second drying treatment, first freeze-drying it at -50°C for 12 h, and then drying it at 40°C for 30 min, to obtain an implant with a calcium phosphate bone induction coating.

[0047] Comparative Example 3 The difference from Example 1 is that the immersion temperature of the implant in the zinc oxide solution containing polyacrylamide is changed.

[0048] (1) pre-treating the titanium or titanium alloy implant, the pre-treating comprising sequentially cleaning, sandblasting, rinsing, ultrasonic cleaning, drying, acid etching, water washing and drying to obtain a pre-treated implant; (2) Adding nano zinc oxide with an average particle size of 30 nm to a polyacrylamide aqueous solution with a mass concentration of 1% (the average molecular weight of polyacrylamide is 8 million), with the mass ratio of nano zinc oxide to polyacrylamide being 3:1, and ultrasonically dispersing for 30 minutes to obtain a zinc oxide solution containing polyacrylamide; soaking the pretreated implant in the zinc oxide solution containing polyacrylamide (at room temperature 28°C) for 5 hours; then taking it out and performing the first drying treatment, freeze-drying it at -15°C for 6 hours; (3) Mixing a phosphate buffer solution with a concentration of 0.1 mol / L and a pH of 7.5-8.5 with 4°C ice water, wherein the volume ratio of the phosphate buffer solution to the ice water is 1:4, and then dripping a calcium salt solution with a concentration of 0.1 mol / L into the resulting mixed solution while stirring. The dripping time is controlled to be 30 min. After the dripping is completed, a calcium phosphate solution with a molar ratio of calcium to phosphorus of 1.6:1 is obtained; cooling the calcium phosphate solution to 4°C, and then placing the implant in step (2) in the calcium phosphate solution and soaking it for 4 h, heating the solution to 35°C and continuing to soak it for 4 h; then taking it out and performing a second drying treatment, first freeze-drying it at -50°C for 12 h, and then drying it at 40°C for 30 min, to obtain an implant with a calcium phosphate bone induction coating.

[0049] Comparative Example 4 The difference from Example 1 is that the implant is not freeze-dried but directly immersed in the calcium phosphate solution.

[0050] (1) pre-treating the titanium or titanium alloy implant, the pre-treating comprising sequentially cleaning, sandblasting, rinsing, ultrasonic cleaning, drying, acid etching, water washing and drying to obtain a pre-treated implant; (2) Adding nano zinc oxide with an average particle size of 30 nm to a polyacrylamide aqueous solution with a mass concentration of 1% (the average molecular weight of polyacrylamide is 8 million), with the mass ratio of nano zinc oxide to polyacrylamide being 3:1, and ultrasonically dispersing for 30 minutes to obtain a zinc oxide solution containing polyacrylamide; soaking the pretreated implant in the zinc oxide solution containing polyacrylamide, with the initial temperature being room temperature 28°C, and gradually cooling it to 4°C by placing it in an ice water bath, with an average cooling rate of 1°C / min, and soaking it at this temperature for 5 hours; then taking it out and performing air drying; (3) Mixing a phosphate buffer solution with a concentration of 0.1 mol / L and a pH of 7.5-8.5 with 4°C ice water, wherein the volume ratio of the phosphate buffer solution to the ice water is 1:4, and then dripping a calcium salt solution with a concentration of 0.1 mol / L into the resulting mixed solution while stirring. The dripping time is controlled to be 30 min. After the dripping is completed, a calcium phosphate solution with a molar ratio of calcium to phosphorus of 1.6:1 is obtained; cooling the calcium phosphate solution to 4°C, and then placing the implant in step (2) in the calcium phosphate solution and soaking it for 4 h, heating the solution to 35°C and continuing to soak it for 4 h; then taking it out and performing a second drying treatment, first freeze-drying it at -50°C for 12 h, and then drying it at 40°C for 30 min, to obtain an implant with a calcium phosphate bone induction coating.

[0051] Comparative Example 5 The difference from Example 1 is that the concentration of the calcium phosphate solution is too high.

[0052] (1) pre-treating the titanium or titanium alloy implant, the pre-treating comprising sequentially cleaning, sandblasting, rinsing, ultrasonic cleaning, drying, acid etching, water washing and drying to obtain a pre-treated implant; (2) Adding nano zinc oxide with an average particle size of 30 nm to a polyacrylamide aqueous solution with a mass concentration of 1% (the average molecular weight of polyacrylamide is 8 million), with the mass ratio of nano zinc oxide to polyacrylamide being 3:1, and ultrasonically dispersing for 30 minutes to obtain a zinc oxide solution containing polyacrylamide; soaking the pretreated implant in the zinc oxide solution containing polyacrylamide, with the initial temperature being room temperature 28°C, and gradually cooling to 4°C by placing it in an ice water bath, with an average cooling rate of 1°C / min, and soaking at this temperature for 5 hours; then taking it out and performing the first drying treatment, freeze-drying it at -15°C for 6 hours; (3) Mixing a phosphate buffer solution with a concentration of 0.4 mol / L and a pH of 7.5-8.5 with 4°C ice water, wherein the volume ratio of the phosphate buffer solution to the ice water is 1:4, and then dripping a calcium salt solution with a concentration of 0.4 mol / L into the resulting mixed solution while stirring. The dripping time is controlled to be 30 min. After the dripping is completed, a calcium phosphate solution with a molar ratio of calcium to phosphorus of 1.6:1 is obtained; cooling the calcium phosphate solution to 4°C, and then placing the implant in step (2) in the calcium phosphate solution and soaking it for 4 h, heating the solution to 35°C and continuing to soak it for 4 h; then taking it out and performing a second drying treatment, first freeze-drying it at -50°C for 12 h, and then drying it at 40°C for 30 min, to obtain an implant with a calcium phosphate osteoinductive coating.

