CN112190767A

CN112190767A - Nano-antibacterial coating material based on nanogold cluster and preparation method thereof

Info

Publication number: CN112190767A
Application number: CN202011003783.0A
Authority: CN
Inventors: 陈云丰; 褚光宇; 张春雷; 崔大祥
Original assignee: Shanghai Sixth Peoples Hospital
Current assignee: Shanghai Sixth Peoples Hospital
Priority date: 2020-09-22
Filing date: 2020-09-22
Publication date: 2021-01-08
Anticipated expiration: 2040-09-22
Also published as: CN112190767B

Abstract

The invention provides a nano-antibacterial coating material based on a nano-gold cluster and a preparation method thereof. The material takes a pMBA-AuNCs cluster as a basic constitutional unit, and then a solution is acidified to obtain a pMBA-AuNCs cluster methanol solution with completely protonated ligand terminal carboxyl; adding a salt solution of IVB group quadrivalent ions to obtain a mixed solution; finally adding Cu into the mixed solution²⁺The solution is used for obtaining the nano antibacterial coating material. The nano antibacterial coating material based on the nanogold cluster regulates the property of the cluster surface ligand, the surface ligand and metal ions form a three-dimensional periodic net-shaped film structure through self-assembly on the surface of a plant in metal through coordination, excellent in-vivo and in-vitro anti-biofilm activity is shown, an integrated treatment concept of preventing a biofilm and controlling infection is provided, and the method is an imminent treatment concept of the integrated treatment of biofilm prevention and infection controlThe bed treatment of the implant-related infection provides a new idea.

Description

Nano-antibacterial coating material based on nanogold cluster and preparation method thereof

Technical Field

The invention relates to the field of metal implant antibacterial coatings, in particular to a nano antibacterial coating material based on a nano gold cluster and a preparation method thereof.

Background

The biofilm (biofilm) is a microbial cell aggregate formed by bacteria adsorbed on the surface of a biological material or a body cavity and wrapped by an Extracellular Polymeric Substrate (EPS). The attachment of microorganisms on the surface of a material and the formation of a biofilm are complex dynamic processes involving a variety of physiological mechanisms, which are influenced by a variety of factors. A great deal of research shows that the drug resistance of bacteria is not only related to the generation of a great amount of drug-resistant strains, but also related to the formation of a biological film in vivo by pathogenic bacteria. The biofilm increases the resistance of bacteria to antibiotics by 10-1000 times compared with planktonic bacteria. The clinical biomembrane can be formed on various medical plants, has extremely strong drug resistance and immune evasion, and is one of the main reasons for clinical infection.

Pure titanium (Ti) and titanium alloys have been widely used as metal implants in the fields of orthopedics, orthopedics and dentistry due to their excellent biosafety and mechanical properties, and the formation of surface biofilms is an important cause of implant infection, and once a biofilm is formed, bacteria in the biofilm exhibit high resistance to antibiotics and host defense, i.e., high drug resistance. For example, Staphylococcus aureus (Staphylococcus aureus), Staphylococcus epidermidis, escherichia coli and the like are main pathogenic bacteria of bone infection related to orthopedic implants, wherein Staphylococcus aureus accounts for more than 50%, and Methicillin-resistant Staphylococcus aureus (MRSA) is common. Inhibiting the bacterial film formation on the surface of the metal built-in material has important clinical requirements for ensuring the health of human bodies, and the current ideal solution for the infection of metal implants is to enable the surface of plants in the metal to have an antibacterial function so as to inhibit or reduce the formation of biofilms. The common method is to modify the plant surface in the metal into an antibacterial coating. The clinical antibacterial coating is mainly a drug-releasing coating, and antibacterial agents are loaded on the coating and are released to kill surrounding bacteria. The antibacterial agent includes: organic antibacterial agents such as antibacterial peptides (AMP), antibiotics, antiseptics, and the like; ② the inorganic antibacterial agent comprises silver (Ag), copper (Cu), zinc (Zn), bismuth (Bi), chlorine (Cl), iodine (I), fluorine (F) and the like.

