Titanium-based implant material with osteogenesis and antibacterial properties and preparation method thereof

Abstract

The present invention discloses a preparation method of CaO2@ZIF-67-HA-ADH nanoparticles with osteopromoting and antibacterial properties, the method comprising: dissolving 2-methylimidazole in N,N-dimethylformamide to prepare solution A; adding CaO2 nanoparticles and Co(NO3)2 to dissolve in N,N-dimethylformamide to prepare solution B; adding solution A to solution B, reacting for 20 to 40 minutes, and preparing an intermediate CaO2@ZIF-67; dissolving HA in triple-distilled water, and activating the HA with EDC and NHS. After activation for a period of time, ADH is added. After a reaction time of 20-30 hours, the mixed solution is dialyzed using a dialysis bag with a molecular weight cutoff of 1.2-1.5 kDa. After dialysis, HA-ADH is produced by freeze-drying. The obtained intermediate CaO2@ZIF-67 is dispersed in a DMF solution, and the obtained HA-ADH is added to the solution. The pH of the solution is adjusted to 8-8.5, and the solution is stirred at room temperature for 20-30 hours. After the reaction is completed, CaO2@ZIF-67-HA-ADH nanoparticles are obtained. These nanoparticles are combined with titanium-based implant materials to form titanium-based implant materials with osteopromoting and antibacterial properties.

Description

Technical Field

[0001] The invention belongs to the field of biomedical materials, and in particular relates to a titanium-based implant material with osteogenesis and antibacterial properties and a preparation method thereof. Background Art

[0002] The information disclosed in this background technology section is only intended to enhance understanding of the overall background of the invention and should not necessarily be regarded as an admission or any form of suggestion that the information constitutes the prior art already known to those skilled in the art.

[0003] Titanium and its alloys are widely used as artificial implants in orthopedics, dentistry, and other hard tissue replacement and repair fields due to their excellent biocompatibility, mechanical properties, and chemical stability. However, the bioinertness and stress shielding effect of the titanium surface result in insufficient bonding with the surrounding bone tissue, which may shorten the service life of Ti implants. At the same time, implant-related infections caused by bacteria such as Staphylococcus aureus are very common complications in orthopedic and dental implant surgery, which may cause patients to experience symptoms such as pain, inflammation, functional impairment, and implant failure, seriously affecting their physical health and quality of life. Therefore, there is an urgent need to develop a titanium-based implant material with both excellent antibacterial and osteogenic properties to achieve ideal bone integration. Summary of the Invention

[0004] To overcome these issues, the present invention provides a titanium-based implant material with both osteopromoting and antibacterial properties and a method for preparing the material. The present invention combines CaO2 nanoparticles, the metal-organic framework ZIF-67, and the chemical conjugate HA-ADH to produce CaO2@ZIF-67-HA-ADH nanoparticles with both osteopromoting and antibacterial properties. These nanoparticles, when combined with a titanium-based implant material, form the titanium-based implant material with both osteopromoting and antibacterial properties.

[0005] In order to achieve the above technical objectives, the present invention adopts the following technical solutions:

[0006] The first aspect of the present invention provides a method for preparing CaO2@ZIF-67-HA-ADH nanoparticles having osteopromoting and antibacterial properties, the method comprising:

[0007] (1) 2-Methylimidazole was dissolved in N,N-dimethylformamide (DMF) to prepare solution A; CaO2 nanoparticles and Co(NO3)2 were dissolved in N,N-dimethylformamide to prepare solution B; solution A was added to solution B and reacted for 20 to 40 minutes to obtain the intermediate CaO2@ZIF-67, which was then washed and collected by centrifugation;

[0008] (2) Sodium hyaluronate (HA) was dissolved in triple-distilled water and activated using 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) and NN-hydroxysuccinimide (NHS). After activation for a period of time, adipic acid dihydrazide (ADH) was added. After reacting for 20 to 30 hours, the mixed solution was dialyzed using a dialysis bag with a molecular weight cut-off of 1.2 to 1.5 kDa. After dialysis, HA-ADH was freeze-dried.

