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

Abstract

The invention discloses a preparation method and application of a zirconium-aluminum bone-promoting electroactive film modified by a magnetron sputtering technology, and relates to the technical field of bone electroactive film materials. The method comprises the following steps: cleaning and blow-drying a PVDF film; and fixing the PVDF film on the glass slide, putting the glass slide into a magnetron sputtering chamber, and depositing a zirconium simple substance and an aluminum simple substance on the PVDF film in a magnetron sputtering manner to prepare the zirconium-aluminum bone-promoting electroactive film modified by the magnetron sputtering technology, in the zirconium-aluminum bone-promoting electroactive film modified by the magnetron sputtering technology, the content of a zirconium element is 46.15 to 46.35 wt%, the content of an oxygen element is 19.25 to 19.45 wt%, the content of a fluorine element is 19.25 to 19.45 wt%, and the content of an aluminum element is 1.77 to 1.97 wt%. The prepared film material has excellent biocompatibility, antibacterial property and osteogenesis induction capability, and the application scene of the film material in the oral cavity and other parts is expanded.

Description

Technical Field

[0001] The invention relates to the technical field of bone electroactive film materials, and in particular to a method for preparing a zirconium-aluminum bone-promoting electroactive film modified by magnetron sputtering technology. Background Art

[0002] Traditional bone tissue engineering is to repair bone defects by inoculating seed cells on scaffold materials, but there are defects such as seed cell apoptosis and immune inflammatory response, which will weaken the repair effect. To solve this problem, some scholars have proposed the concept of "in situ tissue engineering" to construct materials with high osteogenic response, and accelerate the repair process of the defect site through the body's regenerative potential. This method focuses on using material properties to induce endogenous stem cells to the site of injury and mediate osteogenic differentiation to achieve the goal of bone repair. The key is to simulate the natural extracellular physiological regeneration microenvironment of bone. Electroactive materials refer to a class of materials that can generate electrical signals under external stimulation, or can change their physical and chemical properties under electrical signal stimulation. At present, a series of electroactive biomaterials including ferroelectric, piezoelectric, electret and conductive materials and their mediated electrostimulation materials have been developed to simulate the natural physiological electrical microenvironment as a biophysical clue to regulate stem cell fate and regenerative medicine. Electroactive materials are a type of bone repair material that effectively constructs a regenerative electrical microenvironment.

[0003] When PVDF (polyvinylidene fluoride polymer) piezoelectric film is subjected to external force, polarized charges similar to the piezoelectric properties of natural bone tissue will appear on its surface, which is the piezoelectric effect. PVDF bone electroactive film has a wide range of applications in bone tissue engineering, bone repair, biosensors, drug delivery, etc. due to its excellent piezoelectric properties and large specific surface area. However, PVDF bone electroactive film still has problems such as poor electrical signal stability, poor bone regeneration regulation, and slow bone healing in practical applications. Therefore, how to prepare a PVDF bone electroactive film with good biocompatibility and antibacterial activity, which can exist stably in the body and regulate bone regeneration, has become a hot research direction for the application of PVDF materials in electroactive bone repair materials. Summary of the invention

[0004] The present invention provides a preparation method and application of a zirconium aluminum bone-promoting electroactive film modified by magnetron sputtering technology. The prepared film material has excellent biocompatibility, antibacterial properties and bone-inducing ability, which expands its application scenarios in the oral cavity and other parts, and effectively solves the problems of poor bone integration, insufficient bone healing speed and effectiveness of existing PVDF bone electroactive films. It is specifically achieved through the following technologies.

[0005] A method for preparing a zirconium-aluminum bone-promoting electroactive film modified by magnetron sputtering technology, comprising the following steps: washing and drying a PVDF film; fixing the PVDF film on a glass slide and placing the film in a magnetron sputtering chamber, and depositing zirconium and aluminum on the PVDF film by magnetron sputtering to prepare the zirconium-aluminum bone-promoting electroactive film; in the zirconium-aluminum bone-promoting electroactive film, the zirconium content, the oxygen content, the fluorine content, and the aluminum content are 19.25-19.45 wt%, 1.77-1.97 wt%.