The active release antibacterial coating has strong antibacterial power, but the release speed and the release amount of the medicament are difficult to control, the initial burst effect of the active release antibacterial coating can cause higher local concentration, generate cytotoxicity and be difficult to maintain the long-term antibacterial activity.

Disclosure of Invention

The invention aims to overcome the defects in the prior art and provide a nano-antibacterial coating material based on a nano-gold cluster and a preparation method thereof, wherein a pMBA-AuNCs cluster is used as a basic constitutional unit, and a nano-metal oxide material (MM-MONs) with a three-dimensional network structure is formed by in-situ self-assembly with various functional metal ions, so that the function of weak non-covalent interaction in overall coordination characteristics can be facilitated to be clarified.

In order to solve the technical problems, the invention adopts the following technical scheme:

the invention provides a preparation method of a nano antibacterial coating material based on a nano gold cluster, which comprises the following steps:

step one, using freshly prepared HAuCl₄Adding the solution and p-mercaptobenzoic acid (pMBA) solution into ultra-pure water in sequence, and stirring to obtain a white Au (I) -pMBA complex;

step two, adding NaOH solution to dissolve white Au (I) -pMBA complex, and then adding NaBH₄Continuously stirring the solution at room temperature for 2-4h, and collecting and concentrating by using an ultrafiltration tube to obtain a pMBA-AuNCs cluster water solution;

step three, putting the pMBA-AuNCs cluster water solution into a centrifuge tube, adding HCl to ensure that pMBA molecules of the ligand are completely protonated, separating out the pMBA-AuNCs cluster from the water solution, centrifugally collecting and removing redundant HCl and NaCl in supernatant, washing twice by using ultrapure water, drying at room temperature, and adding methanol for dissolution to obtain a pMBA-AuNCs cluster methanol solution with completely protonated ligand terminal carboxyl;

step four, adding a salt solution of IVB group quadrivalent ions into the pMBA-AuNCs cluster methanol solution obtained in the step three, and then carrying out ultrasonic treatment for 5-10min to obtain a mixed solution;

step five, adding Cu into the mixed solution²⁺Salt solution, and ultrasonic treating for 5-10min to obtain pAnd mixing the MBA-AuNCs cluster and metal ion methanol to obtain the nano antibacterial coating material.

Further, the HAuCl described above₄The molar ratio to pMBA was 1: 2.

Further, the IVB group quadrivalent ion in the fourth step is Ti⁴⁺、Zr⁴⁺Or Hf⁴⁺。

Further, the molar ratio of the pMBA-AuNCs cluster to the group IVB tetravalent ion is 1: 10.

Further, the salt solution is a hydrochloride salt solution.

Further, the above-mentioned IVB group tetravalent ion and Cu²⁺In a ratio of 1: (1-2).

Further, the ultrasonic treatment time in the fourth step and the fifth step was 5 min.

The second aspect of the present invention is to provide the nano-gold cluster-based nano-antibacterial coating material prepared by the above preparation method.

The third aspect of the invention provides the application of the nano antibacterial coating material in preparing an antibacterial nano coating.

The fourth aspect of the invention provides a method for preparing an antibacterial nano coating by adopting the nano antibacterial coating material, which comprises the steps of uniformly dripping the nano antibacterial coating material on a dried titanium sheet, and drying at room temperature to form the antibacterial nano coating.

Further, the titanium sheet is treated on the surface by a strong acid oxidation method before use: the polished titanium sheet is placed in 30 percent of H₂O₂And H₂SO₄Oxidizing in the mixed solution with the volume ratio of 1:1 for 10-15min to form hydroxyl-rich TiO on the surface₂A thin layer; finally, washing with a large amount of clear water, and putting into absolute ethyl alcohol for later use.

