A method for preparing a near-beta tlm titanium alloy dealloyed / chronological function surface

Abstract

The application discloses a preparation method of a near-beta TLM titanium alloy dealloying / chronological functional surface, and the method comprises the following steps: 1, heat treatment to obtain a dual-phase microstructure TLM titanium alloy substrate; 2, electrochemical dealloying to obtain a hierarchical micro-nano porous surface structure; 3, preparation of a complex solution; 4, chemical deposition of the complex solution to construct an osteogenesis and vascularization functional layer on the hierarchical micro-nano porous surface structure; and 5, electrostatic adsorption to construct an antibacterial functional layer. The osteogenesis and vascularization functional layer and the antibacterial functional layer are introduced on the dealloyed surface of the TLM titanium alloy, the surface is endowed with the chronological expression characteristics through controllable degradation release, the positive electricity and the porous structure of polylysine are used to realize synergistic antibiosis, the tannic acid induced strontium ion controllable release improves the osteogenesis and vascularization performance, the hierarchical micro-nano porous structure of the surface is beneficial to the formation of good bone bonding, the TLM titanium alloy is endowed with early antibiosis, medium and long-term osteogenesis promotion and excellent bone integration characteristics, and is used for bone defect repair.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of medical titanium and titanium alloy materials, and particularly relates to a preparation method of a near-beta TLM titanium alloy de-alloying / chronological functional surface. BACKGROUND

[0002] Titanium and titanium alloy have irreplaceable advantages in the field of bone defect repair due to their excellent biocompatibility, corrosion resistance and processing performance. However, the results of clinical follow-up studies on titanium alloy show that the research on titanium alloy for bone defect repair still needs to consider the problems of insufficient biomechanical compatibility, bioactivity and antibacterial performance, and titanium alloy with good biomechanical adaptability and biological functionality needs to be developed.

[0003] Ti-3Zr-2Sn-3Mo-25Nb (TLM) titanium alloy is a high-strength low-modulus metastable beta-type medical titanium alloy, which has excellent and controllable comprehensive mechanical properties. Its elastic modulus is about 40 GPa to 80 GPa, which is close to the elastic modulus of bone tissue, and has good biomechanical adaptability. At the same time, the surface micro-nano structure improves the bioactivity of titanium alloy by regulating the adhesion behavior of cells on the surface, and enhances the bone integration between the implant and the newly formed bone tissue. Electrochemical de-alloying technology is a surface modification technology for preparing hierarchical micro-nano porous structure by corroding active elements in the matrix. Compared with traditional surface modification technologies, electrochemical de-alloying technology has the advantages of in-situ preparation, one-step preparation and controllable preparation, which avoids the problems of coating peeling and healing complication caused by the interface, and has wide application prospects in the preparation of porous surface of biomedical metal materials. Although electrochemical de-alloying improves the bioactivity while retaining the good biomechanical adaptability of the matrix, it still lacks the necessary early antibacterial and medium-long-term osteogenic differentiation and vascularization performance during the bone healing process, and the de-alloyed surface needs to be functionalized to induce the chronological expression of biological functional properties.

[0004] Metallic strontium ions can promote bone formation and bone mineralization, enhance bone strength, promote osteoblast differentiation and inhibit osteoclast generation, and promote angiogenesis by enhancing the expression of vascular growth factors. Therefore, by controlling the release amount of strontium ions, the sustained expression of strontium ion biological functionality can be achieved. Tannic acid is a naturally occurring polyphenolic compound with free radical scavenging, anti-inflammatory and antioxidant properties. It can form stable complexes with metal ions and has a pH-responsive release mechanism, thereby controlling the release amount of metal ions. After tannic acid is complexed with metal ions strontium, the release behavior of strontium ions is controlled to endow the matrix material with biological functionalities such as oxidation resistance, osteogenic differentiation and angiogenesis promotion.

[0005] Polylysine is a water-soluble lysine polymer, a natural antibacterial peptide secreted by Streptomyces, which has broad-spectrum antibacterial properties and good inhibitory effect on gram-positive and negative bacteria, yeast and mold. Unlike the drug resistance caused by antibiotic antibacterial and metal toxicity caused by metal ion antibacterial, polylysine causes irreversible damage to bacterial membrane, thereby causing bacterial death, which does not produce drug resistance and can be decomposed into essential lysine in the human body. Polylysine is expected to be widely used in antibacterial modification of bone defect repair implant. SUMMARY

[0006] The technical problem to be solved by the present application is to provide a preparation method of dealloyed / sequential function surface of near-beta TLM titanium alloy, which solves the problems of lack of early antibacterial and long-term osteogenic differentiation and vascularization performance in the process of bone tissue healing of the existing TLM titanium alloy.

