Preparation method and application of an antibacterial and healing-promoting photosensitizer

Abstract

This invention discloses an antibacterial and wound-healing photosensitizer, its preparation method, and its applications. The photosensitizer of this invention has a core-shell structure, with the core being MIL-88B modified with amino / bromodiphenyl ligands embedded in silver nanoparticles, and the shell being MIL-88C grown in situ on the core surface. The photosensitizer of this invention significantly improves the photogenerated carrier separation efficiency and reactive oxygen species generation capacity through its MOF-on-MOF heterostructure. Under light irradiation, it exhibits highly efficient killing effects against Staphylococcus aureus, Escherichia coli, and methicillin-resistant Staphylococcus aureus, while also possessing good biocompatibility, promoting fibroblast migration, and showing significant hemostatic and wound-healing effects. This photosensitizer has broad application prospects in the preparation of photodynamic antibacterial drugs and wound-healing medical materials.

Description

Technical Field

[0001] This invention belongs to the field of biomedical materials technology, specifically relating to a method for preparing an antibacterial and healing-promoting photosensitizer and its application. Background Technology

[0002] The information disclosed in this background section is intended only to enhance understanding of the overall background of the invention and is not necessarily to be construed as an admission or in any way implying that such information constitutes prior art known to those skilled in the art.

[0003] Intact skin serves as a natural barrier against microbial invasion and prevents moisture loss. When skin is damaged, its protective function is lost, allowing microorganisms to easily invade and colonize the wound, leading to serious wound infections. Currently, antibiotics are the primary means of treating infected wounds in clinical practice. However, the long-term and inappropriate use of antibiotics has led to the continuous emergence of drug-resistant strains, which not only reduces treatment effectiveness but may even foster "superbugs," posing a serious threat to public health. Therefore, developing new anti-infection strategies that do not rely on traditional antibiotics and are less likely to induce drug resistance is of significant clinical importance.

[0004] Photodynamic therapy (PDT), as an emerging anti-infective method, has attracted widespread attention due to its low likelihood of inducing drug resistance. PDT works by activating a photosensitizer with a specific wavelength of laser light to generate reactive oxygen species (ROS), thereby killing pathogenic microorganisms. However, existing photosensitizers generally suffer from problems such as narrow light absorption range and easy recombination of photogenerated carriers, which limit their antibacterial efficiency and application effects.

[0005] Metal-organic frameworks (MOFs) are porous crystalline materials formed by the self-assembly of metal ions and organic ligands, possessing ultra-high specific surface area, tunable pore size, and good biocompatibility. Iron-based MOFs (Fe-MOFs) have shown potential in the antibacterial field due to their ability to generate reactive oxygen species (ROS) to inhibit bacterial growth. However, single Fe-MOF materials suffer from poor stability, low ROS generation efficiency, and potential drug burst release. Therefore, how to structurally design and functionally modify Fe-MOFs to enhance their photodynamic properties and achieve synergistic antibacterial and hemostatic functions is a pressing technical problem to be solved in this field. Currently, there are no reports on technologies that construct core-shell Fe-MOF photosensitizers through dual-ligand competitive modification, silver nanoparticle loading, and in-situ growth of a second MOF to achieve synergistic functions of efficient photodynamic antibacterial, cell migration promotion, and rapid hemostasis. Based on this, this invention develops a novel antibacterial and hemostatic MOF-based photosensitizer through structural modification and composite modification, overcoming the shortcomings of existing technologies. Summary of the Invention

[0006] This invention aims to overcome the shortcomings of the prior art and provide an Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer with high efficiency photodynamic antibacterial activity, good biocompatibility and wound healing promotion, as well as its preparation method and application.

[0007] To achieve the above objectives, the present invention provides the following technical solution: In a first aspect, the present invention provides an Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer, characterized in that the photosensitizer has a core-shell structure: The core is MIL-88B modified with amino / bromine dual ligands embedded in silver nanoparticles (AgNPs), namely Ag / NH2 / Br-MIL-88B; the shell is MIL-88C, which is grown in situ on the surface of the core.

