Zirconium oxide osseointegration implant material and preparation method thereof

Abstract

The invention relates to the technical field of dental and orthopedic implants, in particular to a zirconia osseointegration implant material and a preparation method thereof. Aiming at the problems of strong biological inertness and slow early-stage osseointegration of the zirconium oxide ceramic surface, the preparation method comprises the following steps: cleaning a zirconium oxide matrix and carrying out alkaline activation; forming a polydopamine adhesion bottom layer; branched polyethyleneimine is fixed to enhance the amino density; constructing a network layer through alternate coordination of phytic acid and calcium / strontium ions; strontium doped hydroxyapatite nano particles are immobilized; finally, biomimetic mineralization is carried out in simulated body fluid. According to the method, a micro-nano composite active interface is formed, the biological activity, ion release controllability and osseointegration capability of the material are remarkably improved, and osteoblast response is promoted.

Description

Technical Field

[0001] This invention relates to the field of dental and orthopedic implant technology, and more particularly to a zirconia osseointegrated implant material and its preparation method. Background Technology

[0002] Zirconia ceramics are widely used in dental and orthopedic implants due to their excellent aesthetic properties and corrosion resistance. However, the material exhibits significant bioinertness, resulting in weak interaction with bone tissue and difficulty in rapidly forming effective osseointegration after implantation. This inert interface delays osteoblast adhesion and differentiation, which, in clinical applications, especially in complex physiological environments such as oral occlusal micromovements or osteoporosis, can easily lead to poor early stability, affecting the long-term success rate of implants.

[0003] To improve osseointegration performance, conventional techniques often employ physicochemical methods such as sandblasting and acid etching to roughen the surface, increasing specific surface area and roughness. However, these methods are sensitive to the microstructure of zirconia, easily inducing phase transformations or microcracks, thus weakening the mechanical strength of the material. Furthermore, the roughened surface morphology is often uneven, making it difficult to controllably introduce bioactive components, leading to unstable osteogenic signal release or easy coating detachment.

[0004] Furthermore, existing modification strategies often focus on single-level surface treatments, such as simple coatings or ion doping, lacking multi-level synergistic interface design. For example, while hydroxyapatite coatings alone can enhance bioactivity, the coating has weak adhesion to the matrix and is easily degraded in body fluids; while single ion implantation has a short-lived release behavior and cannot maintain continuous osteogenic stimulation. This functional singularity limits the dynamic regulation capability of the interface microenvironment and cannot meet the dual requirements of rapid osteogenic formation and stable anchoring for early osteointegration.

[0005] Overall, existing technologies struggle to construct composite interfaces on the surface of zirconia implants that combine high bonding strength, sustained bioactivity, and mechanical stability. Their bioinert nature and the limitations of roughening methods jointly restrict improvements in clinical performance, necessitating a breakthrough in surface modification strategies. Summary of the Invention

[0006] In view of this, the purpose of this invention is to propose a zirconia osseointegration implant material and its preparation method, so as to solve the problem that existing zirconia implants have insufficient initial stability due to their strong surface bioinertness, slow early osseointegration speed, and the fact that conventional surface roughening methods can easily damage the material phase structure, making it difficult to form a stable osteogenic microenvironment while maintaining mechanical strength.

[0007] To achieve the above objectives, the present invention provides a method for preparing a zirconia bone-integrated implant material, comprising the following steps: (1) The zirconia matrix is ​​cleaned and alkaline activated to obtain an activated zirconia matrix; (2) The activated zirconia matrix is ​​placed in a hydrochloric acid dopamine solution in a weakly alkaline buffer system to allow the hydrochloric acid dopamine to self-polymerize and deposit, forming a polydopamine adhesion layer on the surface of the activated zirconia matrix. (3) To fix the branched polyethyleneimine on the polydopamine adhesive substrate; (4) The zirconia matrix treated in step (3) is alternately treated with phytic acid working solution and working solution containing calcium ions and strontium ions, and this process is repeated two to four times, so that phytic acid molecules coordinate and crosslink with calcium ions and strontium ions to form a coordination network layer containing calcium / strontium ions. (5) The zirconium oxide matrix treated in step (4) is brought into contact with the dispersion of strontium-doped hydroxyapatite nanoparticles, so that the strontium-doped hydroxyapatite nanoparticles are immobilized on the surface of the coordination network layer. (6) The zirconia matrix treated in step (5) is placed in a simulated body fluid for biomimetic mineralization deposition, then washed and dried to obtain zirconia bone integration implant material.

[0008] Preferably, the zirconia matrix is ​​a tetragonal polycrystalline zirconia ceramic matrix containing yttrium oxide stabilizer.

[0009] Preferably, the cleaning in step (1) is performed by ultrasonic cleaning with acetone, anhydrous ethanol and deionized water in sequence, and the ultrasonic time for each solvent is 5 min to 20 min.

[0010] Preferably, the alkaline activation treatment in step (1) is performed by soaking in an aqueous sodium hydroxide solution, wherein the mass fraction of the aqueous sodium hydroxide solution is 1% to 3%, the treatment temperature is 55℃ to 65℃, and the treatment time is 20 min to 40 min.

[0011] Preferably, the weakly alkaline buffer system in step (2) is a tris(hydroxymethyl)aminomethane buffer system, wherein the concentration of tris(hydroxymethyl)aminomethane is 8 mmol / L to 12 mmol / L and the pH of the solution is 8.3 to 8.7.

[0012] Preferably, based on 10 zirconia substrates with a size of Φ10mm×2mm, the concentration of dopamine hydrochloride in step (2) is 1mg / mL to 3mg / mL, the reaction temperature is 23℃ to 27℃, the reaction time is 4h to 8h, and the stirring speed is 150r / min to 220r / min.

