Preparation method and application of selective antibacterial heterojunction coating

Abstract

The invention relates to a preparation method and application of a selective antibacterial heterojunction coating. The selective antibacterial heterojunction coating comprises a metal titanium substrate and a selective antibacterial heterojunction film which is arranged on the surface of the metal titanium substrate and is formed by interaction of mixed metal oxide and noble metal particles, the mixed metal oxide includes a combination of a divalent metal oxide and a trivalent metal oxide.

Description

Technical Field

[0001] The present invention belongs to the technical field of surface modification of metal implant materials, and particularly relates to a preparation method and application of a selective antibacterial heterojunction coating. Background Art

[0002] Titanium is an inert metal with excellent physical and chemical properties, having advantages such as high strength, fatigue resistance, corrosion resistance, and high biocompatibility. It has been widely used as an implant material for hard tissue repair and replacement, and is mostly used for reconstructing bone defects caused by external force damage, disease invasion, etc.

[0003] Implant - associated infections (IAIs) are a serious postoperative complication. Even a small amount of bacteria can form a biofilm on the implant surface, which helps bacteria evade host defenses and antibiotic chemotherapy. Therefore, secondary surgery for debridement is usually required, and it may even ultimately lead to implant failure. Currently, there have been many studies on implant - associated infections. For example: loading a releasable antibacterial agent in a metal substrate, a physical antibacterial structure that can pierce bacteria, inducing thermal damage, generating catalytically active oxygen, and electrical stimulation, etc. However, the mechanisms by which these functionalized surfaces produce antibacterial effects also cause damage to normal cells. Designing an intelligent surface that can effectively kill bacteria without damaging cells remains a challenge.

[0004] By designing an advanced heterojunction structure, the manipulation of charge transfer at the interface can be achieved by using the potential difference between energy bands and the built - in electric field. Research results in recent years have confirmed that some heterojunctions with suitable energy band structures can capture protons in solution or transfer electrons on the surface of the bacterial membrane. However, the above - mentioned material systems not only have poor biocompatibility, but also usually can only manipulate one of proton or electron transfer. The resulting metabolic reprogramming makes it difficult to completely kill bacteria. On the other hand, combining electron capture or proton consumption with thermo - kinetic effects generated by external field stimuli such as light / acoustic can achieve better synergistic antibacterial performance. However, the high temperatures and reactive oxygen species (ROS) generated during these processes will damage normal cells or tissues, thus making it difficult for patients to tolerate the side effects of the treatment. Summary of the Invention

[0005] In order to solve the defect that existing medical titanium implant materials do not have selective antibacterial properties, the purpose of the present invention is to provide a preparation method and application of a modified selective antibacterial heterojunction coating. By designing a selective antibacterial heterojunction on the titanium surface, it has the function of regulating surface proton and electron transfer, and has innovation and clinical application value.

[0006] In a first aspect, the present invention provides a selective antibacterial heterojunction coating, which comprises a titanium metal substrate and a selective antibacterial heterojunction thin film formed by the interaction of a mixed metal oxide and noble metal particles on the surface of the titanium metal substrate; The mixed metal oxide comprises a combination of a divalent metal oxide and a trivalent metal oxide.

[0007] Preferably, the divalent metal includes Mg, Co, Ni, Cu, Zn, and the trivalent metal includes Fe, Ga; Preferably, the molar ratio of the divalent metal oxide to the trivalent metal oxide in the mixed metal oxide is 1:4 - 4:1.

[0008] Preferably, the mixed metal oxide is attached to the titanium metal substrate in the form of vertically grown layered nanosheets to form a nanosheet array; Preferably, the thickness of the layered nanosheets is 5 - 50 nm, and the diameter is 10 - 1000 nm; More preferably, the thickness of the nanosheet array is 50 - 1000 nm.

[0009] Preferably, the noble metal particles include Au, Pt, Rh, Pd, Ag particles; Preferably, the particle size of the noble metal nanoparticles is 1 - 100 nm, more preferably 1 - 10 nm.

[0010] In a second aspect, the present invention provides a preparation method of the above selective antibacterial heterojunction coating, which comprises the following steps: hydrothermally treating the titanium metal substrate in a reaction solution containing trivalent metal ions and a reaction solution containing divalent metal ions respectively, and then performing a heat treatment to form a mixed metal oxide layer; then, attaching noble metal particles to the mixed metal oxide layer by physical vapor deposition to obtain the selective antibacterial heterojunction coating.

[0011] Preferably, the reaction solution containing trivalent metal ions further contains sodium sulfate and polyetherimide PEI; preferably, the concentration of trivalent metal ions in the reaction solution containing trivalent metal ions is 50 - 120 mM, the concentration of sodium sulfate is 0 - 500 mM, preferably 100 mM, and the concentration of polyetherimide PEI is 0 - 576 mg / L, preferably 288 mg / L; The concentration of divalent metal ions in the reaction solution containing divalent metal ions is 30 - 60 mM.

[0012] Preferably, the hydrothermal treatment carried out in the reaction solution containing trivalent metal ions is carried out at a temperature of 120-200 °C, preferably 160 °C; the time is 10-60 minutes, preferably 30-60 minutes.

[0013] Preferably, the hydrothermal treatment carried out in the reaction solution containing divalent metal ions is carried out at a temperature of 120-200 °C, preferably 160 °C; the time is 1-48 hours, preferably 12-24 hours.