By adopting the technical scheme, compared with the prior art, the invention has the following technical effects:

the nano-antibacterial coating material based on the nanogold cluster regulates the property of the cluster surface ligand, the surface ligand and metal ions form a three-dimensional periodic net-shaped film structure through self-assembly on the surface of a plant in metal through coordination, excellent in-vivo and in-vitro anti-biofilm activity is shown, an integrated treatment concept of preventing a biofilm and controlling infection is provided, and a new thought is provided for clinical treatment of related infection of the plant.

Drawings

FIG. 1 is a flow chart of the preparation of a nano-antimicrobial coating in one embodiment of the present invention;

FIG. 2 is an electron microscope scan of a pMBA-AuNCs cluster in accordance with an embodiment of the present invention;

FIG. 3 is a graph showing the results of performance characterization measurements of the nano-antimicrobial coating prepared according to an embodiment of the present invention; wherein: FIG. a is a surface topography of various nano-antimicrobial coatings; FIG. b is a graph of high resolution O1sXPS spectra and corresponding high resolution Ti2p, Zr3d, and Hf4fXPS spectra obtained for fully protonated GNCS, Ti-MONS, Zr-MONS, and Hf-MONS thin films; FIG. c is a HAADF-STEM image and EDX elements of the Zr-Cu MONS thin films formed in copper mesh TEM grids;

FIG. 4 shows in vitro antimicrobial experimental results for Zr-Cu MONs coatings in accordance with an embodiment of the present invention; wherein: FIG. a is a graph of Zr-Cu MONs in vitro MRSA adhesion resistance constructed by copper ions with different concentrations; FIG. b is the influence of in vitro co-culture of Zr-Cu MONs constructed by copper ions with different concentrations and MRSA on the growth of the Zr-Cu MONs; c and d are graphs of the results of Zr-Cu MONs in vitro MRSA adhesion resistant representative CFU counting coated plates and SEM constructed by copper ions with different concentrations;

FIG. 5 shows in vitro results of an antimicrobial assay for Ti-Cu MONs coatings in accordance with an embodiment of the present invention; wherein: FIG. a is a representative photograph of bacterial colonies on an agar plate; FIG. b is a statistical histogram of implants coated with Ti-Cu Mons membranes and inoculated with MRSA bacterial suspensions; FIG. c is a graph of the effect of Ti-Cu MONS coatings on MRSA proliferation at different Ti/Cu blend ratios;

FIG. 6 shows the in vitro antibacterial test results of Hf-Cu MONs coating in one embodiment of the present invention; wherein: FIG. a is a representative photograph of bacterial colonies on an agar plate; FIG. b is a statistical histogram of an implant coated with Hf-Cu Mons membrane and inoculated with MRSA bacterial suspension; FIG. c is a graph of the effect of Hf-Cu MONS coatings on MRSA proliferation at different Hf/Cu blend ratios;

FIG. 7 shows the results of in vivo antibacterial experiments with Zr-Cu MONs coatings in accordance with an embodiment of the present invention; wherein: FIG. a shows a schematic representation of the reduction of residual bacteria on the surface and tissue of an implant in a rat subcutaneous intra-implant infection model for MRSA; panel b shows a titanium sheet placed in a rat on a scale of 20 mm; fig. c shows the statistical results of CFU counts for different implant surfaces; FIG. d shows the corresponding statistical results of CFU counts of the tissue surrounding the implant; panel E shows H & E staining of tissue surrounding the implant, scale bar 150 μm; panel f shows H & E stained sections of rat major organs (heart, liver, spleen, lung, kidney) with uncoated groups and different Cu/Zr ratio coated groups, scale bar 200 μm; denotes p <0.001 compared to the uncoated group.

Detailed Description

The nano-antibacterial coating material based on the nanogold cluster provided by the invention takes the pMBA-AuNCs cluster as a basic constitutional unit, and forms a three-dimensional reticular structure through in-situ self-assembly with a plurality of functional metal ions, thereby being helpful for clarifying the function of weak non-covalent interaction in the overall coordination property. First type of metal ion (tetravalent Zr) participating in the construction of the film⁴⁺、Ti⁴⁺、Hf⁴⁺) Can form stable chemical coordination with the cluster, and the second type metal ion (Cu)²⁺) The film coating has the advantages of dissolving antibacterial property, weak coordination effect with clusters, participation of two types of metal ions in the coordination of the clusters, reasonable utilization of the advantages of the two types of metal ions, development of a film coating with chemical stability and antibacterial ion dissolving property, reduction of cytotoxicity and difficulty in generation of drug resistance.

