Piezoelectric nano-hydroxyapatite modified titanium-based implant as well as preparation method and application thereof

Abstract

The invention provides a piezoelectric nano hydroxyapatite modified titanium-based implant as well as a preparation method and application thereof, and particularly relates to the technical field of biomedical materials. The preparation method of the piezoelectric nano-hydroxyapatite modified titanium-based implant comprises the following steps: dripping a piezoelectric nano-hydroxyapatite solution on a titanium plate with a titanium dioxide nanotube array, then putting the titanium plate into a vacuum drying oven for vacuum adsorption, and repeatedly dripping after the surface of the titanium plate is dried, thereby obtaining the piezoelectric nano-hydroxyapatite modified titanium-based implant. And washing the titanium plate by using a buffer solution, and finally carrying out vacuum drying to obtain the piezoelectric nano-hydroxyapatite modified titanium-based implant. According to the preparation method, through a physical-chemical synergistic effect mechanism, collaborative optimization of surface biological activity, mechanical properties and functional characteristics of the implant is realized, and the preparation method has remarkable clinical application value.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedical materials, and particularly to a piezoelectric nano-hydroxyapatite modified titanium-based implant and its preparation method and application. Background Art

[0002] Orthopedic implants are key medical devices for repairing bone injuries. However, the problem of their long-term failure is still prominent, and postoperative infection and prosthesis loosening are the main causes of implant failure. Currently, the titanium-based implants used clinically lack an active anti-infection design, and the existing surface modification technologies have significant limitations. The passive antibacterial strategy inhibits bacterial adhesion by regulating surface roughness, hydrophilicity / hydrophobicity or charge distribution, but this method will also inhibit the adhesion and proliferation of osteoblasts, resulting in poor bone bonding and increasing the risk of loosening; the active bactericidal strategy relies on the loading and slow release of antibiotics, metal ions or antimicrobial peptides, but the release behavior of the antibacterial components is uncontrollable, making it difficult to achieve long-term on-demand antibacterial effects throughout the life cycle of the implant, and long-term release may cause drug resistance or biological toxicity.

[0003] Sonodynamic therapy (SDT), as a new non-invasive antibacterial technology, generates reactive oxygen species to kill bacteria by activating sonosensitizers with ultrasound, and is particularly suitable for the treatment of deep tissue infections. However, traditional sonosensitizers such as porphyrin compounds have defects such as poor water solubility, low targeting and insufficient biosafety, which limit their application in orthopedic implants.

[0004] Based on the above background, the prior art urgently needs to solve the following core problems: first, how to construct a titanium-based implant surface with both long-term anti-infection and bone integration functions to avoid the mutual antagonism between antibacterial and osteogenic functions; second, how to achieve the on-demand release of antibacterial components, which are only triggered when an infection occurs, so as to extend the service life of the implant and reduce the risk of drug resistance; third, how to develop an inorganic sonosensitizer with high biosafety and high piezoelectric response, which can efficiently antibacterial under ultrasonic stimulation and directly participate in the bone repair process.

[0005] In view of this, the present invention is specifically proposed. Summary of the Invention

[0006] One object of the present invention is to provide a preparation method of a piezoelectric nano-hydroxyapatite modified titanium-based implant, aiming to solve at least one of the above technical problems in the prior art.

[0007] Another object of the present invention is to provide a piezoelectric nano-hydroxyapatite modified titanium-based implant.

[0008] A further object of the present invention is to provide an application of a piezoelectric nano-hydroxyapatite modified titanium-based implant.

[0009] To achieve the above object of the present invention, the following technical solutions are specifically adopted:

[0010] In a first aspect of the present invention, a preparation method of a piezoelectric nano-hydroxyapatite modified titanium-based implant is provided. A piezoelectric nano-hydroxyapatite solution is dropped onto a titanium plate with a titanium dioxide nanotube array, and then the titanium plate is placed in a vacuum drying oven for vacuum adsorption. After the surface of the titanium plate is dried, the dropping is repeated. Then, the titanium plate is rinsed with a buffer solution, and finally vacuum dried to obtain the piezoelectric nano-hydroxyapatite modified titanium-based implant.

[0011] Further, the concentration of the piezoelectric nano-hydroxyapatite solution is 5 - 15 mg / mL.

[0012] Preferably, the solvent in the piezoelectric nano-hydroxyapatite solution includes PBS solution, physiological saline solution, simulated body fluid, deionized water, DPBS solution, HBSS solution or EBSS solution.

[0013] Preferably, the buffer solution is the solvent in the piezoelectric nano-hydroxyapatite solution.

[0014] Preferably, the ratio of the usage amount of the piezoelectric nano-hydroxyapatite solution to the area of the titanium plate is 10 - 50 μL / cm 2 .

[0015] Further, the number of times of repeated dropping is 2 - 8 times.

[0016] Preferably, the number of times of rinsing is 1 - 5 times.

[0017] Further, the temperature of the vacuum drying is 50 - 150 °C, and the time is 30 - 90 minh.