[0014] Preferably, the heating rate of the heat treatment is 1-10 °C / minute, preferably 5 °C / minute; the temperature of the heat treatment does not exceed 800 °C, preferably 200-600 °C; the time of the heat treatment is 10 minutes to 2 hours.

[0015] Preferably, the target of the physical vapor deposition includes the noble metal, the current is 5-20 mA, the deposition time is ≥10 seconds and <300 seconds, preferably 30-120 seconds.

[0016] Beneficial effects (1) The metal implant surface-modified by the method provided by the present invention has good charge transfer function and selective antibacterial performance; (2) The surface-modified titanium implant of the present invention can utilize the heterojunction formed by the metal and the mixed metal oxide to control the proton and electron transfer in the microenvironment, which is beneficial to hindering the energy metabolism of bacteria, resulting in a lack of energy supply, and ultimately leading to the death of bacteria; (3) The present invention solves the problems of low bioactivity, susceptibility to bacterial infection, and poor selectivity of the existing alternative titanium implant materials for hard tissue repair, and can be widely applied to the field of metal medical devices related to bone tissue repair and replacement. Description of the drawings

[0017] Figure 1 (a) in is the surface and cross-section scanning electron microscope morphology photos of MgFeO-Ti prepared in Example 1, (b) is the 3D atomic force microscope morphology photo of MgFeO-Ti prepared in Example 1, (c) is the surface and cross-section scanning electron microscope morphology photos of Au / MgFeO-Ti prepared in Example 1, (d) is the 3D atomic force microscope morphology photo of Au / MgFeO-Ti prepared in Example 1; Figure 2 (a) and (b) in are the high-resolution transmission electron microscope photos of MgFeO-Ti prepared in Example 1 and the corresponding surface scanning photos of Fe, Mg, and O elements in the MgFe-MMO nanosheets, respectively. (c) and (d) are the high-resolution transmission electron microscope photos of Au / MgFeO-Ti prepared in Example 1 and the corresponding surface scanning photos of Au, Fe, Mg, and O elements in the Au / MgFe-MMO nanosheets, respectively; Figure 3 Figure (a) shows the high-resolution XPS spectra of Au 4f in MgFeO-Ti and Au / MgFeO-Ti prepared in Example 1, (b) shows the high-resolution XPS spectra of Fe 2p in MgFeO-Ti and Au / MgFeO-Ti prepared in Example 1, and (c) shows the high-resolution XPS spectra of Mg 1s in MgFeO-Ti and Au / MgFeO-Ti prepared in Example 1; Figure 4 Figure shows the photoluminescence spectra of MgFeO-Ti and Au / MgFeO-Ti in Example 1; Figure 5 Figure shows the changes in local microenvironment pH of Ti, Au-Ti, MgFeO-Ti and Au / MgFeO-Ti in Example 1; Figure 6 Figure (a) shows the colony plate counting photos of Ti, Au-Ti, MgFeO-Ti and Au / MgFeO-Ti in Example 1, (b) shows the corresponding antibacterial rate against Staphylococcus aureus, and (c) shows the corresponding antibacterial rate against Escherichia coli; Figure 7 Figure (a) shows the protein leakage results of Staphylococcus aureus on the surfaces of different samples in Example 1, (b) shows the protein leakage results of Escherichia coli on the surfaces of different samples in Example 1, (c) shows the intracellular reactive oxygen species results of Staphylococcus aureus on the surfaces of different samples in Example 1, and (d) shows the intracellular reactive oxygen species results of Escherichia coli on the surfaces of different samples in Example 1; Figure 8 Figure (a) shows the proliferation activity test results of osteoblasts on the sample surfaces at days 1, 4, and 7 in Example 1, (b) shows the live / dead staining results of osteoblasts on the sample surfaces in Example 1, (c) shows the proliferation activity test results of fibroblasts on the sample surfaces at days 1, 4, and 7 in Example 1, and (d) shows the live / dead staining results of fibroblasts on the sample surfaces in Example 1; Figure 9 Figure (a) shows the surface scanning electron microscopy morphology photos of Pt / MgFeO-Ti in Example 2, (b) shows the elemental surface scanning photos of Pt, Mg, Fe, Ti, and O in the corresponding Pt / MgFeO-MMO nanosheets, and (c) shows the XPS full spectra of Ti, Pt-Ti, and Pt / MgFeO-Ti in Example 2; Figure 10 Figure (a) shows the colony plate counting photos of Staphylococcus aureus of Ti, Pt-Ti, and Pt / MgFeO-Ti in Example 2, and (b) shows the corresponding antibacterial rate; Figure 11Colony plating count photos of Staphylococcus aureus for Ti, MgAlO-Ti, and Au / MgAlO-Ti in Comparative Example 1; Figure 12 In (a) is Au small / MgFeO-Ti and Au large / MgFeO-Ti scanning electron microscope photos, and in (b) are colony plating count photos of Staphylococcus aureus for Ti, Au small / MgFeO-Ti, and Au large / MgFeO-Ti; Figure 13 In (a) are scanning electron microscope photos of Au1 / MgFeO-Ti, Au2 / MgFeO-Ti, Au3 / MgFeO-Ti, and Au4 / MgFeO-Ti, and in (b) are colony plating count photos of Staphylococcus aureus for Ti, Au1 / MgFeO-Ti, Au2 / MgFeO-Ti, Au3 / MgFeO-Ti, and Au4 / MgFeO-Ti. Specific Embodiments

[0018] The present invention is further illustrated by the following embodiments. It should be understood that the following embodiments are only used to illustrate the present invention and do not limit the present invention.