[0006] Furthermore, in the zirconium-aluminum bone-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 method is: under an inert atmosphere, the gas pressure in the magnetron sputtering chamber is 0.5-5Pa, the sputtering distance is 4-10 cm, the sputtering power is maintained at 0-100 W, the substrate temperature is room temperature-300 ° C, the argon gas flow rate is 5-80 sccm, and the duration is 10-20 min.

[0008] Furthermore, the magnetron sputtering method is: 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 ° C, the argon gas flow rate is 80 sccm, and the duration is 20 min.

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

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

[0011] Compared with the prior art, the present invention is beneficial in that:

[0012] 1. Compared with the existing common PVDF bone electroactive film, the present invention effectively modifies the surface of the PVDF bone electroactive film with simple 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 exerting excellent antibacterial activity.

[0013] 2. Compared with the existing common PVDF bone electroactive film, the zirconium aluminum bone-promoting electroactive film prepared by the present invention can promote osteoblasts to express RUNX family transcription factor 2 (RUNX2), indicating that it has good osteoinduction activity and has very good bone bonding and bone repair effects. BRIEF DESCRIPTION OF THE DRAWINGS

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

[0015] Figure 2 This is the overall imaging of the prepared PVDF bone electroactive film in Example 1 using a scanning electron microscope at a magnification of 10,000 times.

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

[0017] Figure 4 This is a graph showing the relative mRNA expression of CD86 after culturing macrophages RAW264.7 using the modified zirconium-aluminum bone-promoting electroactive film in Example 1 and the PVDF bone-promoting electroactive film treated in Comparative Example 1.

[0018] Figure 5 This is a graph showing the relative mRNA expression of IL-1β after culturing macrophages RAW264.7 using the modified zirconium-aluminum bone-promoting electroactive film in Example 1 and the PVDF bone-promoting electroactive film treated in Comparative Example 1.

[0019] Figure 6 This is a graph showing the relative mRNA expression of IRF3 after culturing macrophages RAW264.7 using the modified zirconium-aluminum bone-promoting electroactive film in Example 1 and the PVDF bone-promoting electroactive film treated in Comparative Example 1.

[0020] Figure 7 This is a graph showing the relative mRNA expression of TRAF6 after culturing macrophages RAW264.7 using the modified zirconium-aluminum bone-promoting electroactive film in Example 1 and the PVDF bone-promoting electroactive film treated in Comparative Example 1.

[0021] Figure 8 This is a graph showing the relative mRNA expression of NF-κB after culturing macrophages RAW264.7 using the modified zirconium-aluminum bone-promoting electroactive film in Example 1 and the PVDF bone-promoting electroactive film treated in Comparative Example 1.

[0022] Fig. 9This is a graph showing the relative mRNA expression of RUNX2 after osteoblasts mt-3t3 were cultured using the modified zirconium-aluminum bone-promoting electroactive film in Example 1 and the PVDF bone-promoting electroactive film treated in Comparative Example 1.

[0023] Figure 10-13 The relative mRNA expression diagrams of IL-6, NF-κB, MYD88 and TRAF6 after culturing macrophages RAW264.7 with the bone-promoting electroactive films of Example 1 and Comparative Examples 2 and 3, respectively. DETAILED DESCRIPTION

[0024] The technical solution of the present invention will be described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0025] Example 1

[0026] The zirconium-aluminum bone-promoting electroactive film of this embodiment is Figure 1 As shown, it is prepared by the following preparation method:

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

[0028] (2) Adhere conductive glue to a glass slide, adhere the PVDF film processed in S1 to the conductive glue, and place the glass slide and the PVDF film upside down in a magnetron sputtering chamber.

[0029] (3) Zirconium and aluminum were installed as target materials in the DC target area of ​​magnetron sputtering. After closing the chamber, high-purity argon gas was introduced to keep the gas pressure in the magnetron sputtering chamber at 5 Pa. Zirconium and aluminum were deposited on the PVDF bone electroactive film under argon atmosphere. The sputtering distance was 10 cm, the sputtering power was maintained at 100 W, the substrate temperature was 300 °C, the argon gas flow rate was 5 sccm, and the duration was 20 min. Finally, a modified zirconium-aluminum bone-promoting electroactive film was obtained.