[0018] Further, the preparation method of the titanium plate with a titanium dioxide nanotube array is as follows: An anodic oxidation method is used to prepare a titanium dioxide nanotube array on the surface of the titanium plate, and finally annealed to obtain the titanium plate with a titanium dioxide nanotube array.

[0019] Preferably, in the titanium dioxide nanotube array, the length of the titanium dioxide nanotubes is 3 - 8 μm, and the diameter is 50 - 200 nm.

[0020] Preferably, the heating rate of the annealing is 1 - 3 °C / min.

[0021] Preferably, the annealing is carried out in air, the annealing temperature is 400 - 500 °C, and the time is 1 - 3 h.

[0022] Further, the preparation method of the piezoelectric nano-hydroxyapatite is as follows:

[0023] Slowly drop the phosphorus source solution into the calcium source solution, and after completion, mix evenly to obtain a mixed solution; hydrothermally react the mixed solution at 160-200 °C for 18-24 h, and after the reaction, separate the product from the solid and liquid, wash and vacuum dry to obtain the piezoelectric nano-hydroxyapatite.

[0024] Furthermore, the dropping rate of the phosphorus source solution is 0.5-1.5 mL / min.

[0025] Preferably, the temperature of the vacuum drying is 50-80 °C, the time is 12-24 h, and the vacuum degree ≤ 0.1 MPa.

[0026] Preferably, the preparation method of the piezoelectric nano-hydroxyapatite further includes an ultrasonic dispersion process after solid-liquid separation and before washing.

[0027] Preferably, the power of the ultrasonic dispersion is 100-200 W, and the time is 5-15 min.

[0028] Furthermore, the calcium source in the calcium source solution is calcium chloride.

[0029] Preferably, the phosphorus source in the phosphorus source solution is ammonium dihydrogen phosphate.

[0030] Preferably, the molar ratio of calcium in the calcium source to phosphorus in the phosphorus source is 1:

[0031] (0.6-0.7).

[0032] Preferably, the phosphorus source solution and / or the solvent in the phosphorus source solution is an ammonia water solution.

[0033] The second aspect of the present invention provides a piezoelectric nano-hydroxyapatite modified titanium-based implant, which is prepared by using the preparation method described in the first aspect.

[0034] The third aspect of the present invention provides the application of the piezoelectric nano-hydroxyapatite modified titanium-based implant in the preparation of joint prostheses, bone filling materials or fracture internal fixation materials.

[0035] Compared with the prior art, the present invention has at least the following beneficial effects:

[0036] The preparation method provided by the present invention first uses a vacuum adsorption combined with a repeated dropping strategy to effectively fill the three-dimensional pore structure of the titanium dioxide nanotube array with piezoelectric nano-hydroxyapatite. The vacuum negative pressure drives the deep penetration of the solution, realizing the physical anchoring and chemical bonding of nanoparticles on the surface of the titanium substrate, and forming a mechanical interlocking structure. Secondly, the cyclic process of multiple dropping and vacuum adsorption constructs a gradient structure that transitions from a dense bottom layer to a porous surface layer at the nanoscale. This bionic structure not only retains the topological cell growth promoting effect of the nanotube array, but also forms a dual bioactive interface through the charge stimulation of piezoelectric nano-hydroxyapatite. The buffer solution is used to wash away the weakly bound particles, expose the active crystal plane, and optimize the calcium and phosphate ion release rate. This preparation method realizes the coordinated optimization of the bioactivity, mechanical properties, and functional characteristics of the implant surface through a physical-chemical synergistic mechanism, and has significant clinical application value.

[0037] The piezoelectric nano-hydroxyapatite modified titanium-based implant provided by the present invention realizes the antibacterial-osteogenic synergistic functional design by constructing a titanium dioxide nanotube array on the surface of the titanium plate and loading nano-hydroxyapatite with enhanced piezoelectric properties as a sonosensitizer. This technology utilizes the deep tissue penetration advantage of sonodynamic therapy (SDT) to activate the piezoelectric nano-hydroxyapatite sonosensitizer under ultrasonic stimulation to generate reactive oxygen species (ROS) and efficiently kill bacteria. At the same time, nano-hydroxyapatite, as a natural component of bone tissue, directly participates in the bone repair process by promoting osteoblast adhesion and mineralization, significantly enhancing the bone integration ability of the implant. In addition, the porous high specific surface area structure of the TiO2 nanotube array ensures the long-term stable loading and controlled release of the sonosensitizer, and its piezoelectric properties can be precisely regulated by ultrasound to achieve infection-responsive on-demand treatment, avoiding drug resistance or tissue damage caused by the release of unnecessary antibacterial agents. This technology breaks through the limitations of the existing implant passive antibacterial and active bactericidal strategies, and while ensuring the long-term antibacterial efficacy during the service life, reduces the loosening risk through bionic bone component design, providing an innovative path for the biofunctional surface modification of orthopedic implants.