[0019] First, the present invention provides a selective antibacterial heterojunction coating. Among them, the selective antibacterial heterojunction coating may include: a metal titanium substrate, and a selective antibacterial heterojunction thin film on the surface of the metal titanium substrate; the selective antibacterial heterojunction thin film includes a mixed metal oxide on the surface of the metal titanium substrate, and noble metal particles attached to the surface of the mixed metal oxide.

[0020] In some embodiments, the mixed metal oxide (MMO) may include a combination of divalent metal oxides and trivalent metal oxides; among them, divalent metals may include Mg, Co, Ni, Cu, Zn, and trivalent metals may include Fe, Ga, and the molar ratio of divalent metal oxides to trivalent metal oxides may be 1:4 - 4:1; preferably, the mixed metal oxide may include a magnesium-iron mixed metal oxide (MgFe-MMO).

[0021] Mg is a macronutrient in the human body, and Fe is a trace element with a relatively high content in the human body. Among them, the greater the proportion of the metal with stronger alkalinity (i.e., stronger metallicity), the stronger the ability of MMO to capture protons. For example, the higher the proportion of Mg in MgFe-MMO, the stronger its proton transport ability. However, too high a proportion of alkali metals will cause the MMO to degrade too quickly, which is not conducive to maintaining the stability of the heterojunction.

[0022] In some embodiments, the mixed metal oxide is attached to the titanium metal substrate in the form of vertically grown layered nanosheets to form a nanosheet array; preferably, the thickness of the layered nanosheets can be 5-50 nm, the diameter can be 10-1000 nm, and the thickness of the nanosheet array can be 50-1000 nm.

[0023] In some embodiments, the noble metal particles may include gold (Au), platinum (Pt), rhodium (Rh), palladium (Pd), silver (Ag) particles; preferably, the particle size of the noble metal nanoparticles can be 1-100 nm, more preferably 1-10 nm.

[0024] Among them, the type of noble metal element determines its work function. For example, gold (Au) is 5.1 eV and platinum (Pt) is 5.12 eV. The higher the work function compared to that of MMO, the higher the built-in electric field strength formed and the stronger the electron transport ability. In addition, the size and content of the noble metal particles will change their existence form on the surface of the heterojunction. When the size and content are too high, the formed particles will reduce the electron transport efficiency due to excessive thickness and at the same time hinder the contact between MMO and the external environment, weakening its ability to absorb protons.

[0025] In some embodiments, the loading amount of the noble metal particles can be 1-50 at.%, preferably 10 at.%.

[0026] In some embodiments, the work function of the mixed metal oxide can be controlled to be less than the work function of the noble metal particles.

[0027] The selective antibacterial heterojunction coating provided by the present invention has the ability to consume protons in the environment while transferring electrons. Specifically, after the noble metal particles come into contact with MMO, due to the higher work function of the noble metal, electrons flow from MMO to the noble metal, thereby establishing a built-in electric field pointing from the mixed metal oxide to the metal nanoparticles and forming a Schottky heterojunction. The array film composed of a large number of heterojunctions on the titanium surface can effectively conduct the hot electrons generated by the surface plasmon resonance effect in the noble metal into MMO, forming an electron flow that continuously consumes electrons. At the same time, MMO, as a basic substance, can act as a proton acceptor to capture protons in the environment and can consume protons through the hydrolysis of oxides in an aqueous solution, increasing the pH value of the solution. Therefore, the heterojunctions on the surface of the selective antibacterial heterojunction coating can affect the surface proton and electron transport simultaneously.

[0028] The proton transport ability of MMO is essentially a hydrolysis process of alkali metals, MeO + 2H3O + →Me 2++3H2O. Therefore, in MMO, the stronger the basicity (i.e., metallicity) of the metal, the higher the proportion of the oxide content of the alkali metal (usually divalent metal, such as Mg relative to Fe), the stronger its proton transport ability, the higher the degree of oxidation, the more complete the discharge of water molecules in LDH, and the stronger the hydrolysis ability, i.e., the proton transport ability. On the other hand, the electron transport ability depends on the difference in work functions of the two materials forming the heterojunction. The type of noble metal element determines the size of its work function. For example, gold (Au) is 5.1 eV and platinum (Pt) is 5.12 eV. The higher its work function compared to that of MMO, the higher the built-in electric field strength formed and the stronger the electron transport ability. In addition, the size and content of noble metal particles will change their existence form on the surface of the heterojunction. When the size and content are too high, the formed particles will reduce the electron transport efficiency due to excessive thickness and at the same time hinder the contact between MMO and the external environment, weakening its ability to absorb protons.

[0029] Furthermore, the charge transfer ability of the selective antibacterial heterojunction coating can capture protons and electrons in the bacterial membrane respiratory chain, disrupting various biological processes such as bacterial ATP synthesis and amino acid synthesis, resulting in the death of bacteria due to severe starvation. However, this function will not have an adverse impact on the adhesion, proliferation, and mitochondrial function of osteoblasts and fibroblasts because mammalian cells have a complex organelle structure. The mitochondria responsible for energy metabolism are located inside the cells, and the proton-coupled respiratory electron transport chain is located in the mitochondria inside the cells. Since the mitochondria inside the cells do not directly contact the extracellular environment, they will not be negatively affected by charge transfer.

