Preparation method and application of zirconium-aluminum bone-promoting electroactive film modified by magnetron sputtering technology

Abstract

This invention discloses a method for preparing and applying a magnetron sputtering-modified zirconium-aluminum osteogenic electroactive film, relating to the field of osteogenic film material technology. The method involves cleaning and drying a PVDF film; fixing the PVDF film onto a glass slide and placing it in a magnetron sputtering chamber; and depositing elemental zirconium and aluminum onto the PVDF film using magnetron sputtering to prepare the magnetron sputtering-modified zirconium-aluminum osteogenic electroactive film. The magnetron sputtering-modified zirconium-aluminum osteogenic electroactive film contains 46.15-46.35 wt% zirconium, 19.25-19.45 wt% oxygen, 19.25-19.45 wt% fluorine, and 1.77-1.97 wt% aluminum. The film material prepared by this invention possesses excellent biocompatibility, antibacterial properties, and osteogenic induction ability, expanding its application scenarios in the oral cavity and other parts of the body.

Description

Technical Field

[0001] This invention relates to the field of bone electroactive thin film materials technology, and particularly to a method for preparing zirconium-aluminum bone electroactive thin films modified by magnetron sputtering. Background Technology

[0002] Traditional bone tissue engineering repairs bone defects by seeding cells onto scaffold materials and implanting them into the defect site. However, this method suffers from drawbacks such as seed cell apoptosis and triggering of immune inflammatory responses, which weaken the repair effect. To address these shortcomings, researchers have proposed the concept of "in-situ tissue engineering," constructing materials with high osteogenic response to accelerate the repair process at the defect site through the body's regenerative potential. This method focuses on utilizing material properties to induce endogenous stem cells to the injury site and mediate osteogenic differentiation to achieve bone repair goals. The key lies in mimicking the natural extracellular physiological regenerative microenvironment of bone. Electroactive materials are materials that can generate electrical signals under external stimulation or whose physicochemical properties can be altered under electrical signal stimulation. Currently, a series of electroactive biomaterials, including ferroelectric, piezoelectric, electret, and conductive materials and their mediated electrostimulation materials, have been developed to mimic the natural physiological electrical microenvironment, serving as biophysical clues for regulating stem cell fate and regenerative medicine. Electroactive materials are an effective bone repair material for constructing a regenerative electrical microenvironment.

[0003] When subjected to external forces, PVDF (polyvinylidene fluoride polymer) piezoelectric films exhibit polarized charges on their surface, similar to the piezoelectric properties of natural bone tissue—a phenomenon known as the piezoelectric effect. Due to their excellent piezoelectric properties and large specific surface area, PVDF bone-electroactive films have wide applications in bone tissue engineering, bone repair, biosensors, and drug delivery. However, practical applications of PVDF bone-electroactive films still suffer from problems such as poor electrical signal stability, poor bone regeneration regulation, and slow bone healing. Therefore, how to prepare a PVDF bone-electroactive film with good biocompatibility, antibacterial activity, stable existence in vivo, and the ability to regulate bone regeneration has become a research hotspot for the application of PVDF materials in electroactive bone repair materials. Summary of the Invention

[0004] This invention provides a method for preparing and applying a magnetron sputtering modified zirconium aluminum osteoactive film. The prepared film material possesses superior biocompatibility, antibacterial properties, and osteogenic induction ability, expanding its application scenarios in the oral cavity and other sites. It effectively solves the shortcomings of existing PVDF osteoactive films in terms of poor bone integration, bone healing speed, and effectiveness. Specifically, this is achieved through the following techniques.

[0005] A method for preparing a zirconium-aluminum osteoactive film modified by magnetron sputtering includes the following steps: taking a PVDF film, cleaning and drying it; fixing the PVDF film on a glass slide and placing it in a magnetron sputtering chamber; depositing elemental zirconium and elemental aluminum on the PVDF film by magnetron sputtering to obtain the zirconium-aluminum osteoactive film; wherein the zirconium-aluminum osteoactive film contains 46.15-46.35 wt% zirconium, 19.25-19.45 wt% oxygen, 19.25-19.45 wt% fluorine, and 1.77-1.97 wt% aluminum.

[0006] Furthermore, in the zirconium-aluminum osteo-promoting electroactive film, the zirconium content is 46.25 wt%, the oxygen content is 19.35 wt%, the fluorine content is 19.35 wt%, and the aluminum content is 1.87 wt%.

[0007] Furthermore, the magnetron sputtering is performed under the following conditions: in an inert atmosphere, the gas pressure in the magnetron sputtering chamber is 0.5-5 Pa, the sputtering distance is 4-10 cm, the sputtering power is maintained at 0-100 W, the substrate temperature is room temperature to 300 ℃, the argon flow rate is 5-80 sccm, and the duration is 10-20 min.

