Hollow spherical metal organic framework material capable of effectively resisting bacteria and inhibiting bacterial drug resistance

Abstract

According to the present invention, in the prior art, staphylococcus aureus infection and antibiotic resistance (AMR) are two important factors threatening public health, such that persistent pain and even death may be caused, and the common health can be threatened by the staphylococcus aureus infection and the antibiotic resistance (AMR), such that the common health can be threatened by the staphylococcus aureus infection and the antibiotic resistance (AMR), and the common health can be threatened by the staphylococcus aureus infection and the antibiotic resistance (AMR); therefore, it is urgent to develop alternative antibacterial materials. In the invention, a biocompatible nano system (ZIF-67 HS) based on a hollow spherical zeolite imidazate skeleton is successfully developed, and the biocompatible nano system has a remarkable bactericidal effect on gram-positive staphylococcus aureus in vitro and in vivo. Importantly, the research result shows that S. aureus does not generate secondary drug resistance like traditional antibiotics when being exposed to ZIF-67 HS with sub-minimum inhibitory concentration for a long time; on the contrary, it maintains continuous sensitivity to antibiotic resistant strains. Multi-omics analysis (including transcriptomics, metabonomics and metallomics) is combined with bioassays to prove that ZIF-67 HS destroys a plurality of necessary pathways in S.aures, including amino acid metabolism, iron metabolism, membrane transport, ROS and S.aures infection pathways. According to the invention, a new insight is provided for the potential of nano biological medicines in the aspect of removing bacteria, and a related molecular mechanism is clarified.

Description

Technical Field

[0001] This invention relates to the field of antibacterial materials technology, and in particular to the synthesis and application of a hollow spherical metal-organic framework material that effectively fights bacteria and inhibits bacterial resistance. Background Technology

[0002] Throughout human history, bacteria have profoundly impacted all aspects of human survival, including accelerating food spoilage, damaging the environment, corroding machinery, and even threatening human health. The prevalence and spread of diseases such as endocarditis, bacteremia, osteomyelitis, and soft tissue infections caused primarily by Gram-positive Staphylococcus aureus (S. aureus) are significant causes of patient mortality, prompting increasing research interest in antimicrobial materials. In the 1940s, the introduction of antibiotics such as penicillin and streptomycin effectively addressed the problems of prevalent diseases at the time. However, the overuse of antibiotics led to the emergence of drug-resistant bacteria, exacerbating the burden of disease treatment. Furthermore, with the slow progress of antibiotic development in recent years, novel non-antibiotic drugs have emerged as potential alternatives to traditional treatments, including bacteriophages, antimicrobial peptides (AMPs), antimicrobial enzymes, and nanomaterials.

[0003] Nanomaterials possess a unique ability to interact with multiple targets, particularly conserved bacterial components that can resist gene mutations. These components are easier to modify, reducing the likelihood of them evolving drug resistance. Furthermore, functional modifications can unlock their potential for diverse antibacterial effects. Metal-organic frameworks (MOFs) have been extensively studied for their antibacterial properties due to their high porosity, large specific surface area, tunable pore structure, ease of functionalization, and biocompatibility. More importantly, their unique properties make them suitable for multifunctional antibacterial platforms. MOFs can act as reservoirs, releasing antibacterial metal ions or organic linkers upon degradation to exert their effects on bacteria. Moreover, MOFs can be designed with tunable structures to adapt to different antibacterial methods. For example, MOFs with high porosity and stimuli reactivity can serve as carriers for targeted drug delivery; photosensitizing MOFs generate heat and free radicals under light irradiation; and enzyme-like MOFs exhibit bactericidal properties by mimicking enzymes. Of course, the combination of various properties within MOFs may also synergistically promote bacterial eradication.

[0004] Zeolite imidazole framework-67 (ZIF-67) is a type of zeolite composed of cobalt ions (Co). 2+ZIF-67 is a MOF (MeInductor Flask) formed by the coordination of ZIF-67 with dimethylimidazole (2-MeIM). Due to its biodegradability, low toxicity, and consistent properties with other MOFs, ZIF-67 shows great promise in nanomedicine, particularly in antibacterial therapy. It has already been successfully used to kill bacteria such as Staphylococcus aureus, Escherichia coli, and Pseudomonas putida. However, the rhombic dodecahedral ZIF-67 has been reported to achieve optimal antibacterial effects only when combined with other bioactive materials. Therefore, fully developing the antibacterial properties of ZIF-67 is a key research focus. Summary of the Invention

[0005] The purpose of this invention is to construct novel multi-target antibacterial materials using the unique morphology of ZIF-67, which not only exhibit significant antibacterial effects but also address the problem of antibiotic resistance in microorganisms, elucidate the relevant antibacterial molecular mechanisms, and provide a deeper understanding and new perspective for the discovery of novel antibacterial materials.