[0030] The selective antibacterial heterojunction provided by the present invention is constructed by hydrothermal reaction and magnetron sputtering, endowing the surface of the inert titanium implant with unique charge transfer performance, bactericidal performance, and biological safety. When contacting bacteria, the material itself can simultaneously interfere with proton and electron transfer, showing good application prospects in medical titanium or titanium alloy devices in contact with bone tissue. Different from some of the technical solutions disclosed in patents, although a heterojunction with a built-in electric field is also constructed, not only is the construction method cumbersome and complex, requiring strict annealing in a reducing atmosphere to create oxygen vacancies, but also it requires external field stimulation to exert antibacterial function by forming an electric current.

[0031] Hereinafter, the preparation method of the selective antibacterial heterojunction coating provided by the present invention will be exemplarily described. Among them, the preparation method may include the following steps: Hydrothermally treat the metal titanium substrate in a reaction solution containing trivalent metal ions and a reaction solution containing divalent metal ions respectively, and then perform heat treatment to form a mixed metal oxide layer; then, attach noble metal particles on the mixed metal oxide layer by physical vapor deposition to obtain the selective antibacterial heterojunction coating.

[0032] In some embodiments, the titanium metal matrix may be pretreated before hydrothermal treatment; wherein, the pretreatment may include polishing, cleaning or sandblasting.

[0033] As an example, before hydrothermal treatment, the titanium sheet is subjected to surface acid and / or water cleaning; if the surface of the sample is clean enough, it can be ultrasonically cleaned only with deionized water; other acid treatments or polishing treatments can also be used to remove surface stains. Preferably, ultrasonic cleaning is performed successively with a mixed acid solution (volume ratio of HF:HNO3:H2O = 1:5:34) and deionized water; the number of cleaning times can be adjusted as needed, for example, cleaned twice.

[0034] In some embodiments, the reaction solution containing trivalent metal ions may further contain sodium sulfate and polyetherimide PEI; preferably, the concentration of trivalent metal ions in the reaction solution containing trivalent metal ions can be 50-120 mM, the concentration of sodium sulfate can be 0-500 mM (such as 100 mM), and the concentration of polyetherimide PEI can be 0-576 mg / L (such as 288 mg / L).

[0035] Among them, sodium sulfate can be used as an electrolyte to adjust the ionic strength of the reaction system, and the crystal growth rate can be adjusted by controlling the content of sodium sulfate to avoid the formation of irregular morphologies. Polyetherimide can be used as a polymer surfactant to adsorb on the surface of crystal nuclei to prevent agglomeration. By adjusting its content, not only the crystal growth rate can be adjusted, but also dispersion can be promoted to ensure uniform crystal morphology.

[0036] In some embodiments, the temperature of the hydrothermal treatment carried out in the reaction solution containing trivalent metal ions can be 120-200 °C, preferably 160 °C, the time can be 10-60 minutes, preferably 30-60 minutes; the filling degree of the hydrothermal treatment reaction kettle can be 40-80%. Too high reaction temperature will increase the pressure in the reaction kettle, bringing potential safety hazards; too low will slow down the reaction rate. Too long reaction time will cause excessive crystal growth and trigger its phase change; too short will make the reaction insufficient.

[0037] In some embodiments, the concentration of divalent metal ions in the reaction solution containing divalent metal ions can be 30-60 mM. Too high metal ion concentration will cause uneven dispersion in the solvent; too low will reduce the reaction efficiency.

[0038] In some embodiments, the temperature of the hydrothermal treatment carried out in the reaction solution containing divalent metal ions can be 120-200 °C, preferably 160 °C, and the time can be 1-48 hours, preferably 12-24 hours.

[0039] Among them, between the two hydrothermal treatments, the substrate can be cleaned with deionized water and the surface can be cleaned with an ultrasonic cleaner with a power of 40%.

[0040] It should be noted that the purpose of first performing the hydrothermal reaction of trivalent metal ions and then performing the hydrothermal reaction of divalent metal ions is to ensure that the trivalent metal ions can preferentially form a stable structural framework and provide sites for the insertion of divalent metal ions.

[0041] In some embodiments, the heating rate of the heat treatment can be 1-10 °C / min, preferably 5 °C / min. The temperature of the heat treatment can not exceed 800 °C, preferably 200-600 °C. The time of the heat treatment can be 10 minutes to 2 hours.

[0042] Through the heat treatment, the water molecules in the layered double hydroxide obtained by the hydrothermal reaction can be evaporated, and the hydroxide can be transformed into an oxide. The higher the heat treatment temperature, the more thorough the transformation, and the stronger its proton absorption ability and semiconductor characteristics. If the heat treatment temperature is too high, it will cause the phase transformation of the substrate titanium and reduce the service performance; if it is too low, the water molecules cannot be discharged, reducing its antibacterial performance.

[0043] In some embodiments, the physical vapor deposition is carried out in a magnetron sputtering instrument. The target includes the noble metal, the current is 5-20 mA, the deposition time is ≥10 seconds and <300 seconds, preferably 30-120 seconds.

[0044] Too large current and time will thicken the deposition layer and inhibit the contact between MMO and the external environment. Too small will not be able to construct a heterojunction that can fully exert antibacterial performance. Moreover, in the present invention, by counterintuitively reducing the working current and sputtering time, the purpose of controlling the deposition particle size is achieved, so that it stays in the island growth stage, thereby obtaining a noble metal modified layer with a nanoparticle morphology.

[0045] The selective antibacterial heterojunction coating obtained by the preparation method provided by the present invention can be applied to the manufacture of medical metal devices, especially in the preparation of medical metal devices or implants for hard tissue repair.