[0009] (3) The prepared intermediate CaO2@ZIF-67 was dispersed in DMF solution, and then the prepared HA-ADH was added to the solution, and the pH of the solution was adjusted to 8-8.5. The solution was stirred at room temperature for 20-30 hours. After the reaction was completed, the CaO2@ZIF-67-HA-ADH nanoparticles were collected by centrifugation.

[0010] In the CaO2@ZIF-67-HA-ADH nanoparticles provided by the present invention, calcium peroxide (CaO2) slowly reacts with water in a neutral environment to generate Ca(OH)2 and O2, providing oxygen for bone tissue healing and Ca for the proliferation and differentiation of osteoblasts. 2+ and alkaline microenvironment. Under acidic conditions, the rate of this reaction will be greatly accelerated, thus achieving a bactericidal effect. ZIF-67 has the advantages of large surface area, controllable porosity, good biocompatibility and biodegradability, and shows great advantages in the biomedical field. ADH is grafted to HA to form a chemical conjugate: HA-ADH, which has excellent biocompatibility and stable biodegradability. Therefore, CaO2@ZIF-67-HA-ADH nanoparticles have good biocompatibility, antibacterial properties, and osteogenic properties.

[0011] The second aspect of the present invention provides CaO2@ZIF-67-HA-ADH nanoparticles prepared by the above preparation method.

[0012] A third aspect of the present invention provides a method for preparing a titanium-based implant material having osteogenesis and antibacterial properties, the method comprising:

[0013] S1. Pre-treating the titanium implant, wherein the pre-treatment includes: alkaline heat treatment and ultrasonic cleaning; the alkaline heat treatment includes placing the titanium implant in a sodium hydroxide solution and alkaline heat treatment in a water bath at 80-90° C. for 2-3 hours; the ultrasonic cleaning includes ultrasonic cleaning the titanium implant after alkaline heat treatment using ethanol, acetone, and triple-distilled water, respectively;

[0014] S2. Place the pretreated titanium implant in a dopamine hydrochloride-Tris solution and soak it in the dark for 20 to 30 hours. After taking out the sample, wash it with deionized water and dry it to obtain Ti-PDA.

[0015] S3. The CaO2@ZIF-67-HA-ADH nanoparticles prepared in the first aspect are dissolved in triple-distilled water, and then the Ti-PDA prepared in S2 is added and soaked in the dark for 20 to 30 hours. After taking out the sample, it is washed with deionized water and dried to obtain a titanium-based implant material with osteopromoting and antibacterial properties.

[0016] Dopamine is the main component of the surface coating of the titanium-based implant material provided by the present invention. The presence of dopamine can make the CaO2@ZIF-67-HA-ADH nanoparticles evenly dispersed on the surface of the titanium-based implant material; at the same time, dopamine can enhance the wettability of the titanium surface and promote the adhesion and colonization of osteoblasts.

[0017] The fourth aspect of the present invention provides a titanium-based implant material having osteogenesis and antibacterial properties prepared by the above-mentioned preparation method.

[0018] Beneficial effects of the present invention:

[0019] (1) In the CaO2@ZIF-67-HA-ADH nanoparticles provided by the present invention, calcium peroxide (CaO2) slowly reacts with water in a neutral environment to generate Ca(OH)2 and O2, providing oxygen for bone tissue healing and Ca for the proliferation and differentiation of osteoblasts. 2+ and alkaline microenvironment. Under acidic conditions, the rate of this reaction will be greatly accelerated, thus achieving a bactericidal effect. ZIF-67 has the advantages of large surface area, controllable porosity, good biocompatibility and biodegradability, and shows great advantages in the biomedical field. ADH is grafted to HA to form a chemical conjugate: HA-ADH, which has excellent biocompatibility and stable biodegradability. Therefore, CaO2@ZIF-67-HA-ADH nanoparticles have good biocompatibility, antibacterial properties, and osteogenic properties.

[0020] (2) The titanium-based implant material prepared by the present invention has strong antibacterial properties, with antibacterial rates against Staphylococcus aureus and Pseudomonas aeruginosa of 75.37% and 82.37%, respectively.

[0021] (3) The titanium-based implant material prepared by the present invention has excellent biocompatibility, and the CaO2@ZIF-67-HA-ADH nanoparticles on the surface of the titanium-based implant material can promote the proliferation of preosteoblasts.