[0006] To achieve the above objectives, the technical solution adopted by the present invention is as follows: a hollow spherical zeolite imidazole framework material (ZIF-67 HS) with a uniform particle size distribution of 655.2±34.5 nm. The formation of the hollow spherical material involves the introduction of hexadecyltrimethylammonium bromide (CTAB) surfactant as a surface capping agent and soft template, which is beneficial to the formation of a spherical CTAB / water micelle system with water, thereby realizing the nucleation and growth process.

[0007] This invention provides the application of the aforementioned ZIF-67 HS material in combating Gram-positive Staphylococcus aureus. Time- and dose-dependent growth curve analysis, colony counting, and in vitro SEM observation of bacterial morphology demonstrated the significant bactericidal effect of ZIF-67 HS against Staphylococcus aureus.

[0008] This invention demonstrates the significant potential of ZIF-67 HS in promoting the healing of infected wounds. The in vivo antibacterial efficacy of ZIF-67 HS was evaluated by constructing a mouse dorsal wound infection model with Staphylococcus aureus and applying nanomedicine.

[0009] This invention provides the effect of ZIF-67 HS on the evolution of antibiotic resistance (AMR). By passaged with antibiotics for 15 days to form resistant strains, and comparing them with antibiotics (kanamycin monosulfate and gentamicin sulfate), the study measures whether long-term exposure to ZIF-67 HS also leads to resistance, and whether ZIF-67 HS remains susceptible to antibiotic-resistant bacteria.

[0010] This invention provides a biocompatibility assessment of ZIF-67 HS. The safety and feasibility of ZIF-67 HS as an antibacterial drug were verified through cytotoxicity tests, erythrocyte hemolysis tests, and changes in body weight during animal experiments.

[0011] This invention provides the molecular mechanism of action of ZIF-67 HS in killing Staphylococcus aureus. By combining transcriptomics, metabolomics, metallomics, and related biovalidation experiments, the multiple targets of ZIF-67 HS in acting on bacteria and inducing bacterial death were analyzed.

[0012] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0013] First: The antibacterial nanomaterial ZIF-67 HS provided by the technical solution of the present invention has a special structure, exhibits excellent antibacterial effects in vitro and in vivo, and has good biosafety, is not prone to causing drug resistance, and is even sensitive to antibiotic-resistant bacteria.

[0014] Second, the antibacterial nanomaterial ZIF-67 HS provided by the technical solution of this invention has a multi-target antibacterial mechanism, simultaneously interfering with intracellular amino acid and iron metabolism, blocking membrane transport, inhibiting the secretion of virulence factors, and inducing oxidative stress, all of which jointly induce bacterial damage and even death. These characteristics enable it to exhibit sustained antibacterial effects without secondary drug resistance, providing a new approach to antibacterial novel nanomaterials. Attached Figure Description

[0015] Figure 1 This is a characterization diagram of ZIF-67 HS material in the analytical method of this invention;

[0016] Figure 2 This is a graph showing the antibacterial activity test results of ZIF-67 HS material in the analytical method of this invention;

[0017] Figure 3 These are SEM images of Staphylococcus aureus before and after treatment with different concentrations of ZIF-67 HS in the analytical method of this invention;

[0018] Figure 4 This is a graph showing the comparison of Staphylococcus aureus resistance to ZIF-67 HS, kanamycin monosulfate, and gentamicin sulfate in the analytical method of this invention.

[0019] Figure 5 This is a graph showing the results of ZIF-67 HS promoting wound repair in Staphylococcus aureus infection using the analytical method of this invention.