[0013] Preferably, based on 10 zirconia substrates with a size of Φ10mm×2mm, the weight-average molecular weight of the branched polyethyleneimine in step (3) is 20,000 to 30,000, the working solution concentration is 0.5mg / mL to 1.5mg / mL, the reaction temperature is 20℃ to 30℃, the reaction time is 45min to 1h30min, and the stirring speed is 80r / min to 120r / min.

[0014] Preferably, based on 10 zirconia substrates with a size of Φ10mm×2mm, the pH of the phytic acid working solution in step (4) is 6.8 to 7.2, the phytic acid concentration is 0.3mg / mL to 0.8mg / mL, and the phytic acid adsorption time is 8min to 15min.

[0015] Preferably, in the working solution containing calcium ions and strontium ions in step (4), the calcium salt is selected from at least one of calcium chloride dihydrate, calcium nitrate tetrahydrate and calcium acetate, and the strontium salt is selected from at least one of strontium chloride hexahydrate and strontium nitrate.

[0016] Preferably, based on 10 zirconia substrates with a size of Φ10mm×2mm, the calcium ion concentration in step (4) is 30mmol / L~60mmol / L, the strontium ion concentration is 3mmol / L~10mmol / L, the molar ratio of calcium ion to strontium ion is 5:1~15:1, and the calcium / strontium ion coordination time is 1min~3min.

[0017] Preferably, the strontium doping ratio of the strontium-doped hydroxyapatite nanoparticles in step (5) is 3% to 7%, and the average particle size is 20 nm to 50 nm.

[0018] Preferably, based on 10 zirconium oxide substrates with a size of Φ10mm×2mm, the concentration of nanoparticles in the strontium-doped hydroxyapatite nanoparticle dispersion in step (5) is 0.5mg / mL to 2mg / mL.

[0019] Preferably, in step (5), the ultrasonic dispersion power is 100W to 250W and the ultrasonic time is 8min to 15min.

[0020] Preferably, in step (5), the oscillation or stirring speed during the immobilization process of the strontium-doped hydroxyapatite nanoparticles is 100 r / min to 150 r / min, and the immobilization time is 20 min to 45 min.

[0021] Preferably, the simulated body fluid in step (6) is Solarbio product number G0392, the biomimetic mineralization reaction temperature is 37℃, and the biomimetic mineralization reaction time is 6h~12h.

[0022] Preferably, the zirconia bone-integrated implant material is sterilized with ethylene oxide and analyzed after drying.

[0023] Furthermore, the present invention also provides a zirconia bone-integrated implant material, obtained by the above preparation method.

[0024] The beneficial effects of this invention are: This invention provides a robust organic interface foundation for the zirconia matrix through the synergistic effect of the polydopamine adhesive underlayer and the polyamine-functionalized layer. Under weakly alkaline conditions, polydopamine self-polymerizes to form a uniformly covering adhesive layer, which not only enhances surface hydrophilicity but also forms a strong bond with the matrix through its catechol groups, laying a stable anchoring foundation for subsequent modification steps. The introduction of branched polyethyleneimine further enriches the surface amine group density, enhancing the chelating agent adsorption capacity through electrostatic interactions and covalent coupling, thereby improving interfacial chemical activity.

[0025] In the network layer constructed by alternating coordination of chelating agents and calcium and strontium ions, phytic acid molecules form multi-point coordination with the polyamine substrate through phosphate groups, and efficiently chelate with calcium and strontium ions to form a dense ion storage structure. This network layer not only achieves controllable loading of calcium and strontium ions, but also enhances the mechanical stability of the interface through coordination crosslinking, providing a continuous ion release source for biomimetic mineralization.

[0026] The immobilization of strontium-doped hydroxyapatite nanoparticles further introduced high specific surface area nucleation sites, and their nanoscale effect, together with the strontium doping characteristics, promoted the directional deposition of bone-like apatite. In simulated body fluids, the nanoparticles acted as templates to induce the uniform growth of calcium phosphate salts, forming a micro-nano composite active coating layer, which effectively enhanced the surface biomineralization capacity.

[0027] The biomimetic mineralization process simulates physiological temperature and body fluid environment, enabling nanoparticles and coordination network layers to synergistically guide the orderly deposition of apatite crystals. This process not only enhances the chemical affinity between the interface and bone tissue but also activates osteoblast signaling pathways through the continuous release of strontium ions, thereby promoting early bone contact and integration.

[0028] The overall approach achieves multi-dimensional regulation of interfacial activity through stepwise construction of molecular-level adhesion, ionic-level coordination, and nanoscale nucleation, significantly enhancing the bioresponsiveness and long-term stability of zirconia implants. Attached Figure Description

[0029] To more clearly illustrate the technical solutions in this invention or the prior art, the accompanying drawings used in the description of the embodiments or the prior art will be briefly introduced below.

[0030] Figure 1 The infrared spectra of the samples from Example 1 and Comparative Examples 1, 2 and 4 of this invention are shown below. Figure 2 This is the calcium ion release curve of the sample in Example 1 of the present invention; Figure 3 This is the strontium ion release curve of the sample in Example 1 of the present invention. Detailed Implementation

[0031] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments.