[0022] (4) The titanium-based implant material prepared by the present invention can promote osteogenic differentiation. Through the ALP activity detection of pre-osteoblasts (MC3T3-E1), it was found that the ALP activity of the titanium-based implant material prepared by the present application was always higher than that of the untreated titanium-based implant material. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] The accompanying drawings, which constitute a part of the present invention, are used to provide a further understanding of the present invention. The exemplary embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute improper limitations on the present invention.

[0024] Figure 1 : TEM image of CaO2@ZIF-67-HA-ADH nanoparticles prepared in Example 1;

[0025] Figure 2 Elemental analysis image of CaO2@ZIF-67-HA-ADH nanoparticles prepared in Example 1;

[0026] Figure 3 : SEM image of the titanium-based implant material prepared in Example 2;

[0027] Figure 4 : The antibacterial performance image of the titanium-based implant material prepared in Example 2;

[0028] Figure 5 : Cell proliferation activity diagram of the titanium-based implant material prepared in Example 2;

[0029] Figure 6 : ALP activity diagram of the titanium-based implant material prepared in Example 2. DETAILED DESCRIPTION

[0030] It should be noted that the following detailed descriptions are exemplary and intended to provide further explanation of the present invention. Unless otherwise specified, all technical and scientific terms used in the present invention have the same meanings as those commonly understood by those skilled in the art to which the present invention belongs.

[0031] It should be noted that the terms used herein are only for describing specific embodiments and are not intended to limit the exemplary embodiments according to the present invention. As used herein, unless the context clearly indicates otherwise, the singular form is intended to include the plural form. In addition, it should be understood that when the terms "comprise" and / or "include" are used in this specification, they indicate the presence of features, steps, operations, devices, components and / or combinations thereof.

[0032] A first typical embodiment of the present invention provides a method for preparing CaO2@ZIF-67-HA-ADH nanoparticles having osteopromoting and antibacterial properties, the method comprising:

[0033] (1) 2-Methylimidazole was dissolved in N,N-dimethylformamide (DMF) to prepare solution A; CaO2 nanoparticles and Co(NO3)2 were dissolved in N,N-dimethylformamide to prepare solution B; solution A was added to solution B and reacted for 20 to 40 minutes to obtain the intermediate CaO2@ZIF-67, which was then washed and collected by centrifugation;

[0034] (2) Sodium hyaluronate (HA) was dissolved in triple-distilled water and activated using 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) and NN-hydroxysuccinimide (NHS). After activation for a period of time, adipic acid dihydrazide (ADH) was added. After reacting for 20 to 30 hours, the mixed solution was dialyzed using a dialysis bag with a molecular weight cut-off of 1.2 to 1.5 kDa. After dialysis, HA-ADH was freeze-dried.

[0035] (3) The prepared intermediate CaO2@ZIF-67 was dispersed in DMF solution, and then the prepared HA-ADH was added to the solution, and the pH of the solution was adjusted to 8-8.5. The solution was stirred at room temperature for 20-30 hours. After the reaction was completed, the CaO2@ZIF-67-HA-ADH nanoparticles were collected by centrifugation.

[0036] In one or more embodiments, in step (1), the preparation method of the CaO2 nanoparticles is as follows: CaCl2 and polyvinyl pyrrolidone (PVP) are dissolved in ethanol, ultrasonically dispersed, diluted ammonia water is added and then stirred vigorously for 30 minutes, H2O2 solution is added dropwise to obtain a light yellow precipitate, centrifuged, and vacuum dried overnight to obtain CaO2 nanoparticles. Preferably, the mass ratio of CaCl2 to polyvinyl pyrrolidone is 1:3-4, preferably 1:3.5; preferably, the concentration of CaCl2 in ethanol is 0.6-0.7mM, preferably 0.68mM; preferably, the concentration of the diluted ammonia water is 0.8M, and the concentration of the H2O2 solution is 1M; preferably, the volume ratio of the ethanol, diluted ammonia water, and H2O2 solution is 15:1:0.2.

[0037] In one or more embodiments, in step (1), the mass fraction of 2-methylimidazole in solution A is 2% to 3%, preferably 3%.