[0008] Secondly, the present invention provides a method for preparing the above-mentioned Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer, comprising the following steps: Step S1: Synthesis of the dual-ligand NH2 / Br-MIL-88B (NB-B): Iron source, 2-aminoterephthalic acid and 2-bromoterephthalic acid were dissolved in an organic solvent and reacted with a solvothermal agent to obtain NH2 / Br-MIL-88B; Step S2: Synthesis of Ag / NH2 / Br-MIL-88B (ANB-B): The NH2 / Br-MIL-88B obtained in step S1 is dispersed in a solvent, and a silver salt solution is added. Ag⁺ is reduced to AgNPs through a reduction reaction and anchored on NH2 / Br-MIL-88B to obtain Ag / NH2 / Br-MIL-88B. Step S3: In-situ growth of the MIL-88C shell: The Ag / NH2 / Br-MIL-88B and 2,6-naphthalenedicarboxylic acid obtained in step S2 are dispersed in an organic solvent and subjected to a solvothermal reaction to allow MIL-88C to grow in situ on the surface of Ag / NH2 / Br-MIL-88B, thereby obtaining the Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer.

[0009] As a preferred embodiment, in step S1, the mass ratio of 2-aminoterephthalic acid to 2-bromoterephthalic acid is 9:1. The iron source is FeCl3·6H2O, and the organic solvent is N,N-dimethylformamide (DMF); the solvothermal reaction temperature is 100-120℃, and the time is 12-24 hours.

[0010] As a preferred embodiment, in step S2, the silver salt is silver nitrate, and the mass ratio of silver to NH2 / Br-MIL-88B is (2%-20%):1, preferably 5%:1. The reduction reaction is achieved using DMF, and the reaction is carried out in the dark with stirring for approximately 1 hour.

[0011] As a preferred embodiment, in step S3, the mass ratio of Ag / NH2 / Br-MIL-88B to 2,6-naphthalenedicarboxylic acid is (1-2):1, specifically 640.0 mg:520.0 mg. The solvothermal reaction temperature is 120-140℃, and the reaction time is 0.5-12 hours; by controlling the reaction time, composite materials with different shell coating degrees can be obtained. Preferably, the reaction time is 1 hour, yielding 1-Ag / NH2 / Br-MIL-88B@MIL-88C with a complete core-shell structure and uniform morphology.

[0012] Thirdly, the present invention provides the application of the Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer in the preparation of products with photodynamic antibacterial, cell migration promotion, hemostatic, or wound healing promotion functions.

[0013] Preferably, the bacteria include, but are not limited to, Staphylococcus aureus, Escherichia coli, and methicillin-resistant Staphylococcus aureus (MRSA).

[0014] Compared with the prior art, the technical advantages of the present invention are as follows: 1. Structural Innovation: A novel MOF-on-MOF core-shell heterostructure was constructed by modifying MIL-88B using a dual-ligand competition strategy (NH2 / Br) and embedding AgNPs, followed by in-situ growth of a MIL-88C shell. This structure not only improves the stability of the material but also promotes the separation and migration of photogenerated carriers.

[0015] 2. Excellent photodynamic performance: The 1ANB-B@C material prepared in this invention exhibits significantly enhanced instantaneous photocurrent response, lower electrochemical impedance, higher photocurrent density, and characteristic ESR signals of ¹O2, ·OH, and ·O2⁻ that increase with illumination time, demonstrating its excellent photocatalytic activity.

[0016] 3. Broad-spectrum and highly effective antibacterial activity: Under LED light, this photosensitizer exhibits extremely high kill rates against Gram-positive bacteria (Staphylococcus aureus), Gram-negative bacteria (Escherichia coli), and clinically resistant bacteria (MRSA) (reaching 98.3%, 99.6%, and 92.1%, respectively), overcoming the problem of antibiotic resistance.

[0017] 4. Excellent biocompatibility and multifunctionality: MTT assay confirmed that the material had no significant cytotoxicity to L929 cells (cell viability >80%) and significantly promoted fibroblast migration. Animal experiments showed that the material also had excellent hemostatic ability, significantly reducing blood loss in the tail and liver of mice and accelerating the healing of skin wounds in rats. Attached Figure Description

[0018] Figure 1 XRD patterns of a series of materials.

[0019] Figure 2 SEM images of (a) NH2-B, (b) Br-B, (c) NB-B, (d) 5%ANB-B, (e) MIL-88C and (fl) ANB-B@C series; EDS image of 1ANB-B@C.

[0020] Figure 3 NH2-B, Br-B, NB-B, 5%ANB-B, MIL-88C, and 1ANB-B@C: (a) Instantaneous photocurrent response plot, (b) AC impedance plot, and (c) LSV plot; (d) TEMP- 1 O2, (e) DMPO-‧OH and (f) DMPO-‧O2 - The EPR spectrum.

[0021] Figure 4 Image showing the culture of (a) E. coli, S. aureus and (b) MRSA plates after illumination.