[0032] A zirconia bone-integrated implant material and its preparation method While existing zirconia implants possess excellent aesthetics and corrosion resistance, their strong bioinertness and slow early osseointegration lead to insufficient initial stability in clinical settings such as oral occlusal micromovement and osteoporosis. Furthermore, conventional roughening methods like sandblasting and acid etching are sensitive to the structure and defects of the zirconia phase, making it difficult to create a stable osteogenic microenvironment without sacrificing strength. Therefore, a chemical modification pathway is needed that uses zirconia as a matrix, employing a polydopamine-bonded substrate and strontium-doped hydroxyapatite nanoparticles to construct a micro / nano composite active interface. This pathway aims to achieve high bonding strength, promote osteogenic differentiation, and enhance early bone contact rate, making it suitable for dental implants and maxillofacial bone fixation implants.

[0033] This invention provides a method for preparing a zirconia bone-integrated implant material. The overall framework is as follows: A zirconia matrix is ​​subjected to organic contamination removal and alkaline activation treatment to form hydroxylated / negatively charged groups on the surface, facilitating subsequent deposition. Subsequently, dopamine hydrochloride undergoes self-polymerization in a weakly alkaline buffer system to form a polydopamine adhesion layer on the zirconia surface. Branched polyethyleneimine is then introduced onto this adhesion layer to provide high-density amine groups and positively charged sites, thereby enhancing the adsorption of phosphate-containing chelating agents. An alternating process of chelating agent adsorption and calcium / strontium ion coordination is then repeated several times to construct a multi-point coordination network layer containing calcium / strontium ions on the surface. Based on this, strontium-doped hydroxyapatite nanoparticles are immobilized as nucleation sites, and biomimetic mineralization deposition is performed in a modified simulated body fluid heated to physiological temperature. Finally, after cleaning, drying, and sterilization, a zirconia bone-integrated implant material with a micro / nano composite active interface is obtained. In the above technical approach, branched polyethyleneimine is introduced to increase the amine group density and surface positive charge, thereby enhancing the adsorption of phosphate-containing molecules (such as phytic acid) and improving multi-point coordination ability. Chelating agents are used to alternately coordinate with calcium / strontium ions and repeated several times to form a denser calcium / strontium-containing coordination network layer. Strontium-doped hydroxyapatite nanoparticles are further immobilized on the coordination network layer and biomimetic mineralization is carried out to construct a micro-nano composite active interface of nanoparticle nucleation sites + biomimetic mineralization deposition, which is beneficial to improve the surface apatite deposition ability and improve the activity of early bone integration-related interfaces.

[0034] In the matrix pretreatment (decontamination + alkaline activation) step provided by the present invention, the zirconia matrix is ​​preferably a tetragonal polycrystalline zirconia ceramic matrix containing yttrium oxide stabilizer; the matrix shape can be a dental implant, maxillofacial / orthopedic fixation piece or a standard sheet for evaluation, wherein it is more preferably a sheet or a regular sample for process development and quality comparison, so as to control the total surface area and liquid-solid ratio (using a circular sheet with a diameter of 10 mm and a thickness of 2 mm).

[0035] For cleaning and decontamination, a combination of acetone-anhydrous ethanol-deionized water for sequential ultrasonic cleaning is preferred; wherein, the ultrasonic time is preferably 5 min-20 min for each solvent, more preferably 8 min-12 min, and the volume ratio of the substrate to the cleaning solvent is such that the substrate is completely immersed and there is no stacking or obstruction during the ultrasonic process. Preferably, the volume of solvent corresponding to each substrate is 8 mL-25 mL, more preferably 9 mL-15 mL.

[0036] For alkaline activation, immersion in an aqueous sodium hydroxide solution is preferred. The mass fraction of the sodium hydroxide solution is preferably 0.5%-5%, more preferably 1%-3%, the treatment temperature is preferably 40℃-80℃, more preferably 55℃-65℃ (60℃ in Example 1), and the treatment time is preferably 10min-60min, more preferably 20min-40min. These alkaline activation conditions affect the degree of hydroxylation and surface energy of the zirconia surface, thus affecting the initial nucleation and uniformity of polydopamine coverage. Insufficient alkaline activation may lead to localized defects in the subsequent coating, while excessive alkaline activation may introduce unnecessary surface defects or roughness fluctuations. As an alternative process, an aqueous potassium hydroxide solution or an ammonia system can also be used for alkaline activation, but an aqueous sodium hydroxide solution is preferred to obtain a stable and controllable surface activation effect and repeatability.

[0037] In the polydopamine adhesion substrate construction step provided by the present invention, it is preferred to carry out the self-polymerization deposition of dopamine hydrochloride in a weakly alkaline buffer system; the buffer is preferably tris(hydroxymethyl)aminomethane, the concentration of which is preferably 5 mmol / L-20 mmol / L, more preferably 8 mmol / L-12 mmol / L, and the pH of the solution is preferably 8-9, more preferably 8.3-8.7.

[0038] The preferred concentration of dopamine hydrochloride is 0.5 mg / mL-5 mg / mL, more preferably 1 mg / mL-3 mg / mL; the preferred reaction temperature is 20℃-30℃, more preferably 23℃-27℃; the preferred reaction time is 2h-12h, more preferably 4h-8h; and the preferred stirring speed is 100r / min-250r / min, more preferably 150r / min-220r / min.

[0039] The temperature, time, and stirring conditions described above collectively affect the growth kinetics and coverage integrity of the polydopamine film, which in turn affects the firmness of subsequent polyethyleneimine fixation and interfacial stability. The preferred reaction atmosphere is air to provide the oxidizing environment required for self-polymerization.

[0040] In the polyamine substrate construction (polyethyleneimine introduction) step provided by the present invention, it is preferable to use branched polyethyleneimine to adsorb / covalently couple and fix it on the polydopamine adhesive substrate, so that the surface presents polyamine functional groups and improves the positive charge, thereby enhancing the adsorption of phosphate-containing chelating agents and improving the subsequent multi-point coordination ability.