[0030] The whole zirconium-aluminum bone-promoting electroactive film prepared in this example was imaged using a scanning electron microscope at a magnification of 10,000 times. Figure 2 As shown in Figure 2, it can be seen that the surface of the modified PVDF bone electroactive film is smoother.

[0031] Then, the surface elements of the zirconium-aluminum bone-promoting electroactive film were scanned using the energy spectrum scanning module. The results are as follows: Figure 3 As shown in Table 1, it can be seen that stable content of single zirconium and aluminum is generated. Among them, the zirconium content is 46.25 wt%, the oxygen content is 32.53 wt%, the fluorine content is 19.35 wt%, and the aluminum content is 1.87 wt%.

[0032] Table 1 Spectrum scanning results

[0033] Example 2

[0034] The zirconium-aluminum bone-promoting electroactive film of this embodiment is prepared by the following preparation method:

[0035] (1) Cut the PVDF film into 2 cm × 2 cm square films, clean the PVDF film with anhydrous ethanol by ultrasonic cleaning for 15 min, blow dry with nitrogen, and store in a vacuum.

[0036] (2) Adhere conductive glue to a glass slide, adhere the PVDF film processed in S1 to the conductive glue, and place the glass slide and the PVDF film upside down in a magnetron sputtering chamber.

[0037] (3) Install zirconium and aluminum as target materials in the DC target area of ​​magnetron sputtering. After closing the chamber, introduce high-purity argon gas to keep the gas pressure in the magnetron sputtering chamber at 0.5 Pa, and deposit zirconium and aluminum on the PVDF bone electroactive film under argon atmosphere. The sputtering distance is 10 cm, the sputtering power is maintained at 100 W, the substrate temperature is room temperature, the argon gas flow rate is 5 sccm, and the duration is 30 min. Finally, a modified zirconium-aluminum bone-promoting electroactive film is obtained.

[0038] Example 3

[0039] The zirconium-aluminum bone-promoting electroactive film of this embodiment is prepared by the following preparation method:

[0040] (1) Cut the PVDF film into 2 cm × 2 cm square films, clean the PVDF film with anhydrous ethanol by ultrasonic cleaning for 15 min, blow dry with nitrogen, and store in a vacuum.

[0041] (2) Adhere conductive glue to a glass slide, adhere the PVDF film processed in S1 to the conductive glue, and place the glass slide and the PVDF film upside down in a magnetron sputtering chamber.

[0042] (3) Install zirconium and aluminum as target materials in the DC target area of ​​magnetron sputtering. After closing the chamber, introduce high-purity argon gas to keep the gas pressure in the magnetron sputtering chamber at 0.5 Pa, and deposit zirconium and aluminum on the PVDF bone electroactive film under argon atmosphere. The sputtering distance is 5 cm, the sputtering power is maintained at 50 W, the substrate temperature is 200 ° C, the argon flow rate is 80 sccm, and the duration is 20 min. Finally, a modified zirconium aluminum bone-promoting electroactive film is obtained.

[0043] Comparative Example 1

[0044] The film material of this comparative example was prepared by the following preparation method: the PVDF film was cut into square films of 2 cm×2 cm, the PVDF film was ultrasonically cleaned with anhydrous ethanol for 15 min, dried with nitrogen and stored in a vacuum to obtain the final film material.

[0045] Comparative Example 2

[0046] The difference between the preparation method of the thin film material of this comparative example and that of Example 1 is that only zirconium is installed as a target material in the DC target area of ​​magnetron sputtering, and zirconium is doped into the PVDF film; the zirconium content on the surface of the thin film material is about 48.12%, which is the sum of the zirconium and aluminum contents in Example 1. No aluminum or any other metal / non-metal material is doped.

[0047] Comparative Example 3

[0048] The difference between the preparation method of the thin film material of this comparative example and that of Example 1 is that only aluminum is installed as a target material in the DC target area of ​​magnetron sputtering, and the aluminum is doped into the PVDF film; the aluminum content on the surface of the thin film material is about 48.12%, which is the sum of the zirconium and aluminum contents in Example 1. No zirconium or any other metal / non-metal material is doped.