[0046] The following further gives examples to illustrate the present invention in detail. It should also be understood that the following examples are only used to further illustrate the present invention and cannot be construed as limiting the protection scope of the present invention. Those skilled in the art's non-essential improvements and adjustments based on the above content of the present invention all belong to the protection scope of the present invention. The specific process parameters and the like in the following examples are also only an example within a suitable range, that is, those skilled in the art can make selections within a suitable range through the description in this article, rather than being limited to the specific values in the following examples. If not specifically specified, the technical means used in the examples are conventional means well known to those skilled in the art.

[0047] Example 1

[0048] The preparation method of the selective antibacterial heterojunction coating provided in this example includes the following steps: (1) Pure titanium with lengths, widths, and heights of 10 mm, 10 mm, and 1 mm respectively is ultrasonically cleaned twice with a mixed acid (volume ratio of HF:HNO3:H2O = 1:5:34) and deionized water for 5 minutes each time, and is dried for later use, marked as Ti; On the surface of Ti, titanium modified with gold nanoparticles is prepared by magnetron sputtering. A pure gold target with a purity of 99.99% is installed, the sputtering current of the sputtering instrument is set to 10 mA, and the sputtering time is 60 s. The sample after preparation is marked as Au-Ti; (2) The titanium sheet is placed flat in a polytetrafluoroethylene hydrothermal reaction inner liner, and 50 mL of hydrothermal reaction solution is added. The solvent is pure water, and the solutes are 60 mM of ferric chloride, 100 mM of sodium sulfate, and 288 mg / L of PEI. The stainless steel reaction kettle body is tightly installed; the reaction kettle is heated to 160 °C in an oven, and the reaction time is 30 minutes. During this process, the filling degree of the reaction kettle is maintained at 40%; after the reaction, it is soaked in deionized water and the surface is cleaned with an ultrasonic cleaner with a power of 40% for 1 minute, repeated once, and then rinsed 3 times with deionized water, and the sample is dried; (3) The dried sample in step (2) is placed flat in a clean polytetrafluoroethylene hydrothermal reaction inner liner, and 50 mL of hydrothermal reaction solution is added. The solvent is pure water, and the solute is 30 mM of magnesium chloride. The reaction kettle is heated to 160 °C, and the reaction time is 24 hours. The filling degree is the same as above; after the reaction, it is cleaned with deionized water and dried; (4) The dried sample in step (3) is placed in a muffle furnace and heated to 250 °C at a rate of 5 °C per minute, held for 2 hours, and naturally cooled to room temperature with the furnace. The obtained sample is marked as MgFeO-Ti; (5) The MgFeO-Ti is placed in a magnetron sputtering instrument, a pure gold Au target with a purity of 99.99% is installed, the sputtering current of the sputtering instrument is set to 10 mA, and the sputtering time is 60 s to obtain the selective antibacterial heterojunction coating, denoted as Au / MgFeO-Ti.

[0049] Figure 1 (a) is the surface and cross-section scanning electron microscope morphology photos of MgFeO-Ti prepared in Example 1, (b) is the 3D atomic force microscope morphology photo of MgFeO-Ti prepared in Example 1, (c) is the surface and cross-section scanning electron microscope morphology photos of Au / MgFeO-Ti prepared in Example 1, and (d) is the 3D atomic force microscope morphology photo of Au / MgFeO-Ti prepared in Example 1. From Figure 1As can be seen from (a) and (b), the surface of Ti has been covered by an array film of uniformly grown magnesium-iron mixed metal oxide (MgFe-MMO) with a thickness of about 400 nm. As can be seen from (c) and (d), the surface of Ti is also covered by an array film of uniformly grown magnesium-iron mixed metal oxide (MgFe-MMO), and at the same time, dense nanoparticles are evenly distributed on the surface of the nanosheets.

[0050] Figure 2 Among them, (a) and (b) are the high-resolution transmission electron microscope photos of MgFeO-Ti prepared in Example 1 and the corresponding surface scanning photos of Fe, Mg, and O elements in the MgFe-MMO nanosheets, respectively. (c) and (d) are the high-resolution transmission electron microscope photos of Au / MgFeO-Ti prepared in Example 1 and the corresponding surface scanning photos of Au, Fe, Mg, and O elements in the Au / MgFe-MMO nanosheets, respectively. From the TEM results in Figure (a), the single-layered structure of the MgFe-MMO nanosheets can be seen. From the mapping results in Figure (b), the elemental composition of the modified layer is confirmed to be iron (Fe), magnesium (Mg), and oxygen (O). In addition to observing the gold nanoparticles distributed on the nanosheets in Figure (c) TEM, the lattice belonging to elemental gold (Au) other than MgFe-MMO is also confirmed. In the mapping results in Figure (d), four elements, namely iron (Fe), magnesium (Mg), oxygen (O), and gold (Au), are detected. The above results confirm that an array film of magnesium-iron mixed metal oxide (MgFe-MMO) has been successfully constructed on the surface of MgFeO-Ti in this example, and a heterostructure composed of an array film of magnesium-iron mixed metal oxide (MgFe-MMO) and gold nanoparticles has been successfully constructed on the surface of Au / MgFeO-Ti.