[0022] Figure 5 (a) Cytotoxicity assay of 1ANB-B@C; (b) cytotoxicity assay of NH2-B, Br-B, NB-B, 5%ANB-B, MIL-88C, and 1ANB-B@C; and (c) streak plot of L929 co-culture promoting L929 migration.

[0023] Figure 6 :1. Hemostatic performance test of ANB-B@C and ANB-B@C gel on mouse tail and liver. Detailed Implementation

[0024] To make the objectives and technical solutions of this invention clearer, the following embodiments are provided for further explanation. However, the scope of protection of this invention is not limited to these embodiments; the embodiments are merely for illustrative purposes. Those skilled in the art should understand that any changes or equivalent substitutions that do not depart from the concept of this invention are included within the scope of protection of this invention.

[0025] Example 1: Preparation of a series of materials 1. Synthesis of the single ligand MIL-88B MIL-88B: Dissolve 838.0 mg FeCl3·6H2O and 171.0 mg terephthalic acid in 40 mL N,N-dimethylformamide (DMF). After stirring for 1 hour, heat at 110 °C for 18 hours. After the reaction is complete, wash and dry to obtain MIL-88B.

[0026] NH2-MIL-88B (NH2-B): The synthesis steps are the same as above, except that terephthalic acid is replaced with 2-aminoterephthalic acid.

[0027] Br-MIL-88B (Br-B): The synthesis steps are the same as above, except that terephthalic acid is replaced with 2-bromoterephthalic acid.

[0028] 2. Synthesis of the dual-ligand NH2 / Br-MIL-88B (NB-B) Terephthalic acid was replaced with a mixture of 2-aminoterephthalic acid and 2-bromoterephthalic acid, with the total mass remaining the same as that of terephthalic acid, while keeping the iron source and other conditions constant. A series of NB-B compounds were synthesized by varying the mass ratio of amino to bromine groups (1:9, 3:7, 5:5, 7:3, 9:1). In this example, a mass ratio of 9:1 is preferred.

[0029] 3. Synthesis of Ag / NH2 / Br-MIL-88B (ANB-B) Ag⁺ was reduced in situ to AgNPs using the reducing properties of DMF. 200.0 mg of NB-B was dispersed in DMF. Different masses of silver nitrate (6.3 mg, 15.8 mg, 31.5 mg, and 63.0 mg, corresponding to Ag / NB-B mass ratios of 2%, 5%, 10%, and 20%) were dissolved in deionized water and added to the NB-B solution. The mixture was stirred in the dark for 1 hour. Subsequently, it was centrifuged, washed, and vacuum dried at 80 °C for 12 hours to obtain ANB-B with different Ag loadings (2% ANB-B, 5% ANB-B, 10% ANB-B, and 20% ANB-B).

[0030] 4. Synthesis of MIL-88C 1286.0 mg FeCl3·6H2O and 104.0 mg 2,6-naphthalenedicarboxylic acid were dissolved in 50 mL DMF and placed in a 250 mL round-bottom flask. The mixture was heated at 130 °C for 12 hours with stirring. The product was collected by centrifugation, washed with DMF and anhydrous ethanol, and dried at 60 °C for 12 hours to obtain MIL-88C.

[0031] 5. Synthesis of Ag / NH2 / Br-MIL-88B@MIL-88C (ANB-B@C) 640.0 mg of 5% ANB-B and 520.0 mg of 2,6-naphthalenedicarboxylic acid were placed in 25 mL of LDM and reacted for different times (0.5, 1, 3, 6, 9, and 12 hours) under reflux and stirring in an oil bath at 130 °C. The products were named 0.5ANB-B@C, 1ANB-B@C, 3ANB-B@C, 6ANB-B@C, 9ANB-B@C, and 12ANB-B@C, respectively. Among them, 1ANB-B@C, obtained after 1 hour of reaction, exhibited a complete core-shell structure and the best performance.

[0032] Example 2: Material Characterization The crystal planes and structural information of the sample were obtained by XRD spectroscopy. Figure 1 (a) shows that all samples exhibit similar diffraction peaks in the 5-20° range, which are highly consistent with the diffraction pattern of MIL-88B, confirming that the modified material successfully inherited the crystal structure of the MIL-88B framework. The diffraction peaks at 9.0°, 10.0°, 16.5° and 18.7° correspond to the (002), (101), (103) and (200) crystal planes, respectively. This also indicates that the substitution of -H groups in the ligand framework by other ligands has a negligible effect on the framework structure. This structural stability provides an ideal topological framework for subsequent surface functionalization modification. The significant peak detected at 2θ value 77.3° indicates that the (311) crystal plane of silver ( Figure 1 (b) confirms the successful incorporation of Ag NPs into MIL-88B. The diffraction peaks at 27.8°, 32.3°, and 46.2° may be attributed to the formation of silver chloride (111), (200), and (220). Figure 1 As shown in (c), ANB-B@C exhibits characteristic peaks of ANB-B, as well as peaks at 7.5° and 14.9° corresponding to the (002) and (004) crystal planes of MIL-88C. With prolonged reaction time, the ANB-B peak changes while the MIL-88C peak intensifies, which can be attributed to the in-situ transformation of 5% ANB-B to MIL-88C.