[0020] Figure 6 This is a diagram showing the results of proving the biocompatibility and biosafety of ZIF-67 HS in the analytical method of this invention;

[0021] Figure 7 This is a diagram showing the bioinformatics analysis results of Staphylococcus aureus treated with ZIF-67 HS in the analytical method of this invention;

[0022] Figure 8 The accompanying figure is a summary drawing of the analytical method of this invention. Detailed Implementation

[0023] To better illustrate the purpose, technical solution, and advantages of this invention, the invention will be further described below with reference to specific embodiments. Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods. Unless otherwise specified, the materials and reagents used in the following embodiments are commercially available. The water used in the following embodiments is ultrapure water, treated by a Milli-Q ultrapure water purification system.

[0024] Implementation Example 1. Characterization of Gold Nanoparticle Probes: Preparation and Characterization of ZIF-67 HS

[0025] The specific preparation method is as follows:

[0026] A1. Dissolve 0.729 g of cetyltrimethylammonium bromide (CTAB, 0.1 mol / L) in 140 mL of water and stir magnetically in a water bath at 30°C for 20 min until the solution is clear and transparent;

[0027] A2. Add 750 μL of Co(NO3)2·6H2O solution (0.5 mol / L) and continue stirring for 20 min;

[0028] A3. Quickly pour 15 mL of 2-methylimidazole solution (1.096 mol / L) into the above solution and react at a constant temperature for 2 h;

[0029] A4. The purple product was washed by centrifugation with N,N-dimethylformamide (DMF) and methanol, and dried under vacuum at 60°C to obtain a purple powder;

[0030] A5. The product was analyzed by scanning electron microscopy, transmission electron microscopy, powder X-ray diffraction, X-ray photoelectron spectroscopy, N2 adsorption-desorption isotherm, Zeta potential, and Co. 2+ The release curve is used for characterization.

[0031] Characterization results as follows Figure 1 As shown, ZIF-67 HS was successfully synthesized, exhibiting a hollow spherical structure with a particle size distribution of 655.2 ± 34.5 nm and an actual Co content of 24.99 ± 0.19 (wt.%). It also exhibits type I isotherm characteristics, with a Brunauer-Emmett-Teller (BET) area of ​​1070 m².2 / g. The material has a positively charged surface and is stable in water and culture medium, demonstrating its feasibility as an antibacterial material.

[0032] Implementation Example 2. Antibacterial Activity Evaluation of ZIF-67 HS

[0033] The specific testing steps are as follows:

[0034] B1. Pick a single colony of preserved S. aureus (ATCC 6538) and place it in 5 mL of LB liquid medium and incubate overnight (37°C, 150 rpm).

[0035] B2. Dilute the logarithmic growth phase bacterial culture with LB medium to 1×10⁻⁶. 7 CFU / mL to be used;

[0036] B3. ZIF-67 HS suspension was added to a 96-well plate using a 2-fold dilution method, 100 μL per well. Then, 100 μL of a bacterial concentration of 1×10⁻⁶ was added. 7 The bacterial suspension was prepared at CFU / mL to achieve a final drug concentration of 0.4-200 μg / mL. The control group consisted of 200 μL of pure LB medium and 200 μL of untreated bacterial suspension.

[0037] B4. Incubate the 96-well plate at 37°C and measure the OD every 2 hours. 600 Value, incubate for 12 hours;

[0038] B5. The minimum inhibitory concentration (MIC) is the lowest ZIF-67 HS concentration without significant turbidity;

[0039] B6. Mix 2 mL of ZIF-67 HS at different concentrations with 2 mL of bacterial suspension to a final concentration of 0.5MIC, MIC and 2MIC, and incubate at 37°C for 4, 12 and 24 h;

[0040] B7. After diluting the obtained bacterial suspension by 10,000 times, take 100 μL and inoculate it evenly on LB agar plates. After incubating at 37°C for 24 h, the difference in colony count can be directly observed by plate counting method.

[0041] Test results are as follows Figure 2 As shown, when the ZIF-67 HS concentration is below 3.2 μg / mL, the survival rate of Staphylococcus aureus can exceed 91.62%, while when the concentration reaches 6.25 μg / mL, the survival rate of Staphylococcus aureus is only about 10%. Therefore, the MIC value for ZIF-67 HS to inhibit the growth of S. aureus is 6.25 μg / mL. Plate colony counting results also show that the MIC-treated group had the fewest bacterial growths.