[0008] Furthermore, the magnetron sputtering is performed under the following conditions: in an inert atmosphere, the gas pressure in the magnetron sputtering chamber is 0.5 Pa, the sputtering distance is 5 cm, the sputtering power is maintained at 50 W, the substrate temperature is 200 ℃, the argon flow rate is 80 sccm, and the duration is 20 min.

[0009] The present invention also provides a zirconium aluminum osteogenic electroactive film prepared by any of the above-described preparation methods.

[0010] The present invention also provides the application of the above-mentioned zirconium aluminum osteogenic electroactive film in the preparation of bone defect repair or tissue engineering materials.

[0011] Compared with the prior art, the advantages of the present invention are:

[0012] 1. Compared with existing ordinary PVDF bone electroactive films, this invention effectively modifies the surface of PVDF bone electroactive films using elemental zirconium and aluminum through magnetron sputtering technology. The prepared zirconium-aluminum bone electroactive film can effectively stimulate macrophages to express T lymphocyte activation antigen (CD86), interleukin-1β (IL-1β), interferon regulatory factor 3 (IRF3), TNF receptor-associated factor 6 (TRAF6), and nuclear factor receptor-κB (NF-κB), thereby exhibiting excellent antibacterial activity.

[0013] 2. Compared with existing ordinary PVDF osteoactive films, the zirconium aluminum osteoactive film prepared in this invention can promote the expression of RUNX family transcription factor 2 (RUNX2) in osteoblasts, indicating that it has good osteogenic induction activity and very good bone integration and bone repair effects. Attached Figure Description

[0014] Figure 1 This is a schematic diagram of the zirconium-aluminum osteo-promoting electroactive film modified by magnetron sputtering technology in this invention.

[0015] Figure 2 The image shows the overall imaging of the prepared PVDF bone electroactive film using a scanning electron microscope at 10,000x magnification, as described in Example 1.

[0016] Figure 3 This is the energy spectrum generated by scanning the surface elements of the modified zirconium aluminum osteogenic film using an energy spectrum scanning module in Example 1.

[0017] Figure 4 The graph shows the relative mRNA expression of CD86 in RAW264.7 macrophages after culturing macrophages with the modified zirconium aluminum osteoactive membrane used in Example 1 and the PVDF osteoactive membrane treated in Comparative Example 1.

[0018] Figure 5 The graph shows the relative mRNA expression of IL-1β in RAW264.7 macrophages after culturing macrophages with the modified zirconium aluminum osteoactive membrane used in Example 1 and the PVDF osteoactive membrane treated in Comparative Example 1.

[0019] Figure 6 The graph shows the relative mRNA expression of IRF3 in RAW264.7 macrophages after culturing macrophages with the modified zirconium aluminum osteoactive membrane used in Example 1 and the PVDF osteoactive membrane treated in Comparative Example 1.

[0020] Figure 7 The graph shows the relative mRNA expression of TRAF6 in RAW264.7 macrophages after culturing macrophages with the modified zirconium aluminum osteoactive membrane used in Example 1 and the PVDF osteoactive membrane treated in Comparative Example 1.

[0021] Figure 8 The graph shows the relative mRNA expression of NF-κB in RAW264.7 macrophages after culturing macrophages with the modified zirconium aluminum osteoactive membrane used in Example 1 and the PVDF osteoactive membrane treated in Comparative Example 1.

[0022] Figure 9This is a graph showing the relative mRNA expression of RUNX2 in osteoblasts mt-3t3 after the modified zirconium aluminum osteoactive membrane used in Example 1 and the PVDF osteoactive membrane treated in Comparative Example 1 were cultured.

[0023] Figure 10-13 The images show the relative mRNA expression of IL-6, NF-κB, MYD88, and TRAF6 in RAW264.7 macrophages after culturing the osteogenic active films of Example 1 and Comparative Examples 2 and 3, respectively. Detailed Implementation

[0024] The technical solution of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0025] Example 1

[0026] The zirconium aluminum osteogenic electroactive film of this embodiment, such as Figure 1 As shown, it was prepared using the following method:

[0027] (1) Cut the PVDF (polyvinylidene fluoride) film into 2 cm × 2 cm square films, ultrasonically clean the PVDF film with anhydrous ethanol for 15 min, blow dry with nitrogen and then store in vacuum. The selected PVDF film was purchased from PolyK Company and had a thickness of 28 μm.

[0028] (2) Adhere conductive adhesive to the glass slide, attach the PVDF film treated in S1 to the conductive adhesive, and invert the glass slide and PVDF film into the magnetron sputtering chamber.