[0053] Performance Testing: (1) The surface morphology of the embodiments of the present invention and the comparative examples was observed by scanning electron microscopy (SEM), and the test method was in accordance with GB / T16594-2008 General Rules for Scanning Electron Microscope Measurement of Micrometer-Level Length. The tensile (adhesive) strength of the coating complies with ASTM F1609-03, and the test method was in accordance with ASTM F1147 Standard Test Method for Tensile Test of Calcium Phosphate and Metal Coatings.

[0054] Table 1 Examples Coating tensile (bonding) strength Example 1 50.3MPa Example 2 49.5MPa Example 3 48.7MPa Comparative Example 1 38.1MPa Comparative Example 2 37.3MPa Comparative Example 3 35.8MPa Comparative Example 4 36.4MPa Comparative Example 5 37.6MPa As shown in Table 1, the present invention deposits nano zinc oxide on the surface of the pretreated titanium alloy to provide sites for subsequent calcium phosphate deposition. The added polyacrylamide can improve the bonding strength between the coating and the titanium substrate. Then, a snowflake-shaped bone induction coating is formed on the surface of the implant by depositing calcium phosphate. The snowflake is micro-nano amorphous, and each snowflake is 20 to 200 nm long, which is conducive to the adhesion and proliferation of osteoblasts. The surface can induce the deposition of minerals, increase the mineralization area, and the formed modified layer has a higher bonding strength with the titanium substrate, so the tensile strength of the implant is higher.

[0055] like Figure 1 , Figure 2 and Figure 3 As shown, the implants prepared in Examples 1-3 all have snowflake-like calcium phosphate osteoinductive coatings, and snowflake-like crystals with a length of 20 to 200 nm are randomly distributed and integrated with the surface of the titanium matrix, thereby increasing the osteoinductive specific surface area while having higher strength, being beneficial to the adhesion and proliferation of osteoblasts, being able to accommodate the gradient climbing growth of bone tissue in the pores, and having excellent osteoinductive properties.

[0056] In Comparative Example 1, since zinc oxide and polyacrylamide were not added, the bonding strength between the calcium phosphate coating and the titanium substrate was poor. Figure 4 As shown in FIG. 1 , the surface calcium phosphate crystals are relatively loose, and the physical properties of the coating and titanium and titanium alloy are greatly different, so the tensile strength of the implant is also poor. Similarly, in Comparative Example 2, since polyacrylamide is not added, Figure 5 As shown, it will also lead to the inability to form a snowflake-like morphology and poor tensile strength.

[0057] In Comparative Example 3, since the immersion temperature of the implant in the zinc oxide solution containing polyacrylamide was changed, it was impossible to locally aggregate on the surface of the implant to form a snowflake-like protrusion base, the bonding firmness was also affected, and the tensile strength was reduced. Figure 6 As shown, although the implant surface in Comparative Example 3 also has an uneven rough surface, it cannot form a snowflake-like morphology.

[0058] In Comparative Example 4, the implant was not freeze-dried and directly immersed in a calcium phosphate solution. Figure 7 As shown, although a partial snowflake-like morphology can be formed, the morphology will be affected, and the bonding strength between the coating and the titanium substrate will also be affected, resulting in a decrease in tensile strength.

[0059] The concentration of calcium phosphate solution in Comparative Example 5 is too high. Figure 8 As shown, it will affect the crystallinity, thickness and morphology of calcium phosphate formed on the surface of the implant, thereby affecting the tensile strength of the implant.

[0060] (2) The zoological experiments are as follows: Two days before the experiment, CT scans were taken of the lower jaw teeth on both sides of 10 Beagle dogs.

[0061] Preoperative preparation: Animals were fasted from food and water for 12 hours before surgery.

[0062] Animal anesthesia: After the animals were weighed, they were anesthetized with a combination of Sumixin and Shutai (2:1), with an anesthetic dose of 0.2 mL / kg Sumixin and 0.1 mL / kg Shutai. During the operation, the anesthesia state was maintained with a combination of Sumixin and Shutai (2:1) according to the animal's response (respiration, muscle tension, blink reflex, etc.).

[0063] Canine tooth extraction: After general anesthesia, the fourth mandibular premolars on both sides were extracted (the distance between the mesial root and the distal root was greater than 8 mm).