[0038] Regarding the advantages of the piezoelectric nano-hydroxyapatite modified titanium-based implant provided by the present invention, the application of the piezoelectric nano-hydroxyapatite modified titanium-based implant improves the biofunctionality of downstream products such as joint prostheses, bone filling materials, or fracture internal fixation materials, and promotes the development of downstream industries. Description of the Drawings

[0039] In order to more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the following will briefly introduce the drawings required for the description of the specific embodiments or the prior art. Obviously, the drawings in the following description are some embodiments of the present invention. For those of ordinary skill in the art, other drawings can be obtained based on these drawings without creative efforts.

[0040] Figure 1 Schematic diagram of the structure of the piezoelectric nano-hydroxyapatite modified titanium-based implant provided by the present invention;

[0041] Figure 2 TEM image of piezoelectric nano-hydroxyapatite;

[0042] Figure 3 XPS image of piezoelectric nano-hydroxyapatite;

[0043] Figure 4 X-ray diffraction pattern of piezoelectric nano-hydroxyapatite;

[0044] Figure 5 Graph for measuring the current response of piezoelectric nano-hydroxyapatite;

[0045] Figure 6 Graph of antibacterial effect of plate coating;

[0046] Figure 7 Histogram of bacterial count of plate coating;

[0047] Figure 8 Biocompatibility (graph of CCK8 test results);

[0048] Figure 9 Graph of histological HE staining. Detailed implementation manners

[0049] To make the objectives, technical solutions and advantages of the present invention clearer, the technical solutions in the embodiments 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, rather than all of the embodiments.

[0050] In the following text, the terms "including", "having" and their cognates that can be used in various embodiments of the present invention are only intended to represent specific features, numbers, steps, operations, elements, components or combinations of the foregoing items, and should not be construed as first excluding the existence of one or more other features, numbers, steps, operations, elements, components or combinations of the foregoing items or increasing the possibility of one or more features, numbers, steps, operations, elements, components or combinations of the foregoing items.

[0051] The first aspect of the present invention provides a preparation method of a piezoelectric nano-hydroxyapatite modified titanium-based implant. A piezoelectric nano-hydroxyapatite solution is dropped onto a titanium plate with a titanium dioxide nanotube array, and then the titanium plate is placed in a vacuum drying oven for vacuum adsorption. After the surface of the titanium plate is dried, the dropping is repeated. Then, the titanium plate is rinsed with a buffer solution, and finally vacuum dried to obtain the piezoelectric nano-hydroxyapatite modified titanium-based implant.

[0052] The preparation method provided by the present invention first uses a vacuum adsorption combined with a repeated dropping strategy to effectively fill the three-dimensional pore structure of the titanium dioxide nanotube array with piezoelectric nano-hydroxyapatite. The vacuum negative pressure drives the deep penetration of the solution, realizing the physical anchoring and chemical bonding of the nanoparticles on the surface of the titanium substrate, and forming a mechanical interlocking structure. Secondly, the cyclic process of multiple dropping and vacuum adsorption constructs a gradient structure that transitions from a dense bottom layer to a porous surface layer at the nanoscale. This biomimetic structure not only retains the topological cell growth promoting effect of the nanotube array, but also forms a dual bioactive interface through the charge stimulation of piezoelectric nano-hydroxyapatite. The buffer flushing removes weakly bound particles, exposes the active crystal planes, and optimizes the calcium and phosphate ion release rate. This preparation method realizes the synergistic optimization of the bioactivity, mechanical properties and functional characteristics of the implant surface through a physical-chemical synergistic mechanism, and has significant clinical application value.

[0053] Furthermore, the concentration of the piezoelectric nano-hydroxyapatite solution is 5-15 mg / mL.

[0054] Typically but not restrictively, the concentration of the piezoelectric nano-hydroxyapatite solution can be, for example, 5 mg / mL, 6 mg / mL, 7 mg / mL, 8 mg / mL, 9 mg / mL, 10 mg / mL, 11 mg / mL, 12 mg / mL, 13 mg / mL, 14 mg / mL, 15 mg / mL, or any value within the range of 5 mg / mL to 15 mg / mL.

[0055] Preferably, the solvent in the piezoelectric nano-hydroxyapatite solution includes PBS solution, physiological saline solution, simulated body fluid, deionized water, DPBS solution, HBSS solution or EBSS solution.

[0056] PBS solution is phosphate buffer; simulated body fluid is an artificially synthesized solution whose composition and ion concentration are similar to human body fluids (such as plasma, synovial fluid, etc.), and is mainly used to simulate the physiological environment in the human body; DPBS solution is a modified phosphate buffer; HBSS solution (Hanks' Balanced Salt Solution) and EBSS solution (Earle's Balanced Salt Solution) are both balanced salt solutions (Balanced Salt Solution, BSS).

[0057] Preferably, the buffer is the solvent in the piezoelectric nano-hydroxyapatite solution.