The preparation method of the nano-gold cluster-based nano-antibacterial coating material comprises the following steps:

step three, putting the pMBA-AuNCs cluster water solution into a centrifuge tube, adding HCl to ensure that pMBA molecules of the ligand are completely protonated, separating out the pMBA-AuNCs cluster from the water solution, centrifugally collecting and removing redundant HCl and NaCl in supernatant, washing twice by using ultrapure water, drying at room temperature, and adding methanol for dissolution to obtain a pMBA-AuNCs cluster methanol solution (GNCS) with completely protonated ligand terminal carboxyl;

step five, adding Cu into the mixed solution²⁺Salt solution, and then ultrasonic treatment is carried out for 5-10min to obtain mixed solution of pMBA-AuNCs cluster and metal ion methanol, namely the nano antibacterial coating material MM-MONs.

In a preferred embodiment of the present invention, the HAuCl is₄The molar ratio to pMBA was 1: 2.

In a preferred embodiment of the present invention, the group IVB tetravalent ion in step four is Ti⁴⁺、Zr⁴⁺Or Hf⁴⁺。

In a preferred embodiment of the invention, the molar ratio of the pMBA-AuNCs cluster to the group IVB tetravalent ion is 1: 10.

In a preferred embodiment of the present invention, the salt solution is a hydrochloric acid salt solution.

In a preferred embodiment of the present invention, the IVB group tetravalent ion and Cu are²⁺In a ratio of 1: (1-2); more preferably 1: 2. Precisely, within a certain range of ratios, Cu²⁺The larger the ratio of (A) to (B), the better antibacterial effect of the coating prepared by the material.

In a preferred embodiment of the present invention, the ultrasonic treatment time in the fourth step and the fifth step is 5 min.

The present invention will be described in detail and specifically with reference to the following examples and drawings so as to better understand the present invention, but the following examples do not limit the scope of the present invention.

In the examples, the conventional methods were used unless otherwise specified, and the reagents used were, for example, conventional commercially available reagents or reagents prepared by conventional methods without specifically specified.

Example 1

Referring to fig. 1, the present embodiment provides a nano antibacterial coating based on a nano gold cluster, and the preparation method thereof includes the following steps:

(1) treatment of titanium sheets

Selecting a titanium alloy wafer with the diameter of 1cm and the thickness of 0.5mm as a representative of a metal implant, polishing the surface of the titanium wafer by using silicon carbide waterproof abrasive paper (320,400,600,800,1000 or 1200 meshes) in a water environment until no obvious scratch exists, sequentially putting the titanium alloy wafer into acetone, absolute ethyl alcohol and deionized water, respectively cleaning the titanium alloy wafer for 10min by using an ultrasonic cleaner, and finally soaking the titanium alloy wafer in the absolute ethyl alcohol for later use. Then, the surface of the titanium sheet is treated by a strong acid oxidation method, and the method comprises the following specific steps: the titanium sheet after polishing treatment is placed in H₂O₂(30％)/H₂SO₄Oxidizing in the mixed solution with the volume ratio of 1:1 for 10-15min to form hydroxyl-rich TiO on the surface₂A thin layer. Finally, washing with a large amount of clear water, and putting into absolute ethyl alcohol for later use.