[0007] To solve the above technical problems, the technical scheme adopted by the present application is as follows: a preparation method of dealloyed / sequential function surface of near-beta TLM titanium alloy, characterized in that the method comprises the following steps:

[0008] Step one, heat treating the near-beta TLM titanium alloy to obtain a TLM titanium alloy substrate with a dual-phase microstructure;

[0009] Step two, performing surface electrochemical dealloying treatment on the TLM titanium alloy substrate with a dual-phase microstructure obtained in step one, using the TLM titanium alloy substrate as an anode, a platinum electrode as a cathode, an acid electrolyte and a constant voltage higher than the critical potential to obtain a TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure;

[0010] Step three, mixing tannic acid and strontium salt and then performing complexation reaction to obtain a complex solution;

[0011] Step four, performing chemical deposition treatment on the TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure obtained in step two by using the complex solution obtained in step three to obtain a TLM titanium alloy with an osteogenic and vascularization function layer;

[0012] Step five, constructing an antibacterial function layer on the TLM titanium alloy with an osteogenic and vascularization function layer obtained in step four by electrostatic adsorption to obtain a dealloyed / sequential function surface.

[0013] The present application firstly carries out heat treatment on the near-beta TLM titanium alloy, preferably solid solution aging heat treatment in a vacuum environment, adjusts the microstructure of the near-beta TLM titanium alloy to be a dual-phase microstructure, and uses the sequential dealloying characteristics of the dual-phase microstructure to lay a foundation for forming a hierarchical micro-nano porous surface structure through subsequent surface electrochemical dealloying treatment. Secondly, the present application carries out surface electrochemical dealloying treatment by using the TLM titanium alloy substrate as an anode, using a platinum electrode as a cathode, using an acid electrolyte, and applying a constant voltage higher than a critical potential. During the treatment process, the alpha phase in the dual-phase microstructure of the TLM titanium alloy substrate is preferentially corroded due to its low self-corrosion potential, forming micron-sized grooves, and acid ions such as fluoride ions in the acid electrolyte form nanometer-sized pores by breaking through the TLM titanium alloy surface oxide film, thereby forming a hierarchical micro-nano porous surface structure. The hierarchical micro-nano porous surface structure regulates the surface roughness and hydrophilicity of the TLM titanium alloy substrate, is conducive to the adhesion of osteogenic-related cells, and the micro-nano pores are conducive to the mechanical "riveting" between the implant and the newly formed bone tissue, achieving good bone integration. Then, the present application constructs a tannic acid / strontium complex functional layer (TA / Sr) on the hierarchical micro-nano porous surface structure of the TLM titanium alloy substrate through complexation reaction and chemical deposition treatment, uses the pH-responsive release mechanism of the complex of tannic acid (TA) and metal ion strontium to realize the controlled release of metal active ion strontium ions, and then realizes the sustained expression of the biological functionality of strontium ions, improving the biological functionality of the TLM titanium alloy, such as promoting osteogenic differentiation, inhibiting osteoclastogenesis, antioxidant, and promoting angiogenesis. Finally, the present application uses the electrostatic adsorption method to construct a polylysine antibacterial functional layer (ε-PL) through the electrostatic adsorption between the negative units in tannic acid and the positive units in polylysine, uses the irreversible damage of polylysine to the bacterial membrane layer to realize good antibacterial performance, and the presence of the polylysine functional layer further inhibits the early burst release behavior of strontium ions, further promoting the sustained expression of the biological functionality of strontium ions.

[0014] Based on the time sequence requirements of early antibacterial, long-term bone differentiation promotion and finally good bone combination between the new bone tissue in the clinical application of orthopedic implant devices, the application first prepares a hierarchical micro-nano porous surface structure, then prepares a TA / Sr functional layer through chemical deposition, and finally prepares an epsilon-PL functional layer to obtain a dealloying / time sequence functional surface. In the human body physiological environment, the polylysine of the dealloying / time sequence functional surface causes irreversible damage to the bacterial membrane layer, thereby meeting the antibacterial performance; with the degradation of the polylysine, the tannic acid pH regulates the strontium ion release rate, so that the strontium ion is effectively released for a long time, the strontium ion promotes the osteogenic differentiation of osteoblasts and the vascularization of endothelial cells, which is beneficial to the formation of new bone tissue; finally, the hierarchical micro-nano porous structure of the surface forms mechanical riveting with the new bone, thereby enhancing the stability between the implant and the new bone, and the dealloying / time sequence functional surface is beneficial to realize rapid osseointegration.