[0064] Implant placement: Immediate implantation is used, and the implantation socket is prepared immediately after tooth extraction. The drill bit drills and grinds the implantation socket on the tooth bone at a drilling speed of about 1200r / min and a torque of 35N·cm. During the preparation process, the implantation machine automatically sprays ice saline for cooling. The implant is screwed in at a low speed with a torque of 35N.cm, and then the temporary covering nut is screwed on. Then the mucoperiosteal flap is reset and tightly sutured.

[0065] Postoperative care: The animals were kept in separate cages after surgery, allowed to move freely, and fasted for 12 hours. 1.6 million IU of penicillin was injected intramuscularly for 3 consecutive days after surgery to prevent infection. The oral cavity was cleaned for two weeks to prevent infection. Liquid food was used for 2 weeks and soft food was used for 1 week.

[0066] Termination of the experiment: 6 months after the operation, the experiment was terminated, blood was collected from the animals, and the animals were killed by overdose of anesthesia. Various examinations were performed, gingival tissue from the implant site was collected for histopathological examination, and samples from the implant site were collected for micro-CT and hard tissue sections.

[0067] Table 2 like Figure 8 As shown and illustrated in Table 2, the implant with the snowflake-like calcium phosphate osteoinductive coating was subjected to an animal experiment in beagle dogs, and the results showed that the average bone binding rate of Example 1 was greater than the average bone binding rate of Comparative Example 1, indicating that the solution of the present invention has good cell compatibility and biological activity, and the results show better binding.

[0068] The above are only preferred embodiments of the present invention, and are not intended to limit the patent scope of the present invention. Any equivalent structure or equivalent process transformation made using the specification of the present invention, or directly or indirectly applied in other related technical fields, are also included in the patent protection scope of the present invention.

Claims

1. A method for preparing an implant having a snowflake-like calcium phosphate osteoinductive coating, characterized in that: The steps include: (1) Pre-treating the titanium or titanium alloy implant, wherein the pre-treatment includes sandblasting and acid etching to obtain a pre-treated implant; (2) Soak the pretreated implant in a zinc oxide solution containing polyacrylamide and gradually cool it to 1-5°C for 4-6 hours, then take it out and perform the first drying treatment; (3) Then soak the implant in a 1-5°C calcium phosphate solution for 4-6 hours, raise the temperature to 35-40°C and continue soaking for 3-4 hours, then take it out and dry it for the second time.

2. The method for preparing an implant having a snowflake-like calcium phosphate osteoinductive coating according to claim 1, characterized in that: In step (1), the pretreatment includes sandblasting, ultrasonic cleaning, acid etching and drying performed in sequence.

3. The method for preparing an implant having a snowflake-like calcium phosphate osteoinductive coating according to claim 1 or 2, characterized in that: The sandblasting is performed using aluminum oxide with a particle size of 80-100 meshes, the sandblasting distance is 3-10 mm, and the sandblasting pressure is 0.45-0.6 MPa.

4. The method for preparing an implant having a snowflake-like calcium phosphate osteoinductive coating according to claim 1 or 2, characterized in that: The acid etching is firstly carried out by etching with a hydrofluoric acid solution, and then etching with a mixed aqueous solution of hydrochloric acid and sulfuric acid.

5. The method for preparing an implant having a snowflake-like calcium phosphate osteoinductive coating according to claim 1, characterized in that: In step (2), the preparation of the zinc oxide solution containing polyacrylamide comprises the following steps: adding nano zinc oxide with a particle size of 10-50 nm to a polyacrylamide aqueous solution with a mass concentration of 1-2%, and performing ultrasonic dispersion to obtain the zinc oxide solution containing polyacrylamide.

6. The method for preparing an implant having a snowflake-like calcium phosphate osteoinductive coating according to claim 5, characterized in that: The mass ratio of the nano zinc oxide to the polyacrylamide is 3-4:1; the average molecular weight of the polyacrylamide is 5-10 million.

7. The method for preparing an implant having a snowflake-like calcium phosphate osteoinductive coating according to claim 1, 5 or 6, characterized in that: In step (2), the first drying is performed at -20 to -10°C for 5 to 8 hours.

8. The method for preparing an implant having a snowflake-like calcium phosphate osteoinductive coating according to claim 1, characterized in that: In step (3), the preparation of the calcium phosphate salt solution comprises the following steps: mixing a phosphate buffer solution and ice water, and then dripping a calcium salt solution into the resulting mixed solution while stirring. After the dripping is completed, a calcium phosphate salt solution having a molar ratio of calcium to phosphorus of 1.5-1.7:1 is obtained.

9. The method for preparing an implant having a snowflake-like calcium phosphate osteoinductive coating according to claim 8, characterized in that: The concentration of the phosphate buffer solution is 0.05-0.2 mol / L, and the pH is 7.5-8.5; the concentration of the calcium salt solution is 0.05-0.2 mol / L; and the volume ratio of the phosphate buffer solution to ice water is 1:3.5-4.

5.

10. The method for preparing an implant having a snowflake-like calcium phosphate osteoinductive coating according to claim 1, 8 or 9, characterized in that: In step (3), the second drying is: first drying at -50~-10°C for 12~24h, and then drying at 30~60°C for 20~50min.

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