[0042] Implementation Example 3. Damaging Effects of ZIF-67 HS on Biomembranes

[0043] The specific testing steps are as follows:

[0044] C1. Mix 2 mL of ZIF-67 HS at different concentrations with 2 mL of bacterial suspension to a final concentration of 0, MIC, 2MIC, 4MIC, and 8MIC, and incubate at 37°C for 6 h.

[0045] C2. The obtained bacterial suspension was centrifuged and washed three times with phosphate-buffered saline (PBS) to obtain the bacterial precipitate;

[0046] C3. Add 2.5% glutaraldehyde to the bacterial precipitate and fix it at 4°C for 12 h;

[0047] C4. After fixation, dehydrate the product sequentially using different volume ratios of ethanol (30%, 50%, 70%, 80%, 90% and 100%) for 15 min each.

[0048] C5. Add 1 ml of isoamyl carbonate to replace the residual ethanol and water in the sample;

[0049] C6. The obtained suspension was dropped onto a silicon wafer, dried with carbon dioxide at the critical point, and then gold was sprayed onto the surface. The morphology of the bacteria was observed using a scanning electron microscope.

[0050] Test results are as follows Figure 3 As shown, with increasing ZIF-67 HS dosage, bacterial cells exhibited shrinkage or swelling, eventually rupturing at concentrations of 4 MIC and 8 MIC. Control group cells had smooth surfaces, were spherical, and had intact membranes. Furthermore, nanoparticles were found to interact with bacteria in all drug treatment groups. These significant changes in morphology and membrane integrity, along with the presence of nanoparticles, indicate that ZIF-67 HS can attach to bacteria and cause membrane damage, thus demonstrating significant antibacterial efficacy.

[0051] Implementation Example 4. ZIF-67 HS Overcoming Antimicrobial Resistance (AMR)

[0052] The specific testing steps are as follows:

[0053] D1. The primary MICs of S. aureus treated with ZIF-67 HS, kanamycin monosulfate, and gentamicin sulfate were determined according to the procedure for determining MIC;

[0054] D2. The activated bacteria were diluted at a ratio of 1:1000 and incubated with a drug at a sub-MIC concentration for 12 h, and then the new MIC value was retested.

[0055] D3. After subculturing for 15 consecutive days in the same manner and determining the new MIC values, *S. aureus* (AMR) strains were obtained, namely *S. aureus* (AMR_HS), *S. aureus* (AMR_GS), and *S. aureus* (AMR_KM).

[0056] D4. Calculate the ratio of MIC of each generation of evolved bacteria to that of the primary bacteria, generate resistance curves, and evaluate the resistance status;

[0057] D5. Transfer 50 μL of the drug-resistant strain to 5 mL of medium containing ZIF-67 HS, kanamycin monosulfate and gentamicin sulfate at the primary MIC concentration, and incubate at 37°C for 6 h.

[0058] D6. Dilute the bacterial suspension, inoculate it evenly onto agar plates, count the colonies and calculate the survival rate;

[0059] D7. Test the drug dependence index (RDI) of three drugs against three strains of S. aureus (AMR), RDI = MIC1 / MIC0 (MIC1 and MIC0 correspond to the MIC of the evolutionary treatment strain and the primary strain, respectively).

[0060] Test results are as follows Figure 4 As shown, with increasing passage number, the MIC of *S. aureus* (AMR_GS) increased 64-fold, and the MIC of *S. aureus* (AMR_KM) increased 128-fold. In contrast, the ZIF-67 HS treatment group consistently exhibited bacterial inhibition at a constant concentration. Resistant bacteria, after treatment with antibiotics at MIC0 concentrations, showed significantly higher colony numbers than primary bacteria. Furthermore, antibiotic-resistant bacteria consistently showed sensitivity to ZIF-67 HS. Based on these results, it is demonstrated that ZIF-67 HS possesses sustained antibacterial properties and does not induce secondary resistance.

[0061] Implementation Example 5. ZIF-67 HS promotes the healing process of S. aureus infected wounds.

[0062] The specific testing steps are as follows:

[0063] E1. BALB / c mice (female, 6 weeks old) were selected for modeling and experiments. They were acclimatized for 3 days and fed and hydrated normally.

[0064] E2. Mice were anesthetized with propofol and a 7 mm circular full-thickness lesion was created at the same site on their backs.