[0058] Preferably, the ratio of the usage amount of the piezoelectric nano-hydroxyapatite solution to the area of the titanium plate is 10-50 μL / cm 2 .

[0059] Typically but not restrictively, the ratio of the usage amount of the piezoelectric nano-hydroxyapatite solution to the area of the titanium plate can be, for example, 10 μL / cm 2 , 15 μL / cm 2 , 20 μL / cm 2 , 25 μL / cm 2 , 30 μL / cm 2 , 35 μL / cm 2 , 40 μL / cm 2 , 45 μL / cm 2 , 50 μL / cm 2 , or it can also be 10 μL / cm 2 ~50 μL / cm 2 and can be any value within this range.

[0060] Furthermore, the number of times of repeated dropping is 2 to 8 times.

[0061] Typically but not restrictively, the number of times of repeated dropping can be, for example, 2 times, 3 times, 4 times, 5 times, 6 times, 7 times or 8 times.

[0062] Preferably, the number of times of rinsing is 1 to 5 times.

[0063] Typically but not restrictively, the number of times of rinsing can be, for example, 1 time, 2 times, 3 times, 4 times or 5 times.

[0064] Furthermore, the temperature of the vacuum drying is 50 to 150 °C, and the time is 30 to 90 min.

[0065] Furthermore, the preparation method of the titanium plate with a titanium dioxide nanotube array is as follows: an anodic oxidation method is used to prepare a titanium dioxide nanotube array on the surface of the titanium plate, and finally annealing is carried out to obtain the titanium plate with the titanium dioxide nanotube array.

[0066] Preferably, in the titanium dioxide nanotube array, the length of the titanium dioxide nanotubes is 3 to 8 μmnm, and the diameter is 50 to 200 nm.

[0067] In the actual preparation process, the diameter and length of the nanotubes can be adjusted according to the oxidation voltage and the duration. The higher the voltage, the larger the tube diameter; the longer the time, the longer the nanotubes. The oxidation voltage and time can be adjusted according to specific requirements until the length and diameter of the titanium dioxide nanotubes meet the requirements. The oxidation voltage and time are not specifically limited in the present invention, and any oxidation voltage and time that can prepare titanium dioxide nanotubes meeting the requirements are acceptable.

[0068] Typically but not limited thereto, the length of the titanium dioxide nanotubes can be, for example, 3 μm, 4 μm, 5 μm, 6 μm, 7 μm or 8 μm, or any value within the range of 3 to 8 μm; the diameter of the titanium dioxide nanotubes can be, for example, 50 nm, 100 nm, 150 nm or 200 nm, or any value within the range of 50 nm to 200 nm.

[0069] Preferably, the heating rate of the annealing is 1 to 3 °C / min.

[0070] Typically but not limited thereto, the heating rate of the annealing can be, for example, 1 °C / min, 1.5 °C / min, 2 °C / min, 2.5 °C / min or 3 °C / min, or any value within the range of 1 °C / min to 3 °C / min.

[0071] Preferably, the annealing is carried out in air, the annealing temperature is 400 to 500 °C, and the time is 1 to 3 h.

[0072] Typically but not limited thereto, the annealing is carried out in air, the annealing temperature can be, for example, 400 °C, 410 °C, 420 °C, 430 °C, 440 °C, 450 °C, 460 °C, 470 °C, 480 °C, 490 °C or 500 °C, or any value within the range of 400 °C to 500 °C; the annealing time can be, for example, 1 h, 1.5 h, 2 h, 2.5 h or 3 h, or any value within the range of 1 h to 3 h.

[0073] Furthermore, the preparation method of the piezoelectric nano-hydroxyapatite is as follows:

[0074] The phosphorus source solution is slowly added dropwise to the calcium source solution, and after completion, the mixture is stirred evenly to obtain a mixed solution; the mixed solution is subjected to hydrothermal reaction at 160 to 200 °C for 18 to 24 h. After the reaction is completed, the product is separated by solid-liquid separation, washed and dried in vacuum to obtain the piezoelectric nano-hydroxyapatite.

[0075] Typically but not limited thereto, the hydrothermal reaction of the mixed solution is carried out at a temperature of 160 °C, 165 °C, 170 °C, 175 °C, 180 °C, 185 °C, 190 °C, 195 °C or 200 °C, and the reaction time can be 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours or 24 hours, or any combination within the range of a temperature of 160 °C to 200 °C and a time of 18 hours to 24 hours.

[0076] Furthermore, the dropping rate of the phosphorus source solution is 0.5 to 1.5 mL / min.

[0077] Typically but not limited thereto, the dropping rate of the phosphorus source solution can be, for example, 0.5 mL / min, 0.6 mL / min, 0.7 mL / min, 0.8 mL / min, 0.9 mL / min, 1.0 mL / min, 1.1 mL / min, 1.2 mL / min, 1.3 mL / min, 1.4 mL / min or 1.5 mL / min, or can be any value within the range of 0.5 mL / min to 1.5 mL / min.