[0041] The weight-average molecular weight of branched polyethyleneimine is preferably 10,000-70,000, more preferably 20,000-30,000; the working solution concentration is preferably 0.2 mg / mL-3 mg / mL, more preferably 0.5 mg / mL-1.5 mg / mL; the reaction temperature is preferably 20℃-30℃; the reaction time is preferably 30 min-4 h, more preferably 45 min-1 h30 min; and the stirring speed is preferably 50 r / min-200 r / min, more preferably 80 r / min-120 r / min.

[0042] The molecular weight and concentration affect the surface amine density and polymer layer thickness: insufficient amine density weakens the adsorption of chelating agents and the number of ion coordination sites, while excessively high amine density or excessively thick polymer layers may introduce excessive swelling, affecting the compactness of the coordination network and the uniformity of subsequent particle immobilization. As alternative raw materials, linear polyethyleneimine, polylysine, or modified chitosan can also be used as polymers providing polyamine groups, but branched polyethyleneimine is preferred to obtain higher amine density and stronger electrostatic adsorption capacity.

[0043] In the step of constructing a coordination network layer by adsorption of chelating agents and alternating coordination of calcium / strontium ions provided in this invention, the chelating agent is preferably phytic acid or its water-soluble salt (e.g., sodium phytate), more preferably phytic acid; the pH of the chelating agent working solution is preferably 6-8, more preferably 6.8-7.2, as this pH affects the degree of dissociation and coordination ability of phosphate groups, thereby affecting the efficiency of multi-point coordination crosslinking with calcium / strontium ions and the density of the network layer. The effective concentration of the chelating agent is preferably 0.1 mg / mL-2 mg / mL, more preferably 0.3 mg / mL-0.8 mg / mL.

[0044] The chelating agent adsorption time is preferably 5-30 min, more preferably 8-15 min. The adsorption time affects the coverage of the chelating agent on the polyamine substrate, and thus affects the density of subsequent ion coordination sites. In the calcium / strontium ion working solution, the calcium salt is preferably calcium chloride dihydrate, calcium nitrate tetrahydrate, or calcium acetate, and the strontium salt is preferably strontium chloride hexahydrate or strontium nitrate; more preferably, a combination of calcium chloride dihydrate and strontium chloride hexahydrate is used to obtain a coordination environment with good solubility and stable ionic strength.

[0045] The calcium ion concentration is preferably 20 mmol / L-80 mmol / L, more preferably 30 mmol / L-60 mmol / L; the strontium ion concentration is preferably 1 mmol / L-20 mmol / L, more preferably 3 mmol / L-10 mmol / L; the molar ratio of calcium ions to strontium ions is preferably 2:1-20:1, more preferably 5:1-15:1. The calcium / strontium ion coordination time is preferably 30 s-10 min, more preferably 1 min-3 min.

[0046] The alternating cycle is preferably one to six times, more preferably two to four times. The number of cycles affects the thickness and ion reserve of the coordination network layer: insufficient cycles may result in a loose network layer and low ion content; excessive cycles may result in an overly thick network layer and internal stress accumulation, thereby affecting the uniformity and wash fastness of subsequent nanoparticle immobilization. This alternating process can form an interfacial network with multi-point coordination cross-linking characteristics and introduce calcium and strontium ions, providing an ionic basis for subsequent hydroxyapatite-related mineralization, thereby improving the interfacial activity related to early bone integration.

[0047] Furthermore, as alternative raw materials for chelating agents, any material capable of forming stable adsorption on the polyamine substrate and providing phosphate or phosphonic acid groups that can coordinate with calcium / strontium ions at multiple points can achieve the technical effects of this invention. Preferably, small molecules containing polyphosphate / phosphonic acid groups, or their salts, such as pyrophosphate, tripolyphosphate, hexametaphosphate, oxadiphosphonic acid, or aminotrimethylenephosphonic acid, can be used. More preferably, phytic acid and its salts are used to obtain stronger multi-point chelating ability and a more easily formed dense coordination network interface structure. The above substitutions do not change the core mechanism of the chelating agent adsorption-calcium / strontium ion coordination alternating network formation. However, different chelating agents have different acid strengths and complexation constants. It is preferable to adjust the pH of the working solution and the adsorption time to achieve similar network layer density and washout resistance under the same number of cycles.

[0048] In the strontium-doped hydroxyapatite nanoparticle immobilization + biomimetic mineralization step provided by the present invention, the strontium-doped hydroxyapatite nanoparticles preferably have a strontium doping ratio of 1%-10%, more preferably 3%-7%; the average particle size is preferably 10nm-100nm, more preferably 20nm-50nm.

[0049] The concentration of nanoparticles in the particulate dispersion is preferably 0.2 mg / mL-5 mg / mL, more preferably 0.5 mg / mL-2 mg / mL; the ultrasonic dispersion power is preferably 50 W-400 W, more preferably 100 W-250 W; and the ultrasonic time is preferably 5 min-30 min, more preferably 8 min-15 min.

[0050] The dispersion conditions affect the degree of particle aggregation, which in turn affects the uniformity of particle immobilization and the density of nucleation sites on the coordination network layer. The preferred oscillation / stirring speed during immobilization is 80 r / min-200 r / min, more preferably 100 r / min-150 r / min, and the preferred immobilization time is 10 min-1 h 30 min, more preferably 20 min-45 min.

[0051] In terms of biomimetic mineralization, modified simulated body fluid is preferred, with a concentration preferably 1-2 times, more preferably 1.2-1.8 times, a reaction temperature preferably 35℃-39℃, more preferably 37℃, and a reaction time preferably 4h-24h, more preferably 6h-12h.