(2) Preparation of pMBA-AuNCs cluster with completely protonated ligand carboxyl

Step one, 1.25mL HAuCl prepared in fresh is added₄(20mM) was added to 10mL of ultrapure water followed by 5mL of p-mercaptobenzoic acid (10 mM) solution, and stirred for 5 min. The white Au (I) -pMBA complex was then solubilized by the addition of 1.5mL of 1M NaOH solution, followed by the addition of 0.5mL of freshly prepared 112mM NaBH₄The solution was stirred at room temperature for 3 h. After the reaction is finished, the ultrafiltration tube collects a concentrated material pMBA-AuNCs cluster, and the carboxyl group at the tail end of the pMBA ligand is in a state of coexistence of protonation (-COOH) and deprotonation (-COO-).

And step two, taking 5mL (2mg/mL) of the pMBA-AuNCs cluster aqueous solution into a centrifuge tube, adding about 5mL of 1M HCl, completely protonating the pMBA molecules of the ligand, separating out the pMBA-AuNCs cluster from the aqueous solution, centrifugally collecting, removing redundant HCl and NaCl in the supernatant, washing twice by using ultrapure water, finally drying at room temperature, adding methanol for dissolving, and obtaining the pMBA-AuNCs cluster methanol solution (GNCS) with completely protonated carboxyl at the tail end of the ligand (the scanning electron microscope of the pMBA-AuNCs cluster methanol solution is shown in figure 2).

(3) preparation of mixed solution of pMBA-AuNCs cluster and metal ion methanol

Step one, taking 1mL of the pMBA-AuNCs cluster methanol solution of 2mg/mL prepared in the step one, respectively adding a certain amount of TiCl prepared from methanol into a centrifugal tube₄、ZrCl₄And HfCl₄Solution of pMBA-AuNCs clusters with Ti⁴⁺、Zr⁴⁺、Hf⁴⁺The molar ratio of the mixed solution is 1:10 respectively, and the mixed solution is subjected to ultrasonic treatment for 5min for later use.

Step two, adding CuCl into the mixed solution after ultrasonic treatment₂The solution was then sonicated for 5min to prepare a M-Cu MONs solution.

(4) Preparation of antibacterial nano coating

And (3) respectively sucking 0.15mL of M-Cu MONs solution by a liquid transfer machine, uniformly dripping the M-Cu MONs solution on the treated dried titanium sheet (drop-casting), and drying at room temperature to form a uniform film visible to the naked eye, thereby obtaining the nano-coating modified titanium sheet formed by pMBA-AuNCs cluster and metal ions through coordination bonds. The concentration of the film can be adjusted by adjusting the concentration of the solution and the amount of the liquid to be dropped.

Verification example 1

In this embodiment, the performance of the nano-gold cluster-based nano-antibacterial coating is studied, and in order to facilitate the characterization of the coating performance, a silicon wafer with the same surface rich in hydroxyl groups is used to replace a titanium wafer to prepare a thin film coating, and then the thin film coating is tested. The specific experimental procedures and results are as follows:

1. GNCS-Ti, GNCS-Zr and GNCS-Hf suspensions were deposited on the wafer surface (5 mm. times.5 mm), PMBA: m⁴⁺1:1, Ti-, Zr-, and Hf-MON were formed, respectively, and samples for SEM characterization were prepared. The surface morphology of the film was studied by a scanning electron microscope (SEM, Tescan MAIA3) at an accelerating voltage of 5kV, and the result is shown in FIG. 3a, wherein the formed coating has regular morphology and a thickness of between 100 and 200 nm.

X-ray photoelectron Spectroscopy the chemical states of GNCs films and Ti, Zr or Hf-MONs films were investigated by X-ray photoelectron Spectroscopy using a Krato Axis Ultra DLD instrument equipped with a monochromatic Al K α (1486.6eV) source. Light alignment at 284.8eV with C1SThe spectra were calibrated. The suspension was deposited on the surface of a silicon wafer (5 mm. times.5 mm), pMBA: m⁴⁺：Cu²⁺1: 1:1, samples were prepared and dialyzed 1d against ultrapure water to remove non-coordinated metal ions, the results are shown in FIG. 3 b.