[0051] Figure 3Figure (a) shows the high-resolution XPS spectra of Au 4f in MgFeO-Ti and Au / MgFeO-Ti prepared in Example 1, figure (b) shows the high-resolution XPS spectra of Fe 2p in MgFeO-Ti and Au / MgFeO-Ti prepared in Example 1, and figure (c) shows the high-resolution XPS spectra of Mg 1s in MgFeO-Ti and Au / MgFeO-Ti prepared in Example 1. It can be seen from (a) that the peaks at 84.6 eV and 88.3 eV correspond to the characteristic peaks of Au 4f7 / 2 and Au 4f5 / 2, respectively. Compared with Au-Ti, the binding energy of Au in Au / MgFeO-Ti decreases, indicating that the Au nanoparticles in the Au / MgFe-MMO heterojunction obtain electrons from MgFe-MMO, that is, the introduction of MgFe-MMO reduces the valence state of Au in Au / MgFe MMO; it can be seen from (b) that the peaks at 711.9 and 725.6 eV correspond to the characteristic peaks of Fe 2p3 / 2 and Fe 2p1 / 2, and the binding energy of Fe in the Au / MgFe MMO film is higher than that of MgFe-MMO, indicating that the Fe element in Au / MgFe-MMO loses electrons; a similar phenomenon of increased binding energy of the Mg 1s characteristic peak can also be observed in (c). The above results show that in the Au / MgFe-MMO heterojunction, Mg and Fe lose electrons, while Au gains electrons, proving that interfacial electron transfer occurs between the MgFe-MMO nanosheets and Au nanoparticles, forming a built-in electric field in the heterojunction.

[0052] Figure 4 is the photoluminescence spectrum of MgFeO-Ti and Au / MgFeO-Ti in Example 1, Figure 5 is the change in the local microenvironment pH of Ti, Au-Ti, MgFeO-Ti and Au / MgFeO-Ti in Example 1. From Figure 4 it can be seen that under the stimulation of an excitation light with a wavelength of 300 nm, the emission fluorescence intensity of Au / MgFeO-Ti is stronger than that of MgFeO-Ti, which may be due to the surface plasmon resonance effect of gold Au nanoparticles. Under light illumination, the recombination of electron-hole pairs generated by MgFe-MMMO will produce partial fluorescence, while the gold Au nanoparticles in Au / MgFeO-Ti generate high-energy hot electrons through surface plasmon resonance, and these electrons then pass through the Schottky barrier and enter the conduction band of MgFe-MMO, thus enhancing the excitation light of MgFe MMO. This result shows that under specific excitation conditions, electron flow from gold nanoparticles to the MgFe MMO semiconductor can be achieved. From Figure 5It can be seen that the proton concentration on the surfaces of MgFeO-Ti and Au / MgFeO-Ti is significantly lower than that on Ti and Au-Ti, confirming the proton consumption ability of MgFe-MMO and AuMgFe-MMOs. Since the MgFe MMO nanosheets on Au / MgFeO-Ti are partially covered by Au nanoparticles, the contact area between protons and Au / MgFe MMO nanoparticles is reduced, which weakens their ability to consume protons. The above results confirm the successful construction of the Au / MgFe MMO Schottky heterojunction thin film and its ability to control electron and proton transfer.

[0053] Next, the samples prepared in Example 1 were tested for antibacterial activity as follows: 60 μL of Escherichia coli and Staphylococcus aureus suspensions with a density of 1×10 7 CFU / mL were respectively dropped onto the surfaces of the samples and cultured in an incubator at 37 °C for 12 h, followed by plating and counting.

[0054] Figure 6 (a) in shows the colony plating count photos of Ti, Au-Ti, MgFeO-Ti, and Au / MgFeO-Ti in Example 1, (b) shows the corresponding antibacterial rate against Staphylococcus aureus, and (c) shows the corresponding antibacterial rate against Escherichia coli. From the colony diagrams of Staphylococcus aureus and Escherichia coli on the surfaces of Ti, Au-Ti, MgFeO-Ti, and Au / MgFeO-Ti in (a) after shaking, dilution, and transplantation to agar plates for culture at the end of the culture, as well as the corresponding antibacterial rates of Staphylococcus aureus and Escherichia coli in (b) and (c), it can be seen that compared with Ti, the antibacterial effect of Au-Ti is hardly improved, the number of colonies in MgFeO-Ti is slightly reduced, and no colonies can be seen in the plating results of Au / MgFeO-Ti. The antibacterial rates of Au-Ti, MgFeO-Ti, and Au / MgFeO-Ti against Staphylococcus aureus are -20.882%, 27.787%, and 99.977% respectively, and the antibacterial rates against Escherichia coli are 11.416%, 44.615%, and 99.975% respectively. The above results prove that the heterojunction formed by Au and MgFe-MMO has spectral and efficient antibacterial ability.

[0055] Figure 7In (a), it is the protein leakage results of Staphylococcus aureus on different sample surfaces in Example 1. In (b), it is the protein leakage results of Escherichia coli on different sample surfaces in Example 1. In (c), it is the intracellular reactive oxygen species results of Staphylococcus aureus on different sample surfaces in Example 1. In (d), it is the intracellular reactive oxygen species results of Escherichia coli on different sample surfaces in Example 1. From the protein leakage results of Staphylococcus aureus and Escherichia coli bacteria cultured on the surfaces of Ti, Au-Ti, MgFeO-Ti, and Au / MgFeO-Ti in (a-b), it can be seen that the intracellular protein leakage on Au / MgFeO-Ti is significantly higher than that in the Ti, Au-Ti, and MgFeO-Ti groups. From the intracellular reactive oxygen species (ROS) results of Staphylococcus aureus and Escherichia coli bacteria cultured on the surfaces of Ti, Au-Ti, MgFeO-Ti, and Au / MgFeO-Ti in (c-d), it can be seen that the ROS level of bacteria on Au / MgFeO-Ti is more than 10 times higher than that of other groups. ROS has strong oxidizing properties and can damage the membrane structure, proteins, and NDA substances of bacteria. The above results indicate that the bacteria on the Au / MgFe MMO heterojunction thin film have their energy metabolism disrupted by proton and electron transfer, thereby generating severe oxidative stress, leading to the rupture of their membrane structure and the leakage of cytoplasmic proteins and other substances.