[0065] E3. Instill 150 μL of S. aureus bacterial suspension (10 μL) into the wound. 8(CFU / mL), and a wound infection model was established after 48 h of infection;

[0066] E4. Mice were randomly divided into 3 groups and treated with 50 μL PBS and ZIF-67 HS (MIC), respectively.

[0067] E5. Repeat the treatment for 12 days, and record the wound photos / size and mouse weight daily;

[0068] E6. The wound area and contour were analyzed using ImageJ software.

[0069] Test results are as follows Figure 5 As shown in the wound photographs, the ZIF-67 HS group exhibited significantly faster wound healing, with almost complete closure and new hair growth by day 12, while the control group remained dry and crusted. Quantitative analysis of the wound contour and area also supported these findings; after 9 days of treatment, the wound area in the ZIF-67 HS group was only 26.46%, while the control group still had 60.26% coverage. In conclusion, ZIF-67 HS effectively accelerates wound healing by eliminating bacteria, inhibiting infection and inflammation, and promoting tissue regeneration.

[0070] Implementation Example 6. ZIF-67 HS exhibits good biocompatibility.

[0071] The specific testing steps are as follows:

[0072] Cytotoxicity assay: The cytotoxicity of ZIF-67 HS was evaluated using the CCK-8 assay.

[0073] F1. Human embryonic kidney cells (HEK293) and human liver cancer cells (HepG2) were seeded into 96-well cell culture plates and incubated at 37°C in a humid environment of 5% CO2 for 24 h.

[0074] F2. Add 100 μL of ZIF-67 HS suspension at different concentrations (0.8-400 μg / mL) to each well and incubate for another 24 h;

[0075] F3. Add 10 μL of CCK-8 solution to each well and continue incubation for 1 h;

[0076] F4. Use an ELISA reader to measure absorbance at 450 nm to determine cell viability.

[0077] Hemolysis test: Using rabbit erythrocytes as a model, the blood compatibility of ZIF-67 HS was evaluated.

[0078] F5. Fresh rabbit red blood cells were obtained by centrifugation (8000 rpm, 5 min), washed three times with PBS, and a red blood cell suspension with a volume concentration of 4% was prepared.

[0079] F6. Different concentrations of ZIF-67 HS samples were added to red blood cells to achieve a final volume of 1 ml;

[0080] F7. Incubate at 37°C and 150 rpm for 4 h in a shaker;

[0081] F8. After centrifuging the sample at 4000 rpm for 5 min, collect 200 μL of the supernatant into a 96-well plate and measure the OD using a microplate reader. 540 ;

[0082] F9. Positive controls were treated with 0.2% Triton X-100, and negative controls were treated with PBS.

[0083] Mouse body weight: Changes in mouse body weight during administration can also be used to assess the biosafety of ZIF-67 HS.

[0084] Test results are as follows Figure 6 As shown, compared with the control group, cell viability did not change significantly with increasing drug concentration. Even at a dose of 2 MIC (12.5 μg / mL) of ZIF-67 HS, almost all bacteria were eradicated, and the viability of HepG2 and HEK293 cells remained above 90% and 80%, respectively, indicating negligible cytotoxicity. Regarding blood compatibility, the hemolytic activity of ZIF-67 HS at concentrations between 0.5 MIC and 2 MIC was negligible, with a maximum hemolysis rate of less than 2%. Furthermore, no significant weight loss or even weight gain was observed during ZIF-67 HS treatment in S. aureus-infected wound model mice. Based on this, ZIF-67 HS has been demonstrated to be biocompatible at effective bactericidal doses.

[0085] Implementation Example 7. Molecular Mechanism Analysis of ZIF-67 HS Antibacterial Properties

[0086] The specific testing steps are as follows:

[0087] G1. Dilute the bacterial suspension that has been cultured overnight with culture medium at a ratio of 1:100 and incubate at 37°C for 4 h;

[0088] G2. Co-culture the bacterial culture with 1×MIC ZIF-67 HS for 6 h (the control group was treated the same without drug administration).

[0089] G3. After centrifugation and washing with PBS three times, the obtained samples were flash-frozen in liquid nitrogen for 15 min.

[0090] G4. The sample should be stored at -80°C for further analysis.