[0052] The simulated body fluid concentration, temperature, and time affect the deposition rate, crystal morphology, and coverage of apatite, which in turn affect the stability of the micro-nano composite active interface and the inorganic phase signal required for bone-related cell adhesion / differentiation. Strontium doping can also provide an ionic stimulation pathway at the interface to promote osteogenic differentiation, thereby helping to improve the early bone contact formation rate and stability.

[0053] For sterilization, ethylene oxide sterilization is preferred, along with thorough desorption to reduce residues; gamma ray or electron beam sterilization can also be used as alternatives, but excessively high temperature and humidity conditions that would significantly alter the structure of the surface organic layer and the stability of the coordination network should be avoided.

[0054] The following detailed description is provided with reference to the embodiments, but these should not be construed as limiting the scope of protection of the present invention.

[0055] Example 1: The zirconia matrix was yttrium-stabilized tetragonal polycrystalline zirconia ceramic (3Y-TZP, containing 3 mol% Y2O3); the branched polyethyleneimine was Sigma-Aldrich branched polyethyleneimine, catalog number 408727, with an average Mw of approximately 25,000; the strontium-doped hydroxyapatite nanoparticles were strontium-doped hydroxyapatite from Jiangsu Xianfeng Nanomaterials Technology Co., Ltd., catalog number 104241, with a specification of 5% Sr and an average particle size of 30 nm; the simulated body fluid was modified SBF simulated body fluid from Beijing Solarbio Technology Co., Ltd., 1.5×, sterile, brand Solarbio, catalog number G0392.

[0056] Step S1: Take 10 zirconia substrates (each Φ10mm×2mm), and sonicate them sequentially in 200mL of acetone for 10min, then in 200mL of anhydrous ethanol for 10min, and finally in 200mL of deionized water for 10min. After removal, rinse twice with 200mL of deionized water and air dry in a clean environment for 30min. Then immerse them in 200mL of 2wt% sodium hydroxide aqueous solution, stand at a constant temperature of 60℃ for 30min, remove them, rinse three times with deionized water and air dry for 30min to obtain pretreated zirconia substrates. Step S2: Weigh 121 mg of tris(hydroxymethyl)aminomethane and dissolve it in 100 mL of deionized water. Adjust the pH to 8.5 by titration with 1 mol / L hydrochloric acid solution. Then weigh 200 mg of dopamine hydrochloride and add it to the solution, stirring and dissolving under air to obtain a dopamine self-polymerization reaction solution. Place 10 pretreated zirconia substrates into the dopamine self-polymerization reaction solution and react at 25 °C and 200 rpm for 6 hours. After that, remove the substrates, rinse them three times with deionized water, and air dry them for 30 minutes to obtain a polydopamine-containing adhesive underlayer. Zirconia matrix; 121 mg of tris(hydroxymethyl)aminomethane was dissolved in 100 mL of deionized water, and the pH was adjusted to 8.5 by titration with 1 mol / L hydrochloric acid solution. Then, 100 mg of branched polyethyleneimine was added to prepare polyethyleneimine working solution. Ten zirconia matrices with polydopamine-adhesive bottom layers were immersed in the working solution and reacted at 25 °C and 100 rpm for 1 h. After being removed, they were rinsed three times with deionized water and air-dried for 30 min to obtain zirconia matrix containing polyamine groups. Step S3: Weigh 100 mg of 50 wt% phytic acid aqueous solution and add it to 100 mL of deionized water. Mix well and titrate with 1 mol / L sodium hydroxide solution to adjust the pH to 7 to obtain the phytic acid working solution. Then weigh 661 mg of calcium chloride dihydrate and 133 mg of strontium chloride hexahydrate and add them to 100 mL of deionized water to dissolve, obtaining the calcium / strontium ion working solution. Place 10 zirconia matrices containing polyamine groups in the phytic acid working solution and let them stand at 25°C for 10 min. After removing them, rinse them once with deionized water. Then place them in the calcium / strontium ion working solution and let them stand at 25°C for 2 min. After removing them, rinse them once with deionized water. Repeat the above phytic acid adsorption-calcium / strontium coordination process 3 times. Finally, rinse them 3 times with deionized water and air dry for 30 min to obtain zirconia matrices containing coordination networks. Step S4: Weigh 100 mg of strontium-doped hydroxyapatite nanoparticles and add them to 100 mL of deionized water. Disperse the nanoparticles under ultrasonic conditions of 200 W for 10 min to obtain a nanoparticle dispersion. Immerse 10 zirconia matrices containing coordination networks in the dispersion and shake at 25 °C and 120 rpm for 30 min. Remove the matrices and rinse them once with deionized water to remove weakly adsorbed particles. Then, immerse them in 500 mL of simulated body fluid at 37 °C for 8 h. After rinsing them three times with deionized water and air-drying them for 12 h, sterilize them with ethylene oxide at 50 °C for 7 days to obtain zirconia bone-integrated implant material.

[0057] Example 2: Compared with Example 1: In step S2, the dosage of dopamine hydrochloride was 250 mg and the dosage of tris(hydroxymethyl)aminomethane was 50 mg; the other conditions were the same as in Example 1.

[0058] Example 3: Compared with Example 1: the phytic acid adsorption-calcium / strontium coordination process in step S3 was repeated twice; the other conditions were the same as in Example 1.

[0059] Example 4: Compared with Example 1: the amount of strontium-doped hydroxyapatite nanoparticles used in step S4 was 50 mg; the other conditions were the same as in Example 1.

[0060] Example 5: Compared with Example 1: the amount of strontium-doped hydroxyapatite nanoparticles used in step S4 was 200 mg; the other conditions were the same as in Example 1.