[0038] In one or more embodiments, in step (1), the mass fraction of Co(NO3)2 in solution B is 9% to 10%, preferably 10%.

[0039] In one or more embodiments, in step (1), the mass ratio of 2-methylimidazole to CaO2 nanoparticles and Co(NO3)2 is: 50-60:20:90-100; preferably 56.8:20:98.6.

[0040] In one or more embodiments, in step (1), solution A is added to solution B, and the reaction time is 30 minutes.

[0041] In one or more embodiments, in step (1), the washing condition is to wash three times with a mixed solution of DMF and methanol (volume ratio is 1:1).

[0042] In one or more embodiments, in step (2), the concentration of sodium hyaluronate in the aqueous solution is 0.1% to 0.2%, preferably 0.2%.

[0043] In one or more embodiments, the mass ratio of sodium hyaluronate, 1-ethyl-(3-dimethylaminopropyl)carbodiimide, NN-hydroxysuccinimide, and adipic acid dihydrazide in step (2) is 100-200:200-210:120-130:2500-3000, preferably 200:203.58:129.12:2740.

[0044] In one or more embodiments, the activation time in step (2) is 0.5 to 1 hour, preferably 1 hour.

[0045] In one or more embodiments, the mass ratio of CaO2@ZIF-67 to HA-ADH in step (3) is 3-4:8-10, preferably 4:10.

[0046] In one or more embodiments, the step (3) uses 0.8 M dilute ammonia water to adjust the pH of the solution.

[0047] In one or more embodiments, the stirring time in step (3) is 24 hours.

[0048] A second typical embodiment of the present invention provides CaO2@ZIF-67-HA-ADH nanoparticles prepared by the above preparation method.

[0049] A third typical embodiment of the present invention provides a method for preparing a titanium-based implant material having osteogenesis and antibacterial properties, the method comprising:

[0050] S1. Pre-treating the titanium implant, wherein the pre-treatment includes: alkaline heat treatment and ultrasonic cleaning; the alkaline heat treatment includes placing the titanium implant in a sodium hydroxide solution and alkaline heat treatment in a water bath at 80-90° C. for 2-3 hours; the ultrasonic cleaning includes ultrasonic cleaning the titanium implant after alkaline heat treatment using ethanol, acetone, and triple-distilled water, respectively;

[0051] S2. Place the pretreated titanium implant in a dopamine hydrochloride-Tris solution and soak it in the dark for 20 to 30 hours. After taking out the sample, wash it with deionized water and dry it to obtain Ti-PDA.

[0052] S3. The CaO2@ZIF-67-HA-ADH nanoparticles prepared in the first aspect are dissolved in triple-distilled water, and then the Ti-PDA prepared in S2 is added and soaked in the dark for 20 to 30 hours. After taking out the sample, it is washed with deionized water and dried to obtain a titanium-based implant material with osteopromoting and antibacterial properties.

[0053] In one or more embodiments, in S1, the concentration of the sodium hydroxide solution is 5-8M, preferably 6M.

[0054] In one or more embodiments, in S1, the temperature of the water bath is 90° C., and the time of the alkali heat treatment is 3 hours.

[0055] In one or more embodiments, in S1, the ultrasonic cleaning treatment process is performed for 10 to 20 minutes for each solution, preferably 15 minutes.

[0056] In one or more embodiments, in S2, the volume ratio of the dopamine hydrochloride in the dopamine hydrochloride-Tris solution to the Tris solution is 2-3 mg:1 mL, preferably 2 mg:1 mL.

[0057] In one or more embodiments, in S2, the light-proof immersion time is 24 hours.

[0058] In one or more embodiments, in S2, the method of cleaning with deionized water is: ultrasonic cleaning with deionized water 5 times, each time for 5 minutes.

[0059] In one or more embodiments, in S3, the volume ratio of CaO2@ZIF-67-HA-ADH to triple-distilled water is 2-3 mg:1 mL, preferably 2 mg:1 mL.

[0060] In one or more embodiments, in S3, the light-proof immersion time is 24 hours.