[0033] The morphological changes of MIL-88B during recombination and transformation were analyzed using scanning electron microscopy (SEM). Both NH2-B and Br-B exhibited octahedral structures with smooth surfaces and diameters ranging from approximately 500 nm to 1 μm. Figure 2 (ab)). The binary ligand NB-B retains structural features similar to the monomer. When NB-B is loaded with Ag NPs, the surface of ANB-B exhibits a dense and slightly rough texture. Figure 2 (cd)). Subsequently, using 5% ANB-B as a substrate, without adding any additional iron source, the Fe on the surface of 5% ANB-B was disrupted. 3+ Coordination bonds between Fe and ligands 3+An in-situ reaction with 2,6-naphthalenedicarboxylic acid occurred, and MIL-88C was recombined on the surface of 5% ANB-B to construct an ANB-B@C core-shell structure. The reaction was monitored at different times (…). Figure 2 (fk) Tracking the transformation process: Within 1 hour of the reaction, protrusions appear on the octahedral surface due to the conversion of MIL-88C attaching to the outer surface of 5% ANB-B; after 1 hour, the outer surface of 5% ANB-B is coated with MIL-88C, forming a 1ANB-B@C core-shell structure; as time progresses (1-12 hours), MIL-88C continues to grow on the octahedral surface, and 5% ANB-B gradually disintegrates. Ultimately, blocky ANB-B@C is formed. Figure 2 (f) The basic form of 5% ANB-B can be retained by 1ANB-B@C. Aggregates of 1ANB-B@C are as follows: Figure 2 As shown in (l). In Figure 2 The presence of C, N, O, Fe, Ag and Br elements in (m) further confirms the successful preparation of 1ANB@C.

[0034] Example 3: Evaluation of photocatalytic performance Figure 3 (a) shows the instantaneous photocurrent response of NH2-B, Br-B, NB-B, 5%ANB-B, MIL-88C, and 1ANB-B@C. It can be seen that 1ANB-B@C has the best instantaneous photocurrent response. Figure 3 (b) has the lowest impedance of 1ANB-B@C. Figure 3 (c) shows that 1ANB-B@C has a high photocurrent density, indicating that it has the best photocatalytic activity. Figure 3 The ROS level in (df) was tested to verify the photocatalytic performance of 1ANB-B@C. With prolonged illumination time, the ROS levels in 1ANB-B@C... 1 O2, ·OH and ·O2 - The ESR signal intensity of the materials all showed an increasing trend, indicating that the photocatalytic efficiency of the material has good time-dependent characteristics.

[0035] Example 4: Evaluation of in vitro antibacterial properties Three bacterial strains were selected as model strains and treated with NB-B, 5% ANB-B, MIL-88C, and 1ANB-B@C under dark or light conditions, respectively. Figure 4The results showed that bacteria grew well under both dark and light conditions, indicating that light had little effect on bacterial activity. The addition of NB-B or MIL-88C led to a decrease in colony counts of *S. aureus* and MRSA. This antibacterial effect may be attributed to the adsorption of the material under dark conditions and the relatively low photocatalytic activity under light conditions. The antibacterial efficiency of 5% ANB-B was significantly higher than that of NB-B, confirming that its excellent antibacterial effect stemmed from the bactericidal effect of silver and its good photodynamic antibacterial properties under light. Notably, 1ANB-B@C exhibited the best antibacterial performance against *E. coli* (99.6%, 60 min), *S. aureus* (98.3%, 90 min), and MRSA (92.1%, 60 min), which can be attributed to the synergistic effect of the material's adsorption of bacteria and its highly efficient photodynamic activity.

[0036] Example 5: Cell Viability and Migration Assay like Figure 5 As shown in (ab), the MTT assay demonstrated that the sustained metabolic activity of L929 cells exceeded the 80% survival threshold, confirming the absence of acute cytotoxic effects. The scratch assay was performed using L929 fibroblasts. Figure 5 (c) Compared with the control group, the cells in the 1ANB-B@C treatment group showed significantly enhanced ability to migrate to the scratch area, indicating that 1ANB-B@C can promote cell migration and accelerate wound healing.