[0049] Test Example 1: Performance test of the film materials prepared in Example 1 and Comparative Example 1

[0050] 1. Antibacterial properties

[0051] The zirconium-aluminum bone-promoting electroactive films prepared in Example 1 and Comparative Example 1 were soaked in alcohol, sterilized by ultraviolet irradiation, placed in a six-well cell culture plate, and inoculated with 50% macrophage RAW264.7. Two days later, when the 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 drawing were performed using GraphPad Prism software, and the results are shown in Figure 2. Figure 4-8 As shown. It can be seen that the PVDF bone electroactive film modified by zirconium and aluminum in Example 1 can promote 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 makes them have stronger antibacterial properties. In contrast, the antibacterial properties of the ordinary PVDF film in Comparative Example 1 are very limited.

[0052] The bone-promoting electroactive films prepared in Example 1 and Comparative Examples 2 and 3 were soaked in alcohol, and the same method was used to finally detect the expression of interleukin-6 (IL-6), nuclear factor receptor-κB (NF-κB), MYD88 and TNF receptor-associated factor 6 (TRAF6), and statistical analysis and drawing were performed using ImageJ software. The results are shown in Figure 10-13 As shown in the figure, the bone-promoting electroactive film doped with 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 makes them have stronger antibacterial properties.

[0053] 2. Evaluation of osteogenic activity

[0054] The thin film materials prepared in Example 1 and Comparative Example 1 were soaked in alcohol, sterilized by ultraviolet irradiation, and placed in six-well cell culture plates; osteoblasts mt-3t3 were inoculated at a density of 50%; two days later, when the density increased to 80%, the cells were collected for qPCR experiments to detect the expression of RUNX family transcription factor 2 (RUNX2), and statistical analysis and drawing were performed using GraphPad Prism software. The results are shown in Figure 2. Fig. 9 As shown, the PVDF bone electroactive film modified by zirconium and aluminum in Example 1 can upregulate the expression of osteoblast RUNX2 transcription factor, indicating that it has better osteogenic activity.

[0055] The above specific embodiments describe the implementation of the present invention in detail, but the present invention is not limited to the specific details in the above embodiments. Within the scope of the claims and technical concept of the present invention, the technical solution of the present invention can be modified and changed in many simple ways, and these simple modifications all belong to the protection scope of the present invention.

Claims

1. A method for preparing a zirconium-aluminum bone-promoting electroactive film modified by magnetron sputtering technology, characterized in that: The method comprises the following steps: washing and drying a PVDF film; fixing the PVDF film on the glass slide and placing the film in a magnetron sputtering chamber, and depositing zirconium and aluminum on the PVDF film by magnetron sputtering to prepare the zirconium-aluminum bone-promoting electroactive film; in the zirconium-aluminum bone-promoting electroactive film, the zirconium content, the oxygen content, the fluorine content, and the aluminum content are 19.25-19.45 wt%, 1.77-1.97 wt%, respectively.

2. The method for preparing the zirconium-aluminum bone-promoting electroactive film modified by magnetron sputtering technology according to claim 1, characterized in that: In the zirconium-aluminum bone-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 %.

3. The method for preparing the zirconium-aluminum bone-promoting electroactive film modified by magnetron sputtering technology according to claim 1, characterized in that: The magnetron sputtering method is: under an inert atmosphere, the gas pressure in the magnetron sputtering chamber is 0.5-5Pa, the sputtering distance is 4-10 cm, the sputtering power is maintained at 50-100 W, the substrate temperature is room temperature-300°C, the argon gas flow rate is 5-80sccm, and the duration is 10-20 min.

4. The method for preparing the zirconium-aluminum bone-promoting electroactive film modified by magnetron sputtering technology according to claim 1, characterized in that: The magnetron sputtering method is: 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 ° C, the argon gas flow rate is 80 sccm, and the duration is 20 min.

5. A zirconium-aluminum bone-promoting electroactive film prepared by the preparation method described in any one of claims 1 to 4.

6. Use of the zirconium-aluminum bone-promoting electroactive film according to claim 5 in the preparation of bone defect repair or tissue engineering materials.

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