3. Transmission Electron microscopy High Angle Annular Dark Field (HAADF) Scanning Transmission Electron Microscopy (STEM) images of MM-MONS films were studied, operating at 200kV with FEI Talos F200X G2S/TEM. The element profile of the MM-MONS film was studied using an integrated Super-XEDS detector. Mixing GNCs and MCl₄(M ═ Ti, Zr, Hf) and CuCl₂A mixture in ethanol (about 4 μ L) was deposited onto a carbon film coated copper mesh TEM grid (Cu, 300 mesh) and allowed to evaporate in air to form a MM-MONS thin film for HAADF-STEM measurement and elemental mapping, the results of which are shown in fig. 3 c.

Verification example 2

In this example, MRSA is used as a model strain to study the in vitro antibacterial performance of the nano coating, and the specific test method and results are as follows:

(1) bacterial culture

MRSA monoclonal colonies on Sheep Blood Agar (SBA) plates were picked, soaked in 5mL fresh Tryptase Soy Broth (TSB) medium, and cultured at 37 ℃ at 200rpm to mid-logarithmic phase. Then diluting the obtained bacterial suspension in TSB (pH 7.4) to 1-2 × 10 according to the optical density (OD600) of the suspension at 600nm⁶CFU/mL is ready for use.

(2) Evaluation of ability to form MRSA biofilm on coating surface

Untreated titanium sheets (control group) and 3 types of nano-film antibacterial coating titanium sheets (experimental group) prepared in example 1 were placed in a 24-well plate, 5 multiple wells in each group with the coating facing up, 1mL of the MRSA bacterial liquid from the previous step was taken up with a micro-sampler and added to the 24-well plate, incubated in a 37 ℃ incubator for 24 hours, then the titanium sheets were gently rinsed three times with PBS to remove non-adhered bacteria, then placed in a centrifuge tube and shaken ultrasonically for 1min to disperse the bacteria adhered to the titanium sheets in the PBS solution, 100 μ L of the diluted solution was applied to a culture plate, incubated in a 37 ℃ incubator for 24 hours, and colonies were counted and photographed.

The MRSA biofilm formed on the surface of the titanium sheet coating is evaluated through the antibacterial rate and the adhesion rate: the antibacterial rate is (number of colonies in a control group-number of colonies in an experimental group)/number of colonies in the control group multiplied by 100%; the adhesion rate was defined as the number of colonies in the experimental group/the number of colonies in the control group × 100%.

(3) SEM observation of MRSA biological film on coating surface

The co-culture method of the M-Cu MONs titanium sheets and the bacteria of the experimental group and the control group is the same as the above step, after 6 hours of culture, a pipettor is used for absorbing the culture solution and gently rinsing the titanium sheets with PBS for three times, 2 mL2.5% glutaraldehyde is added respectively and is fixed at 4 ℃ overnight, then the gentle rinsing is carried out with the PBS solution in sequence, the ethanol is used for gradient dehydration (30%, 50%, 70%, 80%, 90%, 100%) for 15min each time, and finally the vacuum drying, the gold spraying and the scanning electron microscope observation are carried out, and the result is shown in the graph 4-6.

As can be seen from FIGS. 4-6, the prepared M-Cu MONs titanium sheet has good antibacterial property, and M is⁴⁺And Cu²⁺Within a certain proportion range, Cu²⁺The higher the content of (A), the better the bacteriostatic ability.

Verification example 3

In this example, the in vivo antibacterial property of the nano-coating provided in example 1 was studied using a rat subcutaneous implant-associated infection animal model.

Among them, the research on the rat subcutaneous endophyte-related infection model was approved by the animal research committee of the sixth national hospital affiliated to Shanghai university of transportation. Male Sprague-Dawley rats (200. + -.10 g) at 6 weeks of age were used for the implant-related infection model and all rats were housed in an SPF environment. The specific test methods and results are as follows:

after anaesthesia with 3% sodium pentobarbital solution (1mL/kg), the hair was removed from the back of the rats with a razor, then a full skin incision of 1cm diameter was made in the back with a scalpel under sterile conditions, and a titanium sheet covered with a Zr-Cu MONs coating was immediately implanted under the skin of the back of each rat (see FIG. 7 b). After implantation, 70. mu.L of methicillin-resistant Staphylococcus aureus suspension (1X 10)⁷CFU/mL) was seeded on the implant surface. Thereafter, the skin was sutured layer by layer. Date for making mouldThe total experimental time was 1 week for day 0, and the implants and tissues surrounding the implants were subjected to bacterial enumeration on

days

1, 3 and 7. Fixing and embedding part of tissues and important organs (heart, liver, spleen, lung and kidney) around the implant at the same time for H&E staining, results are shown in FIG. 7.