[0015] Generally, the application preferably uses SrCl2·6H2O as a strontium salt, which will form a deposit impurity on the surface of the TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure compared to Sr(OH)2, and SrCl2·6H2O is non-toxic to the organism and no impurities are generated, thereby avoiding the generation of cytotoxicity and being suitable for use in the organism implant.

[0016] The preparation method of the near-beta type TLM titanium alloy dealloying / time sequence functional surface described above, characterized in that the heat treatment conditions in step one are as follows: in an environment with a vacuum degree of 1Pa-10Pa, the heating rate is 5℃ / min-10℃ / min, first heating to 760℃-790℃ for primary heat treatment for 1h, water cooling, then heating to 470℃-510℃ for secondary heat treatment for 2h, and air cooling. The application adjusts the microstructure of the TLM titanium alloy substrate to be a dual-phase microstructure through two different temperature heat treatment processes, combines with a proper vacuum environment to effectively avoid the formation of an oxide film during the heat treatment process, and precipitates fine and dispersed alpha phase inside the equiaxed crystal during the two heat treatment processes, thereby controlling the size and content of the precipitated alpha phase by adjusting the heat treatment temperature, time and cooling rate.

[0017] The preparation method of the near-beta TLM titanium alloy dealloying / chronological functional surface has the characteristics that, in the step two, the surface electrochemical dealloying treatment adopts a constant potential instrument to apply a voltage of 1V-5V, and the treatment time is 1h-3h; the acid electrolyte contains 0.2%-0.5% hydrofluoric acid and 2%-5% nitric acid in volume fraction. The voltage value greater than the critical potential is applied by the constant potential instrument to promote the surface electrochemical dealloying treatment process; by controlling the components of the acid electrolyte, the alpha phase in the dual-phase microstructure of the TLM titanium alloy is preferentially corroded due to the low self-corrosion potential, and micron-sized grooves are formed, and the fluorine ions in the acid electrolyte form uniform nanosize by breaking through the surface oxide film of the TLM titanium alloy, so that the surface electrochemical dealloying treatment process of the application can form a micro-nano surface structure by in-situ corrosion control, and has good bonding force with the TLM titanium alloy substrate, and the adhesion behavior of the osteogenic cells on the surface of the TLM titanium alloy material is improved.

[0018] The preparation method of the near-beta TLM titanium alloy dealloying / chronological functional surface has the characteristics that, in the step two, the surface electrochemical dealloying treatment adopts a constant potential instrument to apply a voltage of 1V-5V, and the treatment time is 1h-3h; the acid electrolyte contains 0.2%-0.5% hydrofluoric acid and 2%-5% nitric acid in volume fraction. The voltage value greater than the critical potential is applied by the constant potential instrument to promote the surface electrochemical dealloying treatment process; by controlling the components of the acid electrolyte, the alpha phase in the dual-phase microstructure of the TLM titanium alloy is preferentially corroded due to the low self-corrosion potential, and micron-sized grooves are formed, and the fluorine ions in the acid electrolyte form uniform nanosize by breaking through the surface oxide film of the TLM titanium alloy, so that the surface electrochemical dealloying treatment process of the application can form a micro-nano surface structure by in-situ corrosion control, and has good bonding force with the TLM titanium alloy substrate, and the adhesion behavior of the osteogenic cells on the surface of the TLM titanium alloy material is improved.

[0019] The preparation method of the near-beta TLM titanium alloy dealloying / chronological functional surface has the characteristics that, in the step two, the surface electrochemical dealloying treatment adopts a constant potential instrument to apply a voltage of 1V-5V, and the treatment time is 1h-3h; the acid electrolyte contains 0.2%-0.5% hydrofluoric acid and 2%-5% nitric acid in volume fraction. The voltage value greater than the critical potential is applied by the constant potential instrument to promote the surface electrochemical dealloying treatment process; by controlling the components of the acid electrolyte, the alpha phase in the dual-phase microstructure of the TLM titanium alloy is preferentially corroded due to the low self-corrosion potential, and micron-sized grooves are formed, and the fluorine ions in the acid electrolyte form uniform nanosize by breaking through the surface oxide film of the TLM titanium alloy, so that the surface electrochemical dealloying treatment process of the application can form a micro-nano surface structure by in-situ corrosion control, and has good bonding force with the TLM titanium alloy substrate, and the adhesion behavior of the osteogenic cells on the surface of the TLM titanium alloy material is improved.