[0061] In one or more embodiments, in S3, the method of cleaning with deionized water is: ultrasonic cleaning with deionized water for 3 times, each time for 5 minutes.

[0062] A fourth typical embodiment of the present invention provides a titanium-based implant material having osteogenesis and antibacterial properties prepared by the above preparation method.

[0063] In order to enable those skilled in the art to more clearly understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0064] In a specific embodiment of the present invention, all centrifugal speeds are 12000 rpm / min; and the ultrasonic dispersion and cleaning frequencies are 50 KHz.

[0065] Example 1

[0066] Synthesis of CaO2 Nanoparticles

[0067] Dissolve 0.1 g of CaCl2 and 0.35 g of PVP in 15 mL of ethanol and disperse them ultrasonically. Then add 1 mL of 0.8 M ammonia water and continue stirring vigorously for 30 minutes. Then add 0.2 mL of 1 M H2O2 dropwise to obtain a light yellow precipitate. Collect it by centrifugation (12000 rpm / min) and dry it in vacuum overnight.

[0068] Synthesis of CaO2@ZIF-67:

[0069] 56.8 mg of 2-methylimidazole was dissolved in 2 mL of DMF to prepare solution A; 20 mg of CaO2 and 98.6 mg of Co(NO3)2 were dissolved together in 2 mL of DMF and ultrasonically dispersed at room temperature to prepare solution B; solution A was added to solution B, and the reaction was continued for 30 minutes, followed by washing three times with a mixed solution of DMF and methanol (volume ratio 1:1) and collecting by centrifugation.

[0070] Synthesis of HA-ADH:

[0071] 0.2 g of HA was completely dissolved in 100 mL of triple-distilled water (pH 5.8), followed by the addition of 203.58 mg of EDC and 129.12 mg of NHS. After activation for 1 hour, 2.74 g of adipic acid dihydrazide was added and stirred for 24 hours. The mixed solution was dialyzed using a dialysis bag with a molecular weight cutoff of 1.2 to 1.5 kDa. After dialysis, HA-ADH was obtained by freeze-drying.

[0072] Preparation of CaO2@ZIF-67-HA-ADH nanoparticles:

[0073] 40 mg of CaO2@ZIF-67 was dispersed in 5 mL of DMF solution, and then 100 mg of HA-ADH was added. The pH was adjusted to 8-8.5 using 0.8 M dilute ammonia water. The reaction was stirred at room temperature for 24 hours, and the product was collected by centrifugation to obtain CaO2@ZIF-67-HA-ADH nanoparticles.

[0074] The CaO2@ZIF-67-HA-ADH nanoparticles prepared in Example 1 were characterized. Figure 1 For TEM images, Figure 1 As shown in the figure, the microscopic morphology of CaO2@ZIF-67-HA-ADH nanoparticles is regular and circular. Figure 2 This is an elemental analysis image. Elemental analysis proves that CaO2@ZIF-67-HA-ADH nanoparticles contain elements such as Ca, C, O, Co, and N.

[0075] Example 2

[0076] Preparation of titanium-based implant materials with osteopromoting and antibacterial properties:

[0077] Titanium foil (1 cm x 1 cm x 0.01 mm) was placed in a sodium hydroxide solution and then alkali-heat treated in a 90°C water bath for 3 hours. The alkali-heat treated titanium implants were ultrasonically cleaned using ethanol, acetone, and triple-distilled water, with each solution ultrasonically cleaned for 15 minutes.

[0078] 200 mg of dopamine was dissolved in 200 mL of Tris solution to prepare a dopamine-Tris solution (pH 8.5). The pretreated titanium foil was placed in the dopamine-Tris solution and soaked in the dark for 24 hours. After removal, the sample was ultrasonically cleaned with deionized water five times for 5 minutes each, then dried in a drying oven to obtain Ti-PDA.

[0079] 200mg of CaO2@ZIF-67-HA-ADH nanoparticles were added to 200mL of triple-distilled water, followed by the Ti-PDA, which was then placed in the solution and immersed in the dark for 24 hours. The sample was removed and ultrasonically cleaned in deionized water three times for 5 minutes each time, followed by drying in a drying oven to produce a titanium-based implant with both osteopromoting and antibacterial properties.