[0078] Preferably, the temperature of the vacuum drying is 50 - 80 °C, the time is 12 - 24 h, and the vacuum degree ≤ 0.1 MPa.

[0079] Typically but not limited thereto, the temperature of the vacuum drying can be, for example, 50 °C, 55 °C, 60 °C, 65 °C, 70 °C, 75 °C or 80 °C, or can be any value within the range of 50 °C to 80 °C; the time is 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours or 24 hours, or can be any value within the range of 12 hours to 24 hours; the vacuum degree should be controlled within the range of ≤ 0.1 MPa.

[0080] Preferably, the preparation method of the piezoelectric nano-hydroxyapatite further includes an ultrasonic dispersion process after solid-liquid separation and before washing.

[0081] Preferably, the power of the ultrasonic dispersion is 100 - 200 W, and the time is 5 - 15 min.

[0082] Typically but not limited thereto, the power of the ultrasonic dispersion can be, for example, 100 W, 110 W, 120 W, 130 W, 140 W, 150 W, 160 W, 170 W, 180 W, 190 W or 200 W, or can be any value within the range of 100 W to 200 W; the time of the ultrasonic dispersion can be, for example, 5 minutes, 6 minutes, 7 minutes, 8 minutes, 9 minutes, 10 minutes, 11 minutes, 12 minutes, 13 minutes, 14 minutes or 15 minutes, or can be any value within the range of 5 minutes to 15 minutes.

[0083] Furthermore, the calcium source in the calcium source solution is calcium chloride.

[0084] Preferably, the phosphorus source in the phosphorus source solution is ammonium dihydrogen phosphate.

[0085] Preferably, the molar ratio of calcium in the calcium source to phosphorus in the phosphorus source is 1:

[0086] (0.6 - 0.7).

[0087] Typically but not restrictively, the molar ratio of calcium to phosphorus can be, for example, 1:0.6, 1:0.62, 1:0.64, 1:0.66, 1:0.68 or 1:0.7, or can also be any value within the range of 1:(0.6 to 0.7).

[0088] Preferably, the phosphorus source solution and / or the solvent in the phosphorus source solution is an ammonia water solution.

[0089] The second aspect of the present invention provides a piezoelectric nano-hydroxyapatite modified titanium-based implant, which is prepared by using the preparation method described in the first aspect, and its structural schematic diagram is as Figure 1 shown. A porous titanium dioxide array grows regularly on the titanium plate, and piezoelectric nano-hydroxyapatite is loaded in the porous titanium dioxide array.

[0090] The piezoelectric nano-hydroxyapatite modified titanium-based implant provided by the present invention realizes antibacterial-osteogenic synergistic functional design by constructing a titanium dioxide nanotube array on the surface of the titanium plate and loading nano-hydroxyapatite with enhanced piezoelectric performance as a sonosensitizer. This technology utilizes the deep tissue penetration advantage of sonodynamic therapy (SDT) to activate the piezoelectric nano-hydroxyapatite sonosensitizer to generate reactive oxygen species (ROS) under ultrasonic stimulation, efficiently killing bacteria; at the same time, nano-hydroxyapatite, as a natural component of bone tissue, directly participates in the bone repair process by promoting osteoblast adhesion and mineralization, significantly enhancing the bone integration ability of the implant. In addition, the porous high specific surface area structure of the TiO2 nanotube array ensures the long-term stable loading and controlled release of the sonosensitizer, and its piezoelectric characteristics can be precisely regulated by ultrasound to achieve infection-responsive on-demand treatment, avoiding drug resistance or tissue damage caused by unnecessary release of antibacterial agents. This technology breaks through the limitations of the existing implant passive antibacterial and active bactericidal strategies, while ensuring long-term antibacterial efficacy during the service life, reducing the loosening risk through biomimetic bone component design, and providing an innovative path for the biofunctional surface modification of orthopedic implants.

[0091] The third aspect of the present invention provides the application of the piezoelectric nano-hydroxyapatite modified titanium-based implant in the preparation of joint prostheses, bone filling materials or fracture internal fixation materials.

[0092] The application of the piezoelectric nano-hydroxyapatite modified titanium-based implant provided by the present invention, in view of the advantages of the above-mentioned piezoelectric nano-hydroxyapatite modified titanium-based implant, improves the biofunctionality of downstream products such as joint prostheses, bone filling materials or fracture internal fixation materials, and promotes the development of downstream industries.

[0093] The present invention will be further illustrated below through specific examples and comparative examples. However, it should be understood that these examples are only for more detailed illustration and should not be construed as limiting the present invention in any way. For the raw materials used in the examples and comparative examples of the present invention, unless otherwise specified, they are carried out under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used, unless otherwise indicated by the manufacturer, are all conventional products that can be obtained through commercial purchase.

[0094] Example 1

[0095] In this example, a piezoelectric nano-hydroxyapatite modified titanium-based implant was prepared, and the preparation method was as follows:

[0096] 1. Weigh 0.14702 g (0.001 mol) of calcium chloride dihydrate into a 10 mL centrifuge tube, add 10 mL of water and 150 μL of ammonia water, and ultrasonically mix evenly to obtain a calcium chloride dihydrate solution.