[0020] The preparation method of the near-beta TLM titanium alloy dealloying / chronological functional surface has the characteristics that, in the step two, the surface electrochemical dealloying treatment adopts a constant potential instrument to apply a voltage of 1V-5V, and the treatment time is 1h-3h; the acid electrolyte contains 0.2%-0.5% hydrofluoric acid and 2%-5% nitric acid in volume fraction. The voltage value greater than the critical potential is applied by the constant potential instrument to promote the surface electrochemical dealloying treatment process; by controlling the components of the acid electrolyte, the alpha phase in the dual-phase microstructure of the TLM titanium alloy is preferentially corroded due to the low self-corrosion potential, and micron-sized grooves are formed, and the fluorine ions in the acid electrolyte form uniform nanosize by breaking through the surface oxide film of the TLM titanium alloy, so that the surface electrochemical dealloying treatment process of the application can form a micro-nano surface structure by in-situ corrosion control, and has good bonding force with the TLM titanium alloy substrate, and the adhesion behavior of the osteogenic cells on the surface of the TLM titanium alloy material is improved.

[0021] The preparation method of the near-beta TLM titanium alloy dealloying / chronological functional surface has the characteristics that the process of the electrostatic adsorption in the fifth step is: polylysine powder is dissolved in a hydroxyethylpiperazine ethylthiourea acid buffer solution with a concentration of 25 mM-50 mM to obtain an electrostatic adsorption solution with a mass concentration of 0.5 mg / mL-5 mg / mL, and then 50 muL-200 muL of the mixed solution is point-coated on the TLM titanium alloy surface with the osteogenesis and vascularization functional layer.

[0022] Compared with the prior art, the present application has the following advantages:

[0023] 1. The present application combines the surface hierarchical micro-nano porous structure, tannic acid / strontium osteogenesis and angiogenesis performance and polylysine antibacterial performance by constructing the chronological functional layer on the dealloyed surface, promotes cell adhesion through the hierarchical micro-nano porous structure of the TLM titanium alloy substrate material surface to improve the mechanical stability between the implant and the newly formed bone tissue, controls the strontium ion release behavior through tannic acid to endow the substrate material with good osteogenic differentiation performance and angiogenesis ability, and endows the substrate with excellent antibacterial performance through the irreversible damage of polylysine to the bacterial membrane, and the synergistic effect of the three endows the TLM titanium alloy with antibacterial / osteogenesis performance with chronological expression characteristics and is beneficial to the formation of early bone integration, which is suitable for the field of bone defect repair.

[0024] 2. The present application uses electrochemical dealloying method combined with chemical deposition and electrostatic adsorption to construct the osteogenesis and vascularization functional layer and the antibacterial functional layer from top to bottom on the TLM titanium alloy with hierarchical micro-nano porous surface after dealloying, in a physiological environment, the antibacterial functional layer on the surface first plays an antibacterial role, and with the degradation of polylysine in the antibacterial functional layer, the strontium ions in the intermediate layer, i.e. the osteogenesis and vascularization functional layer, are released slowly and controllably, the strontium ions promote angiogenesis, which is beneficial to the exchange of nutrients, promote in vitro osteogenic differentiation performance by up-regulating the expression of osteogenesis-related genes and proteins, and is beneficial to the formation of new bone tissue, and finally the new bone tissue interacts with the dealloyed micro-nano porous surface to realize good bone integration through mechanical "riveting", so that the dealloyed / chronological functional surface of the present application has the chronological functions of early antibacterial and medium-long term osteogenesis.

[0025] 3、The polylysine in the antibacterial functional layer in the time sequence dealloying / time sequence functional surface designed by the application degrades into lysine necessary for the human body in a physiological environment, and the tannic acid in the bone and vascularization functional layer controls the slow release of strontium ions, avoids the cytotoxicity caused by the burst release of metal ions, and in-situ corrosion forms a hierarchical micro-nano porous surface structure through electrochemical dealloying treatment, the structure has good bonding force with the TLM titanium alloy substrate, avoids the generation of metal abrasion caused by the peeling of an additional coating, and ensures that the dealloying / time sequence functional surface has biological functionality and biological safety.

[0026] 4、The preparation process of the application is easy to operate, is not limited by the structure and shape of the substrate material, can be used for uniform modification of the surface of personalized medical devices, and has good application prospects.