[0061] Comparative Example 1: Compared with Example 1: In step S2, after completing the self-polymerization deposition and cleaning of dopamine hydrochloride, the sample is no longer placed in the branched polyethyleneimine aqueous solution for treatment, but directly proceeds to step S3; the remaining conditions are the same as in Example 1.

[0062] Comparative Example 2: Compared with Example 1, the number of repetitions of the alternating process of phytic acid adsorption-calcium / strontium ion coordination in step S3 was changed from 3 times to 1 time; the other conditions were the same as in Example 1.

[0063] Comparative Example 3: Compared with Example 1: the amount of strontium-doped hydroxyapatite nanoparticles used in step S4 was 0 mg; the other conditions were the same as in Example 1.

[0064] Comparative Example 4: Compared with Example 1: the biomimetic mineralization step in step S4 is cancelled; the remaining conditions are the same as in Example 1.

[0065] Comparative Example 5: Compared with Example 1: the amount of strontium chloride hexahydrate in the calcium / strontium ion working solution in step S3 was changed to 0 mg; the other conditions were the same as in Example 1.

[0066] Performance testing: Fourier transform infrared spectroscopy characterization: Fourier transform infrared spectroscopy was used to identify chemical groups on the surface of the samples from Example 1, Comparative Examples 1, 2, and 4. Air was used as the background before testing, and baseline correction was performed. The results are as follows: Figure 1 As shown.

[0067] Surface roughness: The arithmetic mean roughness Ra of the sample in the example and comparative examples was measured using a stylus profilometer. The stylus tip radius was set to 2 μm, the sampling length was set to 0.8 mm, the evaluation length was set to 4.0 mm, and the scanning speed was set to 0.5 mm / s. Each sample was measured once in two mutually perpendicular directions and the average value was taken. Five samples were taken from each group and the average value within the group was taken.

[0068] Apatite-forming capacity in simulated body fluids: According to YY / T 1447-2016, the sample surface area to solution volume ratio is 0.088 cm². 2 The immersion evaluation was performed using 25.0 mL of simulated body fluid per tablet. The samples from the examples and comparative examples were placed in a 37°C constant-temperature shaker at 100 rpm for 7 days, with the simulated body fluid replaced every 24 hours. After immersion, the samples were rinsed three times with deionized water and dried at 37°C for 12 hours. The mass gain (mg / cm³) was calculated. 2 ).

[0069] Calcium / Strontium ion release: The samples from the examples and comparative examples were placed in a 37°C constant-temperature shaker for release testing, with the shaking speed set to 100 rpm. 10.0 mL of deionized water was added to each sample as the release medium, and the samples were sealed and protected from light. Sampling was conducted on days 1, 3, 7, and 14. At each time point, 2.00 mL of supernatant was collected for testing, and 2.00 mL of fresh deionized water was added to maintain a constant volume. Strontium ion concentration was determined by flame atomic absorption spectrometry. Spectrophotometric determination was performed using a wavelength of 460.7 nm, a lamp current of 5.0 mA, a passband width of 0.5 nm, a burner height of 8.5 mm, and an air-acetylene flame with oxidizing lean combustion. The mass concentration was calculated using the standard curve method. Calcium ion concentration was determined by disodium ethylenediaminetetraacetate complexometric titration, with the concentration of the disodium ethylenediaminetetraacetate standard solution set at 0.0100 mol / L. The cumulative release over 14 days was calculated, and the release curve is shown below. Figure 2 and 3 As shown.

[0070] In vitro cytotoxicity (extraction method): Following GB / T 16886.5-2017, the cytotoxicity of the examples and comparative samples was evaluated using the extraction method. For each group, 10 samples sterilized with ethylene oxide and analyzed for 7 days were used, and 7.33 mL of complete culture medium was added to prepare the extraction solution. The extraction conditions were set at 37℃ for 72 h. After extraction, the extract was filtered through a 0.22 μm filter membrane. Mouse fibroblasts were used for detection, with a cell seeding density of 1.0 × 10⁶ cells / year. 4 Cells per well (96-well plate, 100 μL per well) were cultured for 24 h, then replaced with the corresponding sample extract and cultured for another 24 h and 72 h. Cell viability was determined using the methylthiazolyltetrazole method, with the working solution concentration set at 0.5 mg / mL and the incubation time set at 4 h. Subsequently, dimethyl sulfoxide was added to dissolve formazan, and the absorbance was measured at 570 nm to calculate the relative cell viability.

[0071] Local reaction after implantation: The in vivo local tissue reaction of the sample from the example and the comparative example was evaluated according to GB / T 16886.6-2022. Eight-week-old male Sprague-Dawley rats with a body weight of 250g were selected, and the number of animals in each group was set to 6. Under aseptic conditions, bilateral muscle pouches were established in the back muscles and one sample was implanted in each. The tissue was harvested 12 weeks after the operation. The tissue around the implant was paraffin-embedded, sectioned, and stained with hematoxylin and eosin. The inflammatory cell infiltration, fibrous capsule thickness and necrosis were quantitatively scored.

[0072] The key data of the above test results are shown in Table 1.