[0091] G5. Metabolomics: Accurately weigh 50 mg of bacterial precipitate, add 400 μL of extraction buffer (methanol:water = 4:1 (v:v), with 0.02 mg / mL L-2-chlorophenylalanine as an internal standard), grind, sonicate, vortex, and centrifuge to extract metabolites. Transfer the obtained supernatant to a sample vial for analysis by liquid chromatography-mass spectrometry / LC-MS / MS.

[0092] G6. Transcriptomics: Total RNA was extracted from Staphylococcus aureus using the CTAB method. RNA quality, quantity, and integrity were assessed using a Nanodrop 2000, spectrophotometer, agarose gel electrophoresis, and an Agilent 5300 Fragment Analyzer system. Sequencing libraries were constructed from high-quality samples.

[0093] G7. Metallomics: Bacterial culture samples exposed to different concentrations of ZIF-67 HS (0, 1 / 4 MIC, 1 / 2 MIC, MIC, 2 MIC) were centrifuged and washed three times with PBS. HNO3 (65%) and H2O2 (30%) (1:1, v / v) were added, and the samples were digested and diluted in an 80°C water bath. Elemental analysis was performed by inductively coupled plasma mass spectrometry (ICP-MS).

[0094] G8. Combine the results of transcriptomics, metabolomics and metallomics analysis with relevant validation experiments to explore the molecular mechanism of ZIF-67HS antibacterial activity.

[0095] Test results are as follows Figure 7 As shown, ZIF-67 HS disrupts amino acid and iron balance by promoting amino acid synthesis and metabolism, leading to amino acid accumulation within bacteria. Simultaneously, it utilizes Co ions to replace Fe, creating an iron-limited environment. Furthermore, these nanoparticles interfere with the phosphotransferase system and ABC transporter processes while reducing the secretion of *S. aureus* virulence factors, thereby hindering membrane transport and inhibiting *S. aureus* infection. Finally, increased levels of reactive oxygen species and decreased levels of glutathione impair the antioxidant system, inducing oxidative stress. The results indicate that the combined interaction between ZIF-67 HS and all of the above targets ultimately makes bacteria more susceptible to death.

Claims

1. A zeolite imidazole skeleton-67 antibacterial material (ZIF-67 HS), characterized in that, The antibacterial agent has a hollow spherical structure; the particle size of the antibacterial agent is 655.2±34.5 nm.

2. The ZIF-67 HS antibacterial material according to claim 1, characterized in that, The overall Co content of the material is 24.99±0.19 (wt. %).

3. The ZIF-67 HS antibacterial material according to claim 1, characterized in that, The material surface exhibits a positive charge of 32.26±1.59 mV, indicating that it is more likely to adhere to the negatively charged bacterial surface.

4. The ZIF-67 HS antibacterial material according to claim 1, characterized in that, Materials were tested in LB medium and ultrapure water using Co 2+ The release is stable, and there is a higher release trend in the culture medium.

5. A method for preparing a ZIF-67 HS antibacterial material as described in any one of claims 1-4, characterized in that, Hexadecyltrimethylammonium bromide (CTAB) was dissolved in water under magnetic stirring in a water bath at 30°C until the solution became clear and transparent, forming a spherical CTAB / water micelle system. Then, Co(NO3)2·6H2O solution was added and stirring continued. After uniform dispersion, 2-methylimidazole solution was quickly poured in and reacted at 30°C for 2 h. The resulting purple product was washed by centrifugation with N,N-dimethylformamide and methanol, and then dried under vacuum at 60°C to obtain a purple powder.

6. The ZIF-67 HS antibacterial material according to claim 1, characterized in that, The bacteria in question are Gram-positive Staphylococcus aureus.

7. The ZIF-67 HS antibacterial material according to claim 1, characterized in that, The material exhibits good biocompatibility and biosafety.

8. The ZIF-67 HS antibacterial material according to claim 1, characterized in that, The material achieves its antibacterial effect by acting on bacteria through multiple targets, including interfering with amino acid and iron metabolism, hindering membrane transport, inhibiting Staphylococcus aureus infection, and inducing redox imbalance.

9. The application of the antibacterial nanomaterial as described in claim 1 in effectively killing Staphylococcus aureus without inducing drug resistance.

10. The application of the antibacterial nanomaterial as described in claim 1 in effectively promoting wound healing caused by Staphylococcus aureus infection.

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