[0027] The technical solutions of the application are further described in detail below with reference to the drawings and examples. BRIEF DESCRIPTION OF DRAWINGS

[0028] Figure 1 The SEM image of the dealloying / time sequence functional surface prepared for the example 3 of the application.

[0029] Figure 2 The SEM images of the dealloying / time sequence functional surface prepared for the example 3 of the application and the adhesion of Staphylococcus aureus and Escherichia coli to the TLM titanium alloy.

[0030] Figure 3 The rat bone marrow mesenchymal stem cell alkaline phosphatase (ALP) staining image of the dealloying / time sequence functional surface prepared for the example 4 of the application and the TLM titanium alloy. DETAILED DESCRIPTION

[0031] Example 1

[0032] This example includes the following steps:

[0033] Step one, heat treating the near-beta type TLM titanium alloy to obtain a TLM titanium alloy substrate with a dual-phase microstructure; the heat treatment conditions are as follows: in an environment with a vacuum degree of 1 Pa, the temperature rising speed is 5 ℃ / min, first heat treating at 760 ℃ for 1 h, water cooling, and then heat treating at 470 ℃ for 2 h, air cooling;

[0034] Step two, performing surface electrochemical dealloying treatment on the TLM titanium alloy substrate with a dual-phase microstructure obtained in step one, taking the TLM titanium alloy substrate as an anode, taking a platinum electrode as a cathode, using an acidic electrolyte and applying a voltage of 1 V, and the treatment time is 1 h, to obtain a TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure; the acidic electrolyte contains 0.2% of hydrofluoric acid by volume fraction and 2% of nitric acid by volume fraction.

[0035] Step three, mixing tannic acid TA with strontium salt SrCl2·6H2O, the concentration of tannic acid in the mixed solution system is 1 mg / mL, and the concentration of strontium salt is 0.1 M, and the mixed solution system is placed in a vortex oscillator for 15 min to mix and shake until the solution is light yellow for complexation reaction, to obtain a complex solution;

[0036] Step four, immersing the TLM titanium alloy substrate with hierarchical micro-nano porous surface structure obtained in step two in 1 mL of the complex solution obtained in step three, controlling the environmental temperature to be 25℃ for chemical deposition treatment for 1 h, and cleaning by ultrasonic wave to remove the complex compounds not firmly combined on the surface, to obtain a TLM titanium alloy with a function layer for promoting osteogenesis and vascularization;

[0037] Step five, constructing an antibacterial function layer on the TLM titanium alloy with the function layer for promoting osteogenesis and vascularization obtained in step four by electrostatic adsorption, to obtain a dealloying / chronofunctional surface; the process of electrostatic adsorption is: dissolving polylysine (ε-PL) powder in a hydroxyethylpiperazine ethanesulfonic acid (HEPES) buffer solution with a concentration of 25 mM to obtain an electrostatic adsorption solution with a mass concentration of 0.5 mg / mL, and then taking 50 μL of the mixed solution to be coated on the surface of the TLM titanium alloy with the function layer for promoting osteogenesis and vascularization.

[0038] It is detected that the micropore size of the hierarchical micro-nano porous surface structure of the dealloying / chronofunctional surface prepared on the TLM titanium alloy substrate in the embodiment is 0.58 μm, and the nanopore size is 12.6 nm, and the hierarchical micro-nano porous surface structure is beneficial to early cell adhesion.

[0039] Embodiment 2

[0040] The embodiment includes the following steps:

[0041] Step one, heat treating a near β type TLM titanium alloy to obtain a TLM titanium alloy substrate with a dual-phase microstructure; the heat treatment conditions are: in an environment with a vacuum degree of 1 Pa, the temperature rising speed is 5℃ / min, first heat treating at 760℃ for 1 h, water cooling, and then heat treating at 470℃ for 2 h, air cooling;

[0042] Step two, electrochemically dealloying the TLM titanium alloy substrate with a dual-phase microstructure obtained in step one to obtain a TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure; the TLM titanium alloy substrate is used as an anode, a platinum electrode is used as a cathode, an acid electrolyte is used, and a voltage of 1 V is applied, and the treatment time is 3 h; the acid electrolyte contains 0.2% of hydrofluoric acid and 2% of nitric acid by volume fraction;

[0043] Step three, mixing tannic acid TA and strontium salt SrCl2·6H2O, the concentration of tannic acid in the mixed solution system is 1 mg / mL, and the concentration of strontium salt is 0.1 M, and the mixed solution system is placed in a vortex oscillator for 15 min to mix uniformly until the solution is light yellow for complexation reaction to obtain a complex solution;