[0073] Table 1 Performance Test Results

[0074] Data Analysis: From Table 1 and Figure 2 , 3Data from the embodiments show that the zirconia bone-integrated implant material prepared in this invention exhibits a synergistic improvement in surface roughness, apatite-like formation ability in simulated body fluids, and sustained release of calcium and strontium ions. Simultaneously, relative cell viability remains at a high level, and the local reaction score after implantation is low. This trend indicates that the polydopamine adhesion layer provides a stable interfacial binding basis for the inorganic phase. The polyamine sites introduced by branched polyethyleneimine enhance the adsorption coverage and multi-point coordination efficiency of phytic acid molecules, enabling calcium / strontium ions to construct a denser coordination network layer on the surface and form ion reserves. Based on this, the strontium-doped hydroxyapatite nanoparticles further provide high-density nucleation sites, promoting more continuous and uniform biomimetic mineralization deposition, thereby jointly constructing a stable calcium-phosphorus phase active interface at the micro-nano scale. This active interface enhances the apatite-like deposition tendency in simulated body fluids and provides a continuous inorganic signal at the interface through the slow release of calcium and strontium ions, ultimately balancing interfacial activity, controllable release, and biocompatibility.

[0075] From Table 1 and Figure 2 , 3 Data from Example 1 and Comparative Examples 1 and 2 show that when branched polyethyleneimine is not introduced or the number of repetitions of the alternating process of phytic acid adsorption and calcium / strontium ion coordination is significantly reduced, the surface roughness of the material and its ability to form apatite-like structures in simulated body fluids both decrease. The cumulative release of calcium and strontium ions also decreases simultaneously, and the local reaction score increases after implantation. The main reason is that the high-density amine groups and positively charged sites provided by branched polyethyleneimine are key prerequisites for stable phytic acid adsorption and multi-point coordination network construction. Insufficient cycle counts limit the thickness and ion reserve of the coordination network layer, making it difficult to establish the uniformity of subsequent strontium-doped hydroxyapatite nanoparticle immobilization and the density of nucleation sites. This demonstrates that the multi-amine group enhancement + multiple coordination network construction is not a simple superposition, but rather jointly determines the network density and ion pool capacity at the interface, thereby amplifying the biomimetic mineralization effect and exhibiting a synergistic gain that is difficult to obtain from a single modification.

[0076] From Table 1 and Figure 2 , 3Data from Example 1 and Comparative Example 3 show that when the amount of strontium-doped hydroxyapatite nanoparticles is zero, the apatite-like formation ability and calcium-phosphorus phase-related release behavior in simulated body fluids are significantly weakened, and the surface roughness and biocompatibility-related indicators also deteriorate. The main reason is that although the coordination network layer can introduce calcium / strontium ions and provide a certain nucleation basis, without the "high specific surface area nucleation template" of nanoparticles, biomimetic mineralization deposition relies more on random nucleation and local enrichment, resulting in discontinuous deposition coverage and insufficient nucleation site density, making it difficult to form a stable micro / nano composite active interface. These results indicate that nanoparticle immobilization not only increases roughness, but more importantly, together with the coordination network layer, it constitutes a controllable nucleation-continuous deposition path, enabling the overall interface activity to exhibit an improvement exceeding that of a single network layer, demonstrating a synergistic effect greater than the sum of its parts.

[0077] From Table 1 and Figure 2 , 3 Data from Example 1 and Comparative Example 4 show that when the biomimetic mineralization step is omitted, the material's ability to form apatite-like structures in simulated body fluid decreases, and the release behavior of calcium and strontium ions exhibits a rapid initial release followed by insufficient accumulation. Simultaneously, cell viability and post-implantation local response scores deteriorate. The main reason for this is that the calcium-phosphorus phase capping layer formed by biomimetic mineralization not only provides the osteogenic inorganic phase but also isolates and stabilizes the polyamine basal layer and coordination network layer, reducing the extractable components and regulating ion release into a more gradual, sustained release process. When this capping layer is missing, the nanoparticles and coordination network are more directly exposed to the release medium, leading to an early ion burst release. The lack of a continuous mineralization layer also weakens the interfacial basis for further deposition in simulated body fluid. Therefore, nanoparticle immobilization and biomimetic mineralization deposition have a synergistic effect in regulating release and stabilizing the interface.

[0078] From Table 1 and Figure 2 , 3 Data from Example 1 and Comparative Example 5 show that when strontium chloride hexahydrate is not introduced into the calcium / strontium ion working solution, the cumulative release of strontium ions is significantly reduced, and the ability to form apatite-like structures in the simulated body fluid and the local reaction score after implantation also show adverse changes. The main reason is that, in addition to contributions from strontium-doped hydroxyapatite nanoparticles, the more important source of strontium ions is the calcium / strontium ion reserve in the coordination network layer. This reserve can provide a more durable strontium ion release channel at the interface and influence the nucleation and crystal growth process of the calcium-phosphorus phase during biomimetic mineralization. When the network layer lacks strontium ions, even with strontium-doped nanoparticles, the overall strontium ion release and interface regulation capabilities are still limited, making it difficult to form a complete closed loop of network layer ion pool + nanoparticle nucleation template + mineralized deposition capping layer, thus demonstrating the crucial synergistic role of strontium ions in the multilayer structure.

[0079] from Figure 1 It can be seen that the sample in Example 1 is within the range of 1100-1000 cm. -1 PO4 in the interval 3- v3 characteristic peak (approximately 1090 cm⁻¹) -1 and 1030cm -1 ) and 962cm -1 The intensity of the symmetric stretching vibration peak at v1 is significantly higher than that of Comparative Examples 1, 2, and 4, and is also higher at 602 cm⁻¹. -1 With 563cm -1 PO4 3- The v4 bending vibration peak is also sharper, indicating that its surface is enriched with more calcium phosphate phase; combined with the 7-day simulated body fluid immersion mass increase and Ca 2+ / Sr 2+ The cumulative release was the highest among all groups, suggesting that the dense ionic network layer constructed by the polydopamine adhesion substrate, the branched polyethyleneimine polyamine base layer, and the multiple phytic acid-calcium / strontium coordination steps, along with the biomimetic mineralization steps, synergistically promoted the formation of bone-like apatite. Although biomimetic mineralization was omitted in Comparative Example 4, it still exhibited stronger phosphate characteristic peaks than Comparative Examples 1 and 2, indicating that the pre-constructed coordination network layer and the immobilized strontium-doped hydroxyapatite nanoparticles still possessed certain nucleation and mineralization capabilities. Comparative Example 1, due to the absence of a polyamine base layer, and Comparative Example 2, due to the reduced number of phytic acid-calcium / strontium coordination cycles, showed higher P=O and PO4 content. 3- The relevant peaks all weakened significantly, only reaching 1030 cm⁻¹. -1 The presence of a wide and low shoulder peak nearby proves that the polyamine base layer and the multiple coordination network are key factors in obtaining a stable micro-nano composite active interface.