[0037] Example 6: In vivo hemostasis and wound healing experiment Hemostasis experiment: In mouse tail and liver hemorrhage models, the blood loss in the 1ANB-B@C group (tail weight loss 7.49 mg, liver weight loss 10.19 mg) and the ANB-B@C gel group (tail weight loss 14.42 mg, liver weight loss 13.82 mg) was significantly lower than that in the control group (tail weight loss 43.02 mg, liver weight loss 48.86 mg). Figure 6 As shown.

[0038] Rat skin defect model: A 10 mm diameter full-thickness skin wound was created on the back of rats and infected with Staphylococcus aureus. The experimental groups are shown in Figure 7. On day 12, the 1ANB-B@C (Light) group and the ANB-B@C gel (Light) group had the highest wound healing rates, demonstrating the best wound healing promotion ability.

Claims

1. An Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer, characterized in that, The photosensitizer has a core-shell structure; the core is MIL-88B modified with an amino / bromine dual ligand embedded in silver nanoparticles, namely Ag / NH2 / Br-MIL-88B; the shell is MIL-88C, which is grown in situ on the surface of the core.

2. The Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer according to claim 1, characterized in that, In the amino / bromodiphenyl ligand modified MIL-88B, the amino ligand is 2-aminoterephthalic acid, the bromodiphenyl ligand is 2-bromoterephthalic acid, and the mass ratio of the amino ligand to the bromodiphenyl ligand is 1:9, 3:7, 5:5, 7:3, or 9:

1. Preferably, the mass ratio of the amino ligand to the bromodiphenyl ligand is 9:

1.

3. The Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer according to claim 1 or 2, characterized in that, The mass ratio of the silver nanoparticles to the amino / bromine dual-ligand modified MIL-88B is 2% to 20%:1; preferably, the mass ratio is 5%:

1.

4. A method for preparing the Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer according to any one of claims 1-3, characterized in that, Includes the following steps: Step S1: Synthesis of the dual-ligand NH2 / Br-MIL-88B: Iron source, 2-aminoterephthalic acid and 2-bromoterephthalic acid were dissolved in an organic solvent and reacted with a solvothermal agent to obtain NH2 / Br-MIL-88B; Step S2: Synthesis of Ag / NH2 / Br-MIL-88B: The NH2 / Br-MIL-88B obtained in step S1 is dispersed in a solvent, and a silver salt solution is added. Ag⁺ is reduced to Ag NPs through a reduction reaction and anchored on NH2 / Br-MIL-88B to obtain Ag / NH2 / Br-MIL-88B. Step S3: In-situ growth of the MIL-88C shell: The Ag / NH2 / Br-MIL-88B and 2,6-naphthalenedicarboxylic acid obtained in step S2 are dispersed in an organic solvent and subjected to a solvothermal reaction to allow MIL-88C to grow in situ on the surface of Ag / NH2 / Br-MIL-88B, thereby obtaining the Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer.

5. The preparation method according to claim 4, characterized in that, In step S1, the iron source is FeCl3·6H2O; the organic solvent is N,N-dimethylformamide; the mass ratio of 2-aminoterephthalic acid to 2-bromoterephthalic acid is 1:9~7:3; the solvothermal reaction temperature is 100~120℃, and the time is 12~24 hours.

6. The preparation method according to claim 4, characterized in that, In step S2, the silver salt is silver nitrate; the mass ratio of silver to NH2 / Br-MIL-88B in the silver salt is 2%~20%:1, preferably 5%:1; the reduction reaction is carried out under dark conditions for 0.5~2 hours.

7. The preparation method according to claim 4, characterized in that, In step S3, the mass ratio of Ag / NH2 / Br-MIL-88B to 2,6-naphthalenedicarboxylic acid is 1~2:1; the solvothermal reaction temperature is 120~140℃, and the reaction time is 0.5~12 hours; preferably, the reaction time is 1 hour.

8. The application of the Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer according to any one of claims 1-3 in the preparation of photodynamic antibacterial products.

9. The application according to claim 8, characterized in that, The photodynamic antibacterial product targets at least one of Staphylococcus aureus, Escherichia coli, and methicillin-resistant Staphylococcus aureus.

10. The use of the Ag / NH2 / Br-MIL-88B@MIL-88C photosensitizer according to any one of claims 1-3 in the preparation of drugs or medical materials that promote cell migration, hemostasis, or wound healing.

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