[0044] Step four, immersing the TLM titanium alloy substrate with hierarchical micro-nano porous surface structure obtained in step two in 1 mL of the complex solution obtained in step three, controlling the environmental temperature to be 25℃ for chemical deposition treatment for 3 h, and cleaning by ultrasonic wave to remove the complex compounds not firmly combined on the surface to obtain a TLM titanium alloy with a function layer for promoting osteogenesis and vascularization;

[0045] Step five, constructing an antibacterial function layer on the TLM titanium alloy with the function layer for promoting osteogenesis and vascularization obtained in step four by electrostatic adsorption to obtain a dealloying / chronofunctional surface; the process of electrostatic adsorption is as follows: dissolving polylysine (ε-PL) powder in a hydroxyethylpiperazine ethanesulfonic acid (HEPES) buffer solution with a concentration of 25 mM to obtain an electrostatic adsorption solution with a mass concentration of 0.5 mg / mL, and then taking 50 μL of the mixed solution to spot on the surface of the TLM titanium alloy with the function layer for promoting osteogenesis and vascularization.

[0046] It is detected that the micropore size of the hierarchical micro-nano porous surface structure of the dealloying / chronofunctional surface prepared on the TLM titanium alloy substrate in this embodiment is 1.47 μm, the nanopore size is 14.32 nm, and after immersion in the complex solution and chemical deposition, the deposited complex completely covers the hierarchical micro-nano porous surface structure, and the hierarchical micro-nano porous surface structure is beneficial to early cell adhesion.

[0047] Embodiment 3

[0048] This embodiment includes the following steps:

[0049] Step one, heat treating a near-β type TLM titanium alloy to obtain a TLM titanium alloy substrate with a dual-phase microstructure; the heat treatment conditions are as follows: in an environment with a vacuum degree of 5 Pa, the temperature rising speed is 10℃ / min, first heat treating at 790℃ for 1 h, water cooling, and then heat treating at 510℃ for 2 h, air cooling;

[0050] Step two, electrochemically dealloying the TLM titanium alloy substrate with a dual-phase microstructure obtained in step one to obtain a TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure; the TLM titanium alloy substrate is used as an anode, a platinum electrode is used as a cathode, an acid electrolyte is used, a voltage of 3 V is applied, and the treatment time is 1 h; the acid electrolyte contains 0.5% of hydrofluoric acid by volume fraction and 5% of nitric acid by volume fraction.

[0051] Step 3: Mix tannic acid (TA) with strontium salt (SrCl2·6H2O). The concentration of tannic acid in the mixed solution is 2 mg / mL, and the concentration of strontium salt is 0.2 M. Place the mixed solution in a vortex mixer and shake for 15 min until the solution turns pale yellow to carry out the complexation reaction and obtain the complexed solution.

[0052] Step 4: Immerse the TLM titanium alloy substrate with hierarchical micro-nano porous surface structure obtained in Step 2 in 1 mL of the complexing solution obtained in Step 3, control the ambient temperature at 37℃ for chemical deposition treatment for 1 h, and use ultrasonic cleaning to remove the poorly bonded complexes on the surface to obtain a TLM titanium alloy with a bone-promoting and vascularization functional layer.

[0053] Step 5: Construct an antibacterial functional layer on the TLM titanium alloy with bone-promoting and vascularization functional layer obtained in Step 4 through electrostatic adsorption, resulting in a dealloyed / time-sequential functional surface; The electrostatic adsorption process is as follows: dissolve polylysine (ε-PL) powder in a 50mM hydroxyethylpiperazine thiosulfate (HEPES) buffer solution to obtain an electrostatic adsorption solution with a mass concentration of 2mg / mL, and then take 100μL of the mixed solution and spot-apply it to the surface of the TLM titanium alloy with bone-promoting and vascularization functional layer.

[0054] Testing revealed that the micropore size of the dealloyed / time-sequential functional surface with hierarchical micro-nano porous surface structure prepared on the TLM titanium alloy substrate in this embodiment is 2.3 μm, and the nanopore size is 17.8 nm. This hierarchical micro-nano porous surface structure is beneficial for early cell adhesion.

[0055] Figure 1 SEM images of the dealloyed / time-sequential functional surface prepared in this embodiment, from Figure 1 It can be seen that the dealloyed / time-sequential functional surface still retains the micron-level pore structure after dealloying, which is beneficial to the osteogenic integration between the implant and the newly formed bone tissue.