[0080] Those skilled in the art should understand that the discussion of any of the above embodiments is merely exemplary and is not intended to imply that the scope of the invention is limited to these examples; within the framework of the invention, the technical features of the above embodiments or different embodiments can also be combined, the steps can be implemented in any order, and there are many other variations of the different aspects of the invention as described above, which are not provided in detail for the sake of brevity.

Claims

1. A method for preparing a zirconia bone-integrated implant material, characterized in that, Includes the following steps: (1) The zirconia matrix is ​​cleaned and alkaline activated to obtain an activated zirconia matrix; (2) The activated zirconia matrix is ​​placed in a hydrochloric acid dopamine solution in a weakly alkaline buffer system to allow the hydrochloric acid dopamine to self-polymerize and deposit, forming a polydopamine adhesion layer on the surface of the activated zirconia matrix. (3) To fix the branched polyethyleneimine on the polydopamine adhesive substrate; (4) The zirconia matrix treated in step (3) is alternately treated with phytic acid working solution and working solution containing calcium ions and strontium ions, and this process is repeated two to four times, so that phytic acid molecules coordinate and crosslink with calcium ions and strontium ions to form a coordination network layer containing calcium / strontium ions. (5) The zirconium oxide matrix treated in step (4) is brought into contact with the dispersion of strontium-doped hydroxyapatite nanoparticles, so that the strontium-doped hydroxyapatite nanoparticles are immobilized on the surface of the coordination network layer. (6) The zirconia matrix treated in step (5) is placed in a simulated body fluid for biomimetic mineralization deposition, then washed and dried to obtain zirconia bone integration implant material; Based on 10 zirconia substrates with a size of Φ10mm×2mm, the calcium ion concentration in step (4) is 30mmol / L~60mmol / L, the strontium ion concentration is 3mmol / L~10mmol / L, the molar ratio of calcium ion to strontium ion is 5:1~15:1, and the calcium / strontium ion coordination time is 1min~3min; the concentration of nanoparticles in the strontium-doped hydroxyapatite nanoparticle dispersion in step (5) is 0.5mg / mL~2mg / mL.

2. The method for preparing the zirconia bone-integrated implant material according to claim 1, characterized in that, The zirconia matrix is ​​a tetragonal polycrystalline zirconia ceramic matrix containing yttrium oxide stabilizer.

3. The method for preparing the zirconia bone-integrated implant material according to claim 1, characterized in that, The weakly alkaline buffer system in step (2) is a tris(hydroxymethyl)aminomethane buffer system, wherein the concentration of tris(hydroxymethyl)aminomethane is 8 mmol / L to 12 mmol / L and the pH of the solution is 8.3 to 8.

7.

4. The method for preparing the zirconia bone-integrated implant material according to claim 1, characterized in that, Based on 10 zirconia substrates with a size of Φ10mm×2mm, the concentration of dopamine hydrochloride in step (2) is 1mg / mL~3mg / mL, the reaction temperature is 23℃~27℃, the reaction time is 4h~8h, and the stirring speed is 150r / min~220r / min.

5. The method for preparing the zirconia bone-integrated implant material according to claim 1, characterized in that, Based on 10 zirconia substrates with a size of Φ10mm×2mm, the weight-average molecular weight of the branched polyethyleneimine in step (3) is 20,000 to 30,000, the working solution concentration is 0.5mg / mL to 1.5mg / mL, the reaction temperature is 20℃ to 30℃, the reaction time is 45min to 1h30min, and the stirring speed is 80r / min to 120r / min.

6. The method for preparing the zirconia bone-integrated implant material according to claim 1, characterized in that, Based on 10 zirconium oxide substrates with a size of Φ10mm×2mm, the pH of the phytic acid working solution in step (4) is 6.8~7.2, the phytic acid concentration is 0.3mg / mL~0.8mg / mL, and the phytic acid adsorption time is 8min~15min.

7. The method for preparing the zirconia bone-integrated implant material according to claim 1, characterized in that, In step (4), the working solution containing calcium and strontium ions contains calcium salts selected from at least one of calcium chloride dihydrate, calcium nitrate tetrahydrate, and calcium acetate, and strontium salts selected from at least one of strontium chloride hexahydrate and strontium nitrate.

8. The method for preparing the zirconia bone-integrated implant material according to claim 1, characterized in that, The strontium doping ratio of the strontium-doped hydroxyapatite nanoparticles in step (5) is 3% to 7%, and the average particle size is 20 nm to 50 nm.

9. The method for preparing the zirconia bone-integrated implant material according to claim 1, characterized in that, The simulated body fluid in step (6) is from Solarbio, catalog number G0392. The biomimetic mineralization reaction temperature is 37℃ and the biomimetic mineralization reaction time is 6h to 12h.

10. A zirconia bone-integrated implant material, characterized in that, It is obtained by the preparation method of the zirconia bone integration implant material according to any one of claims 1-9.

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