[0056] Next, mouse osteoblast MC3T3-E1 and mouse fibroblast L929 were cultured in vitro to evaluate the effects of the samples obtained by the modification treatment in Example 1 on the proliferation and survival of normal cells. The specific method is as follows: 1) The samples were sterilized with 75% alcohol for 2 hours (changing the liquid every 0.5 hours), placed in a 24-well cell culture plate (with the modified side facing up), and the cell suspension was added. 2) The cell culture plate was placed in a cell culture incubator with 5% CO2 and saturated humidity and cultured at 36.5 °C. 3) After culturing for 1 day, 4 days, and 7 days, 4 samples were taken from each group, the original culture medium was aspirated, and new culture medium containing 10% AlamarBlue dye was added. After culturing the culture plate in the incubator for 2 hours, it was pipetted evenly, and 100 μL of the culture medium was taken from each well and placed in a black 96-well plate. 4) Test using an enzyme-linked immunosorbent assay (ELISA) reader. 5) After culturing for 7 days, 1 sample was taken from each group, rinsed twice with PBS buffer for 10 minutes each time, then the cell viability dye was added, incubated in an oven at 37 °C for 30 minutes, and then observed and photographed using a fluorescence microscope.

[0057] Figure 8(a) shows the results of the proliferation activity test of osteoblasts on the surface of the samples in Example 1 on the 1st, 4th, and 7th days. Compared with Ti, Au-Ti has a certain inhibitory effect on cell proliferation, while the proliferation activity on the surfaces of MgFeO-Ti and Au / MgFeO-Ti is significantly increased. This may be because the alkaline microenvironment formed after MMO absorbs protons promotes the proliferation of osteoblasts. (b) shows the live-dead staining results of osteoblasts on the surface of the samples in Example 1. A large number of live cells can be observed in each group, while dead cells are hardly visible. (c) shows the results of the proliferation activity test of fibroblasts on the surface of the samples in Example 1 on the 1st, 4th, and 7th days. Compared with Ti, there is little difference in the cell proliferation activity on the surfaces of Au-Ti, MgFeO-Ti, and Au / MgFeO-Ti. (d) shows the live-dead staining results of fibroblasts on the surface of the samples in Example 1. Similar to osteoblasts, fibroblasts also survive in large numbers on the surfaces of each group of samples. The above results indicate that the Au / MgFe MMO heterojunction film constructed on the titanium surface has good cytocompatibility with both osteoblasts and fibroblasts.

[0058] Example 2

[0059] The preparation method of the selective antibacterial heterojunction coating provided in this example refers to Example 1. The main difference is that the gold Au target in the magnetron sputtering step is replaced with a platinum Pt target; the corresponding obtained samples are respectively labeled as Ti, Pt-Ti, and Pt / MgFeO-Ti.

[0060] Figure 9 (a) shows the scanning electron microscope (SEM) topographic photo of the surface of Pt / MgFeO-Ti in Example 2, (b) shows the elemental surface scanning photos of Pt, Mg, Fe, Ti, and O in the corresponding Pt / MgFeO-MMO nanosheets, and (c) shows the XPS full spectra of Ti, Pt-Ti, and Pt / MgFeO-Ti in Example 2. It can be seen from the SEM photo of Pt / MgFeO-Ti in (a) that fine Pt nanoparticles can be seen on the surface of the MgFe-MM nanosheets. It can be seen from the mapping results of Pt / MgFeO-Ti in (b) that the elements Pt, Mg, Fe, and O are detected on the surface of the sample. It can be seen from the XPS test results of Ti, Pt-Ti, and Pt / MgFeO-Ti in (c) that the characteristic peaks of Pt4f, Fe 2p, and Mg 1s. The above results prove that the Pt / MgFe-MMO heterojunction has been successfully prepared on the titanium surface.

[0061] Next, the antibacterial test is carried out on the samples prepared in Example 2: 60 μL of a density of 1×10 7E. coli and S. aureus bacterial suspensions at CFU / mL were cultured in a constant temperature incubator at 37°C for 12 h and then plated for counting.

[0062] Figure 10 Among them, (a) is the photo of the S. aureus colony plate counting of Ti, Pt-Ti, and Pt / MgFeO-Ti in Example 2, and (b) is the corresponding antibacterial rate. It can be seen from the figure that the combination of platinum Pt and MgFe-MMO also has a significant synergistic antibacterial effect on S. aureus and E. coli.

[0063] Comparative Example 1

[0064] The preparation method of the coating provided in this comparative example refers to Example 1, and the main difference is that: In step (2), the trivalent ion is aluminum Al 3+ , and the obtained samples were labeled as MgAlO-Ti and Au / MgAlO-Ti.

[0065] Figure 11 is the photo of the S. aureus colony plate counting of Ti, MgAlO-Ti, and Au / MgAlO-Ti in Comparative Example 1. It can be seen from the figure that when the trivalent metal oxide is changed to Al2O3, the Au / MgAlO heterojunction constructed on the titanium surface also has antibacterial effect, but the antibacterial rate is lower than that of Au / MgFeO.