[0097] Weigh 0.077 g (0.00067 mol) of ammonium dihydrogen phosphate into a 10 mL centrifuge tube, add 10 mL of water and 150 μL of ammonia water, and ultrasonically mix evenly to obtain an ammonium dihydrogen phosphate solution.

[0098] Transfer the calcium chloride dihydrate solution to the inner liner of a 20 mL autoclave, and slowly drop the ammonium dihydrogen phosphate solution into it. Stir at 300 rmp for 10 min; transfer the well-stirred solution to a 20 mL autoclave, set the temperature to 180 °C, and the reaction time to 20 h. After the reaction is completed and cooled to room temperature, transfer the product in the autoclave to a 50 mL centrifuge tube, centrifuge to discard the supernatant, with a rotation speed of 10000 r / min and a centrifugation time of 10 min. Wash the precipitate with water, ultrasonically disperse the solid evenly in the liquid, centrifuge again, remove the precipitate and then wash and centrifuge once more; place the washed precipitate in a vacuum drying oven and let it stand for drying to obtain piezoelectric nano-hydroxyapatite.

[0099] 2. Prepare a titanium dioxide nanotube array on the surface of the titanium alloy by anodic oxidation. The length of the TiO2 nanotubes is 5 μm and the diameter is 100 nm. Anneal the titanium plate with TiO2 nanotubes in an air atmosphere at 450 °C for 2 h, with a heating rate of 2 °C·min -1 , to obtain a titanium plate with a titanium dioxide nanotube array.

[0100] 3. Cut the titanium plate with a titanium dioxide nanotube array into a circle with a diameter of 1 cm, and then drop 25 μL of a piezoelectric nano-hydroxyapatite solution (concentration: 10 mg / mL, solvent: PBS solution) onto the titanium plate. Place the titanium plate in a vacuum drying oven for vacuum adsorption, take it out after the surface is dry, and repeat the cycle 5 times. Use a pipette to aspirate 0.5 mL of PBS buffer to rinse both sides of the titanium plate 3 times each, and then put it back into the vacuum drying oven and dry it at 100 °C for 60 min to obtain a TiO2 nanotube array loaded with piezoelectric nano-hydroxyapatite.

[0101] Example 2

[0102] In this example, a piezoelectric nano-hydroxyapatite modified titanium-based implant is prepared, and the preparation method is as follows:

[0103] 1. The same step as in Example 1.

[0104] 2. The same step as in Example 1.

[0105] 3. Cut the titanium plate with a titanium dioxide nanotube array into a circle with a diameter of 1 cm, and then drop 50 μL of a piezoelectric nano-hydroxyapatite solution (concentration: 5 mg / mL, solvent: PBS solution) onto the titanium plate. Place the titanium plate in a vacuum drying oven for vacuum adsorption, take it out after the surface is dry, and repeat the cycle 5 times. Use a pipette to aspirate 0.5 mL of PBS buffer to rinse both sides of the titanium plate 3 times each, and then put it back into the vacuum drying oven and dry it at 100 °C for 60 min to obtain a TiO2 nanotube array loaded with piezoelectric nano-hydroxyapatite.

[0106] Example 3

[0107] In this example, a piezoelectric nano-hydroxyapatite modified titanium-based implant is prepared, and the preparation method is as follows:

[0108] 1. The same step as in Example 1.

[0109] 2. The same step as in Example 1.

[0110] 3. Cut the titanium plate with a titanium dioxide nanotube array into a circle with a diameter of 1 cm, and then drop 10 μL of a piezoelectric nano-hydroxyapatite solution (concentration: 15 mg / mL, solvent: PBS solution) onto the titanium plate. Place the titanium plate in a vacuum drying oven for vacuum adsorption, take it out after the surface is dry, and repeat the cycle 5 times. Use a pipette to aspirate 0.5 mL of PBS buffer to rinse both sides of the titanium plate 3 times each, and then put it back into the vacuum drying oven and dry it at 100 °C for 60 min to obtain a TiO2 nanotube array loaded with piezoelectric nano-hydroxyapatite.

[0111] Example 4

[0112] In this example, a piezoelectric nano-hydroxyapatite modified titanium-based implant was prepared. Different from Example 1, in step 2, the length of the TiO2 nanotubes was controlled to be 3 μm and the diameter was 50 nm. The remaining preparation methods were the same as those in Example 1 and will not be elaborated here.

[0113] Example 5

[0114] In this example, a piezoelectric nano-hydroxyapatite modified titanium-based implant was prepared. Different from Example 1, in step 2, the length of the TiO2 nanotubes was controlled to be 8 μm and the diameter was 200 nm. The remaining preparation methods were the same as those in Example 1 and will not be elaborated here.