[0029] (3) Zirconium and aluminum were installed as targets in the DC target region of magnetron sputtering. After the chamber was closed, high-purity argon gas was introduced to maintain the gas pressure in the magnetron sputtering chamber at 5 Pa. Zirconium and aluminum were deposited on the PVDF osteoactive film under the argon atmosphere. The sputtering distance was 10 cm, the sputtering power was maintained at 100 W, the substrate temperature was 300 ℃, the argon gas flow rate was 5 sccm, and the duration was 20 min, finally obtaining the modified zirconium-aluminum osteoactive film.

[0030] The zirconium-aluminum osteogenic electroactive film prepared in this embodiment was imaged using a scanning electron microscope at 10,000x magnification. The results are as follows. Figure 2 As shown, the surface of the modified PVDF bone electroactive film is relatively smooth.

[0031] Subsequently, the surface elements of the zirconium aluminum osteogenic electroactive film were scanned using an energy dispersive spectroscopy (EDS) module, and the results are as follows: Figure 3 As shown in Table 1, stable amounts of elemental zirconium and aluminum were generated. The zirconium content was 46.25 wt%, the oxygen content was 32.53 wt%, the fluorine content was 19.35 wt%, and the aluminum content was 1.87 wt%.

[0032] Table 1 Energy Spectrum Scan Results

[0033]

[0034] Example 2

[0035] The zirconium aluminum osteo-promoting electroactive film of this embodiment was prepared using the following method:

[0036] (1) Cut the PVDF film into square films of 2 cm × 2 cm, ultrasonically clean the PVDF film with anhydrous ethanol for 15 min, blow it dry with nitrogen and then store it in a vacuum.

[0037] (2) Adhere conductive adhesive to the glass slide, attach the PVDF film treated in S1 to the conductive adhesive, and invert the glass slide and PVDF film into the magnetron sputtering chamber.

[0038] (3) Zirconium and aluminum were installed as targets in the DC target region of magnetron sputtering. After the chamber was closed, high-purity argon gas was introduced to maintain the gas pressure in the magnetron sputtering chamber at 0.5 Pa. Zirconium and aluminum were deposited on the PVDF osteoactive film under the argon atmosphere. The sputtering distance was 10 cm, the sputtering power was maintained at 100 W, the substrate temperature was room temperature, the argon gas flow rate was 5 sccm, and the duration was 30 min, finally obtaining the modified zirconium-aluminum osteoactive film.

[0039] Example 3

[0040] The zirconium aluminum osteo-promoting electroactive film of this embodiment was prepared using the following method:

[0041] (1) Cut the PVDF film into square films of 2 cm × 2 cm, ultrasonically clean the PVDF film with anhydrous ethanol for 15 min, blow it dry with nitrogen and then store it in a vacuum.

[0042] (2) Adhere conductive adhesive to the glass slide, attach the PVDF film treated in S1 to the conductive adhesive, and invert the glass slide and PVDF film into the magnetron sputtering chamber.

[0043] (3) Zirconium and aluminum were installed as targets in the DC target region of magnetron sputtering. After the chamber was closed, high-purity argon gas was introduced to maintain the gas pressure in the magnetron sputtering chamber at 0.5 Pa. Zirconium and aluminum were deposited on the PVDF osteoactive film under the argon atmosphere. The sputtering distance was 5 cm, the sputtering power was maintained at 50 W, the substrate temperature was 200 °C, the argon flow rate was 80 sccm, and the duration was 20 min, finally obtaining the modified zirconium-aluminum osteoactive film.

[0044] Comparative Example 1

[0045] The film material of this comparative example was prepared by the following method: the PVDF film was cut into 2 cm × 2 cm square films, the PVDF film was ultrasonically cleaned with anhydrous ethanol for 15 min, dried with nitrogen and stored in vacuum as the final film material.

[0046] Comparative Example 2

[0047] The difference between the preparation method of the thin film material in this comparative example and that in Example 1 is that only elemental zirconium is installed as the target material in the DC target region of magnetron sputtering, and elemental zirconium is doped onto the PVDF thin film; the zirconium content on the surface of the thin film material is approximately 48.12%, which is the sum of the zirconium and aluminum content in Example 1. No elemental aluminum or any other metallic / non-metallic materials are doped.

[0048] Comparative Example 3

[0049] The difference between the preparation method of the thin film material in this comparative example and that in Example 1 is that only elemental aluminum is installed as the target material in the DC target region of magnetron sputtering, and elemental aluminum is doped onto the PVDF thin film; the aluminum content on the surface of the thin film material is approximately 48.12%, which is the sum of the zirconium and aluminum content in Example 1. No elemental zirconium or any other metallic / non-metallic materials are doped.