[0056] Figure 2 Figure 1 shows SEM images of Staphylococcus aureus and Escherichia coli adhering to the de-alloyed / time-sequentially functionalized surfaces prepared in this embodiment. Figure 2 shows the SEM image of Staphylococcus aureus adhering to the surface of the control group TLM titanium alloy (polished with 2000# sandpaper only), Figure 3 shows the SEM image of Staphylococcus aureus adhering to the surface of the experimental group de-alloyed / time-sequentially functionalized surfaces, Figure 4 shows the SEM image of Escherichia coli adhering to the surface of the control group TLM titanium alloy (polished with 2000# sandpaper only), and Figure 5 shows the SEM image of Escherichia coli adhering to the surface of the experimental group de-alloyed / time-sequentially functionalized surfaces. Figure 2It can be seen that the de-alloying / timing functional surface significantly reduces the Staphylococcus aureus (gram-positive bacteria) and Escherichia coli (gram-negative bacteria) compared with the control group, showing excellent antibacterial performance, which shows that the polylysine in the antibacterial functional layer of the de-alloying / timing functional surface prepared in this embodiment has inhibitory effect on gram-positive bacteria and gram-negative bacteria.

[0057] Example 4

[0058] This embodiment includes the following steps:

[0059] Step one, heat treatment is performed on the near-beta TLM titanium alloy to obtain a TLM titanium alloy substrate with a dual-phase microstructure; the heat treatment conditions are as follows: in an environment with a vacuum degree of 10 Pa, the temperature is raised at a rate of 5 ℃ / min, first heat-treated at 770 ℃ for 1 h, water-cooled, then heat-treated at 500 ℃ for 2 h, air-cooled;

[0060] Step two, the TLM titanium alloy substrate with a dual-phase microstructure obtained in step one is subjected to surface electrochemical de-alloying treatment, taking the TLM titanium alloy substrate as an anode and a platinum electrode as a cathode, using an acidic electrolyte and applying a voltage of 5 V, and the treatment time is 1 h, to obtain a TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure; the acidic electrolyte contains 0.3% hydrofluoric acid and 3% nitric acid by volume fraction;

[0061] Step three, tannic acid TA is mixed with strontium salt SrCl2·6H2O, the concentration of tannic acid in the mixed solution system is 4 mg / mL, and the concentration of strontium salt is 0.4 M, the mixed solution system is placed in a vortex oscillator for 15 min to mix and shake until the solution is light yellow for complexation reaction, to obtain a complex solution;

[0062] Step four, the TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure obtained in step two is immersed in 1 mL of the complex solution obtained in step three, the environmental temperature is controlled at 37 ℃ for chemical deposition treatment for 1 h, and ultrasonic cleaning is performed to remove the complex that is not firmly combined on the surface, to obtain a TLM titanium alloy with a bone and vascularization promoting functional layer;

[0063] Step five, the TLM titanium alloy with a bone and vascularization promoting functional layer obtained in step four is subjected to electrostatic adsorption to construct an antibacterial functional layer, to obtain a de-alloying / timing functional surface; the process of electrostatic adsorption is as follows: polylysine (ε-PL) powder is dissolved in a hydroxyethylpiperazine ethanesulfonic acid (HEPES) buffer solution with a concentration of 50 mM to obtain an electrostatic adsorption solution with a mass concentration of 5 mg / mL, and then 200 μL of the mixed solution is spotted on the surface of the TLM titanium alloy with a bone and vascularization promoting functional layer.

[0064] It is detected that the micropore size of the hierarchical micro-nano porous surface structure of the de-alloying / aging functional surface prepared on the TLM titanium alloy substrate in the embodiment is 2.79 μm, and the nanopore size is 29.27 nm, and the hierarchical micro-nano porous surface structure is beneficial to early cell adhesion.