[0115] Example 6

[0116] In this example, a piezoelectric nano-hydroxyapatite modified titanium-based implant was prepared. Different from Example 1, in step 2, the length of the TiO2 nanotubes was controlled to be 15 μm and the diameter was 100 nm. The remaining preparation methods were the same as those in Example 1 and will not be elaborated here.

[0117] Comparative Example 1

[0118] In this comparative example, a hydroxyapatite modified titanium-based implant was prepared. Different from Example 1, ordinary hydroxyapatite was used to replace the piezoelectric nano-hydroxyapatite, and the remaining raw materials and preparation methods were the same as those in Example 1 and will not be elaborated here.

[0119] Test Example 1

[0120] The piezoelectric nano-hydroxyapatite prepared in Example 1 was observed and photographed for particle images using an electron projection microscope (TEM) to obtain Figure 2 ; the energy spectrum was detected using X-ray photoelectron spectroscopy (XPS) to obtain Figure 3 ; the crystal structure was detected using X-ray diffraction to obtain Figure 4 ; the current response of the electrode made of this piezoelectric nano-hydroxyapatite was measured under ultrasonic vibration to obtain Figure 5 .

[0121] From Figure 2 and Figure 5 it can be seen that the piezoelectric nano-hydroxyapatite is rod-shaped, the aspect ratio is about 4:1, the dominant crystal plane is the 211 plane, the calcium-phosphorus wall is 1.34:1, and it can exhibit good piezoelectric effects under the action of current.

[0122] Test Example 2: In vitro antibacterial experiment

[0123] Staphylococcus aureus (S. aureus, X387) was inoculated into 10 mL of tryptone soy broth (TSB) and cultured in a constant temperature incubator (5% CO2, 37°C) for 16 h. Then, it was centrifuged at 8000 rpm for 10 min, the supernatant was discarded, 5 mL of PBS solution was added, and the concentration was adjusted to 1×10 6 CFU / mL for later use. The plate coating method was used to evaluate the effectiveness of the titanium-based implants (titanium-based implants provided by the examples and comparative examples respectively) against S. aureus under ultrasonic stimulation.

[0124] The titanium-based implants (titanium-based implants provided by the examples and comparative examples respectively) were placed in a 24-well plate, and 2 wells were kept blank as control group 1 and control group 2, and another well was inoculated with the titanium-based implant of Example 1 as control group 3. 1 mL of bacterial solution was inoculated into each well and subjected to ultrasonic stimulation (1.0 MHz, 1.5 W / cm 2 , 100% duty cycle) for 10 minutes (control group 1 and control group 3 were not subjected to ultrasonic waves). The plate coating method was used to culture the bacterial growth on an agar plate at 37°C for 12 hours to quantify the colony-forming units (CFU). The data obtained were recorded in Table 1.

[0125] Table 1

[0126] Antibacterial rate Antibacterial rate Example 1 99％ Example 4 98％ Example 2 99％ Example 5 98％ Example 3 98％ Example 6 90％ Comparative example 1 30％ Control group 1 0％ Control group 2 30％ Control group 3 0％

[0127] Figure 6 is the antibacterial effect diagram of plate coating; Figure 7 is the bar chart of bacterial count of plate coating.

[0128] Through Figure 6 and Figure 7 it can be seen that the surface of the piezoelectric nano-hydroxyapatite modified titanium-based implant of the present invention has a significant antibacterial effect under the stimulation of ultrasonic waves.

[0129] Test Example 3: In vitro biocompatibility experiment

[0130] MC3T3-E1 cells were cultured with α-minimum essential medium (α-MEM, Gibcom, USA), including 10% fetal bovine serum (FBS, Gibcom, USA) and 1% penicillin-streptomycin solution. The culture was incubated in a humidified incubator at 37°C with 5% CO2 in the culture medium, and co-cultured with the titanium-based implant provided in Example 1. The cell viability and proliferation were determined by the CCK-8 method. The optical density (OD) at 450 nm was measured with an enzyme-linked immunosorbent assay (ELISA) reader on the 1st, 3rd, and 5th days.

[0131] Through Figure 8 it can be seen that the surface of the piezoelectric nano-hydroxyapatite modified titanium-based implant of Example 1 has high biocompatibility and has the effect of promoting the proliferation of osteoblasts.

[0132] Test Example 4: Animal Experiment

[0133] In order to study the bone repair promoting effect of the piezoelectric nano-hydroxyapatite modified titanium-based implant surface in Example 1, the following groups were divided: a blank surgery group, a vancomycin group, and an implant combined with ultrasound group.

[0134] In this experiment, a rat tibial osteomyelitis model was used, and samples were collected 4 weeks after surgery to compare the degree of bone destruction and bone regeneration among the groups.