[0066] Comparative Example 2

[0067] The preparation method of the coating provided in this comparative example refers to Example 1, and the main difference is that: In step (5), the particle size of Au particles was controlled to be >100 nm by adjusting the magnetron sputtering parameters, and it was labeled as Au large / MgFeO-Ti, and Au / MgFeO-Ti in Example 1 was further labeled as Au small / MgFeO-Ti.

[0068] Figure 12 Among them, (a) is the SEM photo of Au small / MgFeO-Ti and Au large / MgFeO-Ti, and (b) is the photo of the S. aureus colony plate counting of Ti, Au small / MgFeO-Ti, and Au large / MgFeO-Ti. It can be seen from the figure that when the size of the noble metal particles on the surface is too large, the antibacterial effect decreases accordingly.

[0069] Comparative Example 3

[0070] The preparation method of the coating provided in this comparative example refers to Example 1, and the main difference is that in step (5), during the magnetron sputtering process, the sputtering time is gradually increased from 30 s to 60 s, 120 s, and 300 s, which are respectively marked as Au1 / MgFeO-Ti, Au2 / MgFeO-Ti, Au3 / MgFeO-Ti, and Au4 / MgFeO-Ti.

[0071] Figure 13 In (a) are the scanning electron microscope photos of Au1 / MgFeO-Ti, Au2 / MgFeO-Ti, Au3 / MgFeO-Ti, and Au4 / MgFeO-Ti, and in (b) are the photos of the plate count of Staphylococcus aureus colonies of Ti, Au1 / MgFeO-Ti, Au2 / MgFeO-Ti, Au3 / MgFeO-Ti, and Au4 / MgFeO-Ti. It can be seen from the figure that as the sputtering time prolongs, the thickness of the gold Au deposition layer on the surface of MgFeO increases accordingly. However, when the sputtering time is too long (300 s), its antibacterial rate decreases.

[0072] Although the content of the present invention has been introduced in detail through the above preferred embodiments, it should be recognized that the above description should not be considered as a limitation of the present invention. After those skilled in the art read the above content, various modifications and alternatives to the present invention will be obvious. Therefore, the protection scope of the present invention should be defined by the appended claims.

Claims

1. A selective antibacterial heterojunction coating, characterized in that, The selective antibacterial heterojunction coating includes: a titanium metal substrate, and a selective antibacterial heterojunction thin film formed by the interaction of a mixed metal oxide and noble metal particles on the surface of the titanium metal substrate; The mixed metal oxide includes a combination of a divalent metal oxide and a trivalent metal oxide.

2. The selective antibacterial heterojunction coating according to claim 1, wherein The divalent metals include Mg, Co, Ni, Cu, Zn, and the trivalent metals include Fe, Ga; Preferably, the molar ratio of the divalent metal oxide to the trivalent metal oxide in the mixed metal oxide is 1:4 - 4:

1.

3. The selective antibacterial heterojunction coating according to claim 1 or 2, characterized in that, The mixed metal oxide adheres to the titanium metal substrate in the form of vertically grown layered nanosheets to form a nanosheet array; Preferably, the thickness of the layered nanosheets is 5 - 50 nm, and the diameter is 10 - 1000 nm; More preferably, the thickness of the nanosheet array is 50 - 1000 nm.

4. The selective antibacterial heterojunction coating according to any one of claims 1-3, characterized in that, The noble metal particles include Au, Pt, Rh, Pd, Ag particles; Preferably, the particle size of the noble metal nanoparticles is 1 - 100 nm, more preferably 1 - 10 nm.

5. A method for preparing the selective antibacterial heterojunction coating according to any one of claims 1-4, characterized in that, The preparation method includes the following steps: subjecting the titanium metal substrate to hydrothermal treatment in a reaction solution containing trivalent metal ions and a reaction solution containing divalent metal ions respectively, and then performing heat treatment to form a mixed metal oxide layer; then, attaching noble metal particles to the mixed metal oxide layer by physical vapor deposition to obtain the selective antibacterial heterojunction coating.

6. The preparation method according to claim 5, characterized in that, The reaction solution containing trivalent metal ions further contains sodium sulfate and polyetherimide PEI; preferably, the concentration of trivalent metal ions in the reaction solution containing trivalent metal ions is 50 - 120 mM, the concentration of sodium sulfate is 0 - 500 mM, preferably 100 mM, and the concentration of polyetherimide PEI is 0 - 576 mg / L, preferably 288 mg / L; The concentration of divalent metal ions in the reaction solution containing divalent metal ions is 30 - 60 mM.

7. The preparation method according to claim 5 or 6, characterized in that, The temperature of the hydrothermal treatment in the reaction solution containing trivalent metal ions is 120 - 200 °C, preferably 160 °C; the time is 10 - 60 minutes, preferably 30 - 60 minutes.

8. The preparation method according to any one of claims 5-7, characterized in that The temperature of the hydrothermal treatment in the reaction solution containing divalent metal ions is 120 - 200 °C, preferably 160 °C; the time is 1 - 48 hours, preferably 12 - 24 hours.

9. The preparation method according to any one of claims 5-8, characterized in that, The heating rate of the heat treatment is 1 - 10 °C / minute, preferably 5 °C / minute; the temperature of the heat treatment does not exceed 800 °C, preferably 200 - 600 °C; the time of the heat treatment is 10 minutes to 2 hours.

10. The preparation method according to any one of claims 5-9, characterized in that, The target material for physical vapor deposition includes the noble metal, the current is 5 - 20 mA, the deposition time is ≥10 seconds and <300 seconds, preferably 30 - 120 seconds.

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