[0080] The titanium-based implant material prepared in this embodiment was characterized using SEM. Figure 3 As shown, CaO2@ZIF-67-HA-ADH nanoparticles can maintain their original morphology and be distributed in an orderly manner on the surface of titanium-based implant materials.

[0081] Example 3

[0082] In vitro antibacterial performance test of the titanium-based implant material prepared in Example 2

[0083] The antibacterial properties of the titanium-based implant material prepared in Example 2 were verified using Gram-positive Staphylococcus aureus (S. aureus) and Gram-negative Pseudomonas aeruginosa (P. aeruginosa) as representative bacteria.

[0084] Under a sterile environment, Staphylococcus aureus or Pseudomonas aeruginosa were revived and inoculated on LB agar plates (containing 10 g / L tryptone, 5 g / L yeast, 10 g / L sodium chloride, and 15 g / L agar) and placed in a 37°C incubator. After 24 hours, Staphylococcus aureus or Pseudomonas aeruginosa with good growth were inoculated into LB liquid medium (containing 10 g / L tryptone, 5 g / L yeast, and 10 g / L sodium chloride). The bacterial concentration was determined using a bacterial turbidity meter and prepared into 1×10 6 A bacterial suspension with a CFU / mL was prepared for later use. 1 mL of the bacterial suspension was inoculated onto the surface of the titanium-based implant material prepared in Example 2 and the surface of the titanium-based material sample without any treatment in a 24-well plate. After incubation at 37°C for 24 hours, Staphylococcus aureus or Pseudomonas aeruginosa was eluted from the sample surface. The eluate was diluted 10,000 times and then spread on an LB agar plate. After incubation at 37°C for 24 hours, the bacteria on each LB agar plate were counted, and the antibacterial rate was calculated.

[0085] Antibacterial rate = (number of colonies in the control group - number of colonies in the experimental group) ÷ number of colonies in the control group × 100%.

[0086] The results are as follows Figure 4 As shown, compared with the untreated titanium-based material, the titanium-based implant material prepared in Example 2 had an antibacterial rate of 75.37% against Staphylococcus aureus and 82.37% against Pseudomonas aeruginosa, respectively. This demonstrates that the titanium-based implant material prepared in Example 2 has excellent antibacterial properties and has a strong antibacterial effect against both Gram-positive and Gram-negative bacteria.

[0087] Example 4

[0088] The cell proliferation activity of the titanium-based implant material prepared in Example 2 was detected

[0089] The cell proliferation activity of the titanium-based implant preosteoblasts (MC3T3-E1) prepared in Example 2 was evaluated by CCK-8 assay. Specifically: MC3T3-E1 (1×10 4The titanium-based implant material prepared in Example 2 and the titanium-based material without any treatment were inoculated (per well) in a sterile incubator for 4 and 7 days, respectively. The culture medium was discarded and the samples were gently washed 3 times with PBS. 500 ml of pre-prepared 10% CCK-8 reagent was added to each well. The 24-well plate was returned to the incubator and incubated at 37 ° C in the dark for 2 hours. After the incubation time was reached, 100 μL of the reaction solution was aspirated and placed in a 96-well plate. Finally, the absorbance value (OD value) was read at 450 nm using a microplate reader.

[0090] The results are as follows Figure 5 As shown, on the 4th and 7th days, the cell proliferation activity of the titanium-based implant material prepared in Example 2 was higher than that of the titanium-based material group without any treatment. Therefore, it can be proved that the titanium-based implant material prepared in Example 2 can promote the proliferation of preosteoblasts and has excellent biocompatibility.

[0091] Example 5

[0092] The titanium-based implant material prepared in Example 2 was tested for osteogenic differentiation:

[0093] In a 24-well plate, MC3T3-E1 was plated at 2 × 10 4 Cells were seeded on the coating surface at a density of 10 cells / well and then cultured in osteogenic induction medium for 7 or 14 days. Finally, the ALP activity of the cells was measured using an ALP assay kit (Beyotime, China).

[0094] The results are as follows Figure 5 As shown, whether it is 4 days or 7 days, the ALP activity of the titanium-based implant material prepared in Example 2 is always higher than that of the titanium-based material group without any treatment, which indicates that the titanium-based implant material prepared in Example 2 has excellent osteogenic properties.