[0135] Figure 9 The scale bar in the upper row of images is 400 μm, and the scale bar in the lower row of images is 100 μm. Figure 9 HE staining showed that the blank surgery group had local hyperplasia of fibrous connective tissue, which was loosely arranged and accompanied by a small amount of bleeding; more necrotic fragments were seen; and there were few bone marrow cells in the bone marrow cavity. The trabecular morphology of the bone tissue in the vancomycin group and the implant combined with ultrasound group was normal, and the morphology and structure of the bone cells were normal; there was no significant change in the size of the bone marrow cavity, and a large number of bone marrow cells and a small number of fat cells were seen in the bone marrow cavity; no obvious necrosis and inflammatory cell infiltration were observed. The bone marrow cells in the bone marrow cavity of the implant combined with ultrasound group were more densely arranged, and the degree of bone regeneration was higher.

[0136] Finally, it should be noted that the above-described embodiments are only specific implementations of the present invention, which are used to illustrate the technical solutions of the present invention, rather than to limit them. The protection scope of the present invention is not limited thereto. Although the present invention is described in detail with reference to the above-described embodiments, ordinary technicians in the field should understand that any technician familiar with the technical field can still modify the technical solutions recorded in the above-described embodiments within the technical scope disclosed by the present invention, or can easily think of changes, or make equivalent replacements for some of the technical features therein; and these modifications, changes or replacements do not make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of the embodiments of the present invention, and should be included in the protection scope of the present invention. Therefore, the protection scope of the present invention shall be based on the protection scope of the claims.

Claims

1. A preparation method of a piezoelectric nano-hydroxyapatite modified titanium-based implant, characterized in that, Drop the piezoelectric nano-hydroxyapatite solution on the titanium plate with a titanium dioxide nanotube array, then place the titanium plate in a vacuum drying oven for vacuum adsorption. After the surface of the titanium plate is dry, repeat the dropping. Then rinse the titanium plate with a buffer solution, and finally vacuum dry to obtain the piezoelectric nano-hydroxyapatite modified titanium-based implant.

2. The preparation method according to claim 1, wherein The concentration of the piezoelectric nano-hydroxyapatite solution is 5-15 mg / mL; Preferably, the solvent in the piezoelectric nano-hydroxyapatite solution includes PBS solution, physiological saline solution, simulated body fluid, deionized water, DPBS solution, HBSS solution or EBSS solution; Preferably, the buffer solution is the solvent in the piezoelectric nano-hydroxyapatite solution; Preferably, the ratio of the usage amount of the piezoelectric nano-hydroxyapatite solution to the area of the titanium plate is 10-50 μL / cm 2 .

3. The preparation method according to claim 1, characterized in that, The number of times of repeated dropping is 2-8 times; Preferably, the number of times of rinsing is 1-5 times.

4. The preparation method according to claim 1, characterized in that, The temperature of the vacuum drying is 50-150 °C, and the time is 30-90 min.

5. The preparation method according to any one of claims 1 to 4, characterized in that, The preparation method of the titanium plate with a titanium dioxide nanotube array is: use an anodic oxidation method to prepare a titanium dioxide nanotube array on the surface of the titanium plate, and finally anneal to obtain the titanium plate with a titanium dioxide nanotube array; Preferably, in the titanium dioxide nanotube array, the length of the titanium dioxide nanotubes is 3-8 μm, and the diameter is 50-200 nm; Preferably, the heating rate of the annealing is 1-3 °C / min; Preferably, the annealing is carried out in air, the annealing temperature is 400-500 °C, and the time is 1-3 h.

6. The preparation method according to any one of claims 1 to 4, characterized in that, The preparation method of the piezoelectric nano-hydroxyapatite is: Slowly drop the phosphorus source solution into the calcium source solution, and after completion, mix evenly to obtain a mixed solution; hydrothermally react the mixed solution at 160-200 °C for 18-24 h. After the reaction is completed, separate the product from the solid-liquid, wash and vacuum dry to obtain the piezoelectric nano-hydroxyapatite.

7. The preparation method according to claim 6, characterized in that, The dropping rate of the phosphorus source solution is 0.5-1.5 mL / min; Preferably, the temperature of the vacuum drying is 50-80 °C, the time is 12-24 h, and the vacuum degree ≤ 0.1 MPa; Preferably, the preparation method of the piezoelectric nano-hydroxyapatite further includes an ultrasonic dispersion process after solid-liquid separation and before washing; Preferably, the power of the ultrasonic dispersion is 100-200 W, and the time is 5-15 min.

8. The preparation method according to claim 6, characterized in that, The calcium source in the calcium source solution is calcium chloride; Preferably, the phosphorus source in the phosphorus source solution is ammonium dihydrogen phosphate; Preferably, the molar ratio of calcium in the calcium source to phosphorus in the phosphorus source is 1: (0.6～0.7)； Preferably, the phosphorus source solution and / or the solvent in the phosphorus source solution is an ammonia water solution.

9. A piezoelectric nano-hydroxyapatite modified titanium-based implant, characterized in that, Prepared by using the preparation method described in any one of claims 1-8.

10. Use of the piezoelectric nano-hydroxyapatite modified titanium-based implant described in claim 9 in the preparation of joint prostheses, bone filling materials or fracture internal fixation materials.

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