As can be seen from FIGS. 7c-7f, the prepared Zr-Cu MONs titanium sheet has good antibacterial property, and Zr⁴⁺And Cu²⁺Within a certain proportion range, Cu²⁺The higher the content of (A), the better the bacteriostatic ability.

The embodiments of the present invention have been described in detail, but the embodiments are merely examples, and the present invention is not limited to the embodiments described above. Any equivalent modifications or alterations to this practice will occur to those skilled in the art and are intended to be within the scope of this invention. Accordingly, equivalent changes and modifications made without departing from the spirit and scope of the present invention should be covered by the present invention.

Claims

1. A preparation method of a nano antibacterial coating material based on a nano gold cluster is characterized by comprising the following steps:

step one, using freshly prepared HAuCl₄Adding the solution and the pMBA solution into ultrapure water in sequence, and stirring to obtain a white Au (I) -pMBA complex;

step two, adding NaOH solution to dissolve the white Au (I) -pMBA complex, and then adding NaBH₄Continuously stirring the solution at room temperature for 2-4h, and collecting and concentrating by using an ultrafiltration tube to obtain a pMBA-AuNCs cluster water solution;

step three, putting the pMBA-AuNCs cluster water solution into a centrifuge tube, adding HCl to ensure that pMBA molecules of the ligand are completely protonated, separating out the pMBA-AuNCs cluster from the water solution, centrifugally collecting and removing redundant HCl and NaCl in supernatant, washing twice by using ultrapure water, drying at room temperature, and adding methanol to dissolve to obtain a pMBA-AuNCs cluster methanol solution with completely protonated ligand terminal carboxyl;

step five, adding Cu into the mixed solution²⁺Salt solution, and then carrying out ultrasonic treatment for 5-10min to obtain a mixed solution of the pMBA-AuNCs cluster and metal ion methanol, namely the nano antibacterial coating material.

2. The method according to claim 1, wherein the group IVB tetravalent ion in step IV is Ti⁴ ⁺、Zr⁴⁺Or Hf⁴⁺。

3. The method according to claim 1, wherein the molar ratio of the pMBA-AuNCs cluster to the group IVB tetravalent ion in step four is 1: 10.

4. The method of claim 1, wherein the salt solution is a hydrochloride solution.

5. The method of claim 1, wherein the group IVB tetravalent ion is reacted with Cu²⁺In a ratio of 1: (1-2).

6. The method according to claim 1, wherein the sonication time in the fourth and fifth steps is 5 min.

7. A nano-gold cluster-based nano-antibiotic coating material prepared by the preparation method as set forth in any one of claims 1 to 6.

8. Use of the nano-antibacterial coating material according to claim 7 for the preparation of antibacterial nano-coatings.

9. A method for preparing an antibacterial nano coating by using the nano antibacterial coating material as claimed in claim 7, characterized in that the nano antibacterial coating material is uniformly dropped on a dried titanium sheet and dried at room temperature to form an antibacterial nano coating.

10. The method according to claim 9, characterized in that the titanium sheet is treated before use with a strong acid oxidation process for the surface treatment: the polished titanium sheet is placed in 30 percent of H₂O₂And H₂SO₄Oxidizing in the mixed solution with the volume ratio of 1:1 for 10-15min to form hydroxyl-rich TiO on the surface₂A thin layer; finally, washing with a large amount of clear water, and putting into absolute ethyl alcohol for later use.