[0095] The foregoing description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Those skilled in the art will readily appreciate that various modifications and variations of the present invention are possible. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention are intended to be within the scope of protection of the present invention.

Claims

1. A method for preparing CaO2@ZIF-67-HA-ADH nanoparticles with osteogenic and antibacterial properties, characterized in that: The method comprises: (1) 2-Methylimidazole was dissolved in N,N-dimethylformamide (DMF) to prepare solution A; CaO2 nanoparticles and Co(NO3)2 were dissolved in N,N-dimethylformamide to prepare solution B; solution A was added to solution B and reacted for 20-40 min to obtain the intermediate CaO2@ZIF-67, which was then washed and collected by centrifugation; (2) Sodium hyaluronate HA is dissolved in triple-distilled water and activated using 1-ethyl-(3-dimethylaminopropyl)carbodiimide EDC and NN-hydroxysuccinimide NHS. After activation for a period of time, adipic acid dihydrazide ADH is added. After reacting for 20-30 hours, the mixed solution is dialyzed using a dialysis bag with a molecular weight cutoff of 1.2-1.5 KDA. After dialysis, HA-ADH is freeze-dried. (3) The prepared intermediate CaO2@ZIF-67 was dispersed in DMF solution, and then the prepared HA-ADH was added to the solution. The pH of the solution was adjusted to 8-8.5 and stirred at room temperature for 20-30 hours. After the reaction was completed, the CaO2@ZIF-67-HA-ADH nanoparticles were collected by centrifugation.

2. The preparation method according to claim 1, wherein In the step (1), the preparation method of the CaO2 nanoparticles is as follows: CaCl2 and polyvinylpyrrolidone (PVP) are dissolved in ethanol, ultrasonically dispersed, diluted ammonia water is added and then stirred vigorously for 30 minutes, H2O2 solution is added dropwise to obtain a light yellow precipitate, and after centrifugation, vacuum drying is carried out overnight to obtain CaO2 nanoparticles.

3. The preparation method according to claim 1, wherein In the step (1), the mass fraction of 2-methylimidazole in the solution A is 2% to 3%.

4. The preparation method according to claim 1, wherein In the step (1), the mass fraction of Co(NO3)2 in the solution B is 9% to 10%.

5. The preparation method according to claim 1, wherein In the step (1), the mass ratio of the 2-methylimidazole to the CaO2 nanoparticles and the Co(NO3)2 is 50-60:20:90-100.

6. The preparation method according to claim 1, wherein In the step (1), solution A is added to solution B, and the reaction time is 30 minutes.

7. The preparation method according to claim 1, wherein In the step (1), the washing condition is to wash three times with a mixed solution of DMF and methanol in a volume ratio of 1:

1.

8. The preparation method according to claim 2, wherein The mass ratio of CaCl2 to polyvinyl pyrrolidone is 1:3~4.

9. The preparation method according to claim 2, wherein The mass ratio of CaCl2 to polyvinyl pyrrolidone is 1:3.

5.

10. The preparation method according to claim 2, wherein The concentration of CaCl2 in ethanol is 0.6~0.7mM.

11. The preparation method according to claim 2, wherein The concentration of CaCl2 in ethanol was 0.68 mM.

12. The preparation method according to claim 2, wherein The concentration of the dilute ammonia solution is 0.8 M, and the concentration of the H2O2 solution is 1 M.

13. The preparation method according to claim 2, wherein The volume ratio of the ethanol, dilute ammonia water and H2O2 solution is 15:1:0.

2.

14. The preparation method according to claim 1, wherein The mass fraction of 2-methylimidazole in the solution A is 3%.

15. The preparation method according to claim 1, wherein The mass fraction of Co(NO3)2 in the solution B is 10%.

16. The preparation method according to claim 1, wherein The mass ratio of the 2-methylimidazole to the CaO2 nanoparticles and the Co(NO3)2 is 56.8:20:98.

6.