[0065] Figure 3 The rat bone marrow mesenchymal stem cell alkaline phosphatase (ALP) staining diagram of the de-alloying / aging functional surface prepared in the embodiment and the TLM titanium alloy is shown, wherein Fig. (a) is the rat bone marrow mesenchymal stem cell alkaline phosphatase (ALP) staining diagram of the control group TLM titanium alloy (only polished by 2000# sandpaper), and Fig. (b) is the rat bone marrow mesenchymal stem cell alkaline phosphatase (ALP) staining diagram of the experimental group de-alloying / aging functional surface, from which it can be seen that the de-alloying / aging functional surface prepared in the embodiment has a significantly improved alkaline phosphatase expression, which indicates that the de-alloying / aging functional surface significantly improves the in-vitro osteogenic differentiation trend of the rat bone marrow mesenchymal stem cells, and the polylysine has antibacterial ability. Figure 3 It can be seen that the de-alloying / aging functional surface prepared in the embodiment has a significantly improved alkaline phosphatase expression, which indicates that the de-alloying / aging functional surface significantly improves the in-vitro osteogenic differentiation trend of the rat bone marrow mesenchymal stem cells, and the polylysine has antibacterial ability.

[0066] The above is only a preferred embodiment of the present application, and does not limit the present application. Any simple modification, change and equivalent change made according to the technical essence of the present application to the above embodiment are still within the protection scope of the technical solution of the present application.

Claims

1. A method of producing a dealloyed / aging functional surface of near-beta TLM titanium alloy, characterized in that, The method comprises the following steps: Step one, heat treatment is performed on the near-beta TLM titanium alloy to obtain a TLM titanium alloy substrate with a dual-phase microstructure; Step two, surface electrochemical dealloying treatment is performed on the TLM titanium alloy substrate with a dual-phase microstructure obtained in step one, a platinum electrode is used as a cathode, an acid electrolyte is used, and a constant voltage higher than a critical potential is applied to obtain a TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure; Step three, tannic acid and strontium salt are mixed to perform a complexation reaction to obtain a complex solution; Step four, chemical deposition treatment is performed on the TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure obtained in step two by using the complex solution obtained in step three to obtain a TLM titanium alloy with a function layer for promoting osteogenesis and vascularization; Step five, an antibacterial function layer is constructed on the TLM titanium alloy with the function layer for promoting osteogenesis and vascularization obtained in step four by electrostatic adsorption to obtain a dealloyed / sequential functional surface.

2. A method of producing a dealloyed / aged functional surface of near-beta TLM titanium alloy according to claim 1, characterized in that, In step one, the heat treatment is performed in a vacuum environment with a vacuum degree of 1 Pa to 10 Pa, the temperature is raised at a speed of 5 ℃ / min to 10 ℃ / min, the temperature is first raised to 760 ℃ to 790 ℃ for one-time heat treatment for 1 h, water cooling is performed, then the temperature is raised to 470 ℃ to 510 ℃ for two-time heat treatment for 2 h, and air cooling is performed.

3. A method of producing dealloyed / aging functional surface of near β TLM titanium alloy according to claim 1, characterized in that, In step two, the surface electrochemical dealloying treatment is performed by applying a voltage of 1 V to 5 V by using a constant potential instrument, and the treatment time is 1 h to 3 h; the acid electrolyte contains hydrofluoric acid with a volume fraction of 0.2% to 0.5% and nitric acid with a volume fraction of 2% to 5%.

4. A method of producing dealloyed / aging functional surface of near β TLM titanium alloy according to claim 1, characterized in that, In step two, the micropores of the TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure have a pore size of 0.5 μm to 2.8 μm, and the nanopores have a pore size of 12 nm to 27 nm.

5. A method of producing dealloyed / aging functional surface of near β TLM titanium alloy according to claim 1, characterized in that, In step three, the concentration of tannic acid in the mixed solution system is 1 mg / mL to 4 mg / mL, the concentration of strontium salt is 0.1 M to 0.4 M, and the complexation reaction process is that the mixed solution system is placed in a vortex oscillator and shaken for 15 min to mix the solution until the solution is light yellow.

6. A method of producing dealloyed / aging functional surface of near β TLM titanium alloy according to claim 1, characterized in that, In step four, the chemical deposition treatment process is that the TLM titanium alloy substrate with a hierarchical micro-nano porous surface structure is soaked in the complex solution, the environmental temperature is controlled to be 25 ℃ to 37 ℃, and the chemical deposition is performed for 1 h to 3 h.

7. A method of producing dealloyed / aging functional surface of near β TLM titanium alloy according to claim 1, characterized in that, In step five, the electrostatic adsorption process is that polylysine powder is dissolved in a hydroxyethylpiperazine ethylthiourea acid buffer solution with a concentration of 25 mM to 50 mM to obtain an electrostatic adsorption solution with a mass concentration of 0.5 mg / mL to 5 mg / mL, and then 50 μL to 200 μL of the mixed solution is spotted on the surface of the TLM titanium alloy with the function layer for promoting osteogenesis and vascularization.

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