[0050] Experimental Example 1: Performance Testing of Thin Film Materials Prepared in Example 1 and Comparative Example 1

[0051] 1. Antibacterial properties

[0052] The zirconium-aluminum osteogenic electroactive films prepared in Example 1 and Comparative Example 1 were soaked in alcohol, respectively, and then sterilized by ultraviolet irradiation. They were then placed in six-well cell culture plates and seeded with RAW264.7 macrophages at a density of 50%. After two days, when the cell density increased to 80%, the cells were collected for qPCR experiments to detect the expression of T lymphocyte activation antigen (CD86), interleukin-1β (IL-1β), interferon regulatory factor 3 (IRF3), TNF receptor-associated factor 6 (TRAF6), and nuclear factor receptor-κB (NF-κB). Statistical analysis and plotting were performed using GraphPad Prism software. The results are shown below. Figure 4-8 As shown, the zirconium and aluminum-modified PVDF osteoactive film of Example 1 promotes the expression of inflammatory factors such as CD86, IL-1β, IRF3, TRAF6, and NF-κB in macrophages, indicating that it enhances the anti-inflammatory activity of macrophages and gives them stronger antibacterial properties. In contrast, the antibacterial properties of the ordinary PVDF film in Comparative Example 1 are very limited.

[0053] The osteogenic electrolytic films prepared in Example 1 and Comparative Examples 2 and 3 were soaked in alcohol using the same method. The expression levels of interleukin-6 (IL-6), nuclear factor receptor-κB (NF-κB), MYD88, and TNF receptor-associated factor 6 (TRAF6) were then detected, and statistical analysis and plotting were performed using ImageJ software. The results are as follows: Figure 10-13 As shown, the osteogenic active film doped with elemental zirconium and aluminum can promote the expression of inflammatory factors such as IL-6, NF-κB, MYD88 and TRAF6 in macrophages, indicating that it enhances the anti-inflammatory activity of macrophages and gives them stronger antibacterial properties.

[0054] 2. Evaluation of osteogenic activity

[0055] The film materials prepared in Example 1 and Comparative Example 1 were soaked in alcohol and then sterilized by ultraviolet irradiation. They were then placed in six-well cell culture plates and seeded with osteoblast mt-3t3 cells at a density of 50%. Two days later, when the cell density reached 80%, the cells were collected for qPCR experiments to detect the expression of RUNX family transcription factor 2 (RUNX2). Statistical analysis and plotting were performed using GraphPad Prism software. The results are shown below. Figure 9 As shown, the PVDF osteogenic membrane modified with zirconium and aluminum in Example 1 upregulated the expression of the RUNX2 transcription factor in osteoblasts, suggesting that it has better osteogenic activity.

[0056] The above detailed embodiments describe the implementation of the present invention; however, the present invention is not limited to the specific details described in the above embodiments. Within the scope of the claims and technical concept of the present invention, various simple modifications and changes can be made to the technical solution of the present invention, and these simple modifications all fall within the protection scope of the present invention.

Claims

1. A method for preparing a zirconium-aluminum osteo-promoting electroactive thin film modified by magnetron sputtering, characterized in that, The process includes the following steps: cleaning and drying a PVDF film; fixing the PVDF film onto a glass slide and placing it in a magnetron sputtering chamber; depositing elemental zirconium and aluminum onto the PVDF film using magnetron sputtering to prepare the zirconium-aluminum osteoactive film; wherein the zirconium-aluminum osteoactive film contains 46.15-46.35 wt% zirconium, 19.25-19.45 wt% oxygen, 19.25-19.45 wt% fluorine, and 1.77-1.97 wt% aluminum. The magnetron sputtering method is as follows: under an inert atmosphere, the gas pressure in the magnetron sputtering chamber is 0.5-5 Pa, the sputtering distance is 4-10 cm, the sputtering power is maintained at 50-100 W, the substrate temperature is room temperature to 300 ℃, the argon flow rate is 5-80 sccm, and the duration is 10-20 min.

2. The method for preparing the magnetron sputtered modified zirconium-aluminum osteo-promoting electroactive thin film according to claim 1, characterized in that, The zirconium-aluminum osteo-promoting electroactive film contains 46.25 wt% zirconium, 19.35 wt% oxygen, 19.35 wt% fluorine, and 1.87 wt% aluminum.

3. The method for preparing the magnetron sputtered modified zirconium-aluminum osteo-promoting electroactive thin film according to claim 1, characterized in that, The magnetron sputtering method is as follows: under an inert atmosphere, the gas pressure in the magnetron sputtering chamber is 0.5 Pa, the sputtering distance is 5 cm, the sputtering power is maintained at 50 W, the substrate temperature is 200 ℃, the argon flow rate is 80 sccm, and the duration is 20 min.

4. A zirconium-aluminum osteo-active film prepared by the preparation method according to any one of claims 1-3.

5. The application of the zirconium-aluminum osteo-electroactive thin film of claim 4 in the preparation of bone defect repair or tissue engineering materials.

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