17. The preparation method according to claim 1, wherein In step (2), the concentration of sodium hyaluronate in the aqueous solution is 0.1% to 0.2%; Alternatively, the mass ratio of sodium hyaluronate, 1-ethyl-(3-dimethylaminopropyl)carbodiimide, NN-hydroxysuccinimide, and adipic acid dihydrazide in step (2) is 100-200:200-210:120-130:2500-3000; Alternatively, the activation time is 0.5 to 1 h.

18. The preparation method according to claim 1, wherein In step (2), the concentration of sodium hyaluronate in the aqueous solution is 0.2%.

19. The preparation method according to claim 1, wherein The mass ratio of sodium hyaluronate, 1-ethyl-(3-dimethylaminopropyl)carbodiimide, NN-hydroxysuccinimide, and adipic acid dihydrazide in step (2) is 200:203.58:129.12:2740.

20. The preparation method according to claim 1, wherein The activation time is 1 h.

21. The preparation method according to claim 1, wherein In the step (3), the mass ratio of CaO2@ZIF-67 to HA-ADH is 3-4:8-10; Alternatively, the pH of the solution can be adjusted using 0.8 M dilute ammonia solution; Alternatively, the stirring time in step (3) is 24 hours.

22. The preparation method according to claim 1, wherein In the step (3), the mass ratio of CaO2@ZIF-67 to HA-ADH is 4:

10.

23. CaO2@ZIF-67-HA-ADH nanoparticles prepared by the preparation method according to any one of claims 1 to 22.

24. A method for preparing a titanium-based implant material with osteopromoting and antibacterial properties, characterized in that: The method comprises: S1. Pre-treating the titanium implant, wherein the pre-treatment includes: alkaline heat treatment and ultrasonic cleaning; the alkaline heat treatment includes placing the titanium implant in a sodium hydroxide solution and alkaline heat treatment in a water bath at 80-90°C for 2-3 hours; the ultrasonic cleaning includes ultrasonic cleaning the titanium implant after alkaline heat treatment using ethanol, acetone, and triple-distilled water, respectively; S2. Place the pretreated titanium implant in a dopamine hydrochloride-Tris solution and soak it in the dark for 20-30 hours. After taking out the sample, wash it with deionized water and dry it to obtain Ti-PDA. S3. Dissolve the CaO2@ZIF-67-HA-ADH nanoparticles described in claim 23 in triple-distilled water, then add the Ti-PDA prepared in S2, soak in the dark for 20 to 30 hours, take out the sample, wash it with deionized water, and dry it to obtain a titanium-based implant material with osteopromoting and antibacterial properties.

25. The preparation method according to claim 24, characterized in that In S1, the concentration of the sodium hydroxide solution is 5-8M; Alternatively, the temperature of the water bath is 90° C., and the alkali heat treatment time is 3 h; Alternatively, during the ultrasonic cleaning process, the ultrasonic time of each solution is 10 to 20 minutes.

26. The preparation method according to claim 24, wherein In S1, the concentration of the sodium hydroxide solution is 6M.

27. The preparation method according to claim 24, wherein During the ultrasonic cleaning process, the ultrasonic time of each solution was 15 minutes.

28. The preparation method according to claim 24, wherein In S2, the volume ratio of dopamine hydrochloride to Tris solution in the dopamine hydrochloride-Tris solution is 2-3 mg:1 mL; Or, the light-proof soaking time is 24 hours; Alternatively, the method of cleaning with deionized water is: ultrasonic cleaning with deionized water 5 times, each time for 5 minutes.

29. The preparation method according to claim 24, wherein In S2, the volume ratio of the dopamine hydrochloride in the dopamine hydrochloride-Tris solution to the Tris solution is 2 mg:1 mL.

30. The preparation method according to claim 24, wherein In S3, the volume ratio of CaO2@ZIF-67-HA-ADH to triple-distilled water is 2-3 mg:1 mL; Or, the light-proof soaking time is 24 hours; Alternatively, the method of cleaning with deionized water is: ultrasonic cleaning with deionized water 3 times, each time for 5 minutes.

31. The preparation method according to claim 24, wherein In S3, the volume ratio of CaO2@ZIF-67-HA-ADH to triple-distilled water is 2 mg:1 mL.

32. A titanium-based implant material prepared by the preparation method according to any one of claims 24 to 31.

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