An antibacterial material with calcium ion mediation and photothermal effect and a preparation method and application thereof

Abstract

The application discloses antibacterial material with calcium ion mediation and photothermal effect, and a preparation method and application thereof, and belongs to the technical field of biological medicine. The antibacterial material is calcium ion loaded porous covalent organic framework polymer BQ-Ca-COF. 2,3,5,6-tetraaminocyclohex-2,5-diene-1,4-diketone is subjected to a solvothermal reaction with cyclohexanhexanone octahydrate to obtain the porous covalent organic framework polymer; under a protective atmosphere, the porous covalent organic framework polymer is subjected to a metallization reaction with calcium chloride to obtain the BQ-Ca-COF. The BQ-Ca-COF prepared in the application has controllable release of Ca 2+ and synergistic effect of photothermal irradiation under 638 nm wavelength laser irradiation, good photothermal conversion, release of calcium ions under infrared (NIR) laser irradiation, increased permeability of bacterial membranes, and synergistically enhanced efficiency of subsequent photothermal effect, so that gram-positive and gram-negative bacteria can be effectively killed.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to an antibacterial material with calcium ion-mediated and photothermal effects, its preparation method, and its application. Background Technology

[0002] Drug-resistant infections directly cause over a million deaths annually. The continued spread of resistance genes is constantly weakening the efficacy of traditional antibiotics, and the biofilm-forming ability of bacteria exacerbates this crisis. Biofilms endow bacteria with remarkable antibiotic resistance and anti-host immunity, making related infections extremely difficult to eradicate, often leading to chronic diseases, prolonged hospital stays, and increased mortality. The dual threat of free-floating "superbugs" and persistent biofilms underscores the urgency of developing non-antibiotic treatment strategies. To address this challenge, researchers have explored various alternative antimicrobial approaches; however, each method faces significant bottlenecks. While metal ions are potent, they raise concerns about long-term biosafety and environmental accumulation. Organic antimicrobial agents are often limited by volatility, photodegradability, and short-lived effects. Gas therapy faces challenges in precise delivery and dosage control. Crucially, most of these strategies lack specificity against bacterial cells, potentially inadvertently damaging host tissues, thus delaying wound healing and causing secondary damage. Furthermore, these methods generally have limitations in their effectiveness against biofilms.

[0003] Photothermal therapy (PTT) has shown great potential as a non-invasive alternative to traditional antibiotics. By converting light energy into localized heat, PTT physically disrupts bacterial membrane integrity, inducing rapid cell death. Its unique mechanism of action makes it less prone to inducing drug resistance; however, its efficacy has several inherent limitations. To overcome these challenges, pre-sensitization of bacterial membranes has become a feasible strategy. Increasing bacterial membrane permeability can lower the temperature threshold required for effective bactericidal action, thereby reducing the risk of damage to surrounding healthy tissues. Currently, there are various methods to increase bacterial membrane permeability, such as using cationic polymers or cationic molecules to interact with the bacterial membrane through cationic groups, thereby disrupting the bacterial membrane; or using lysozyme (which degrades peptidoglycan) or enzymes targeting the outer membrane of Gram-negative bacteria to disrupt the chemical stability of the membrane structure; and also using calcium ions (Ca... 2+ Biological ions such as those involved in cell membrane destruction can cause damage.

[0004] Ca 2+ It can interact with phospholipids in bacterial membranes, disrupting their structural integrity and enhancing permeability. However, the key to this strategy lies in material design, requiring the ability to precisely control Ca... 2+ The release and action of Ca. This ensures Ca 2+Applying the appropriate dosage at the right time to the bacterial membrane enhances the membrane disruption effect while avoiding unnecessary biological interference. Synergistically combining this with photothermal effects can create a more potent and precise antibacterial system. Therefore, it is necessary to achieve Ca... 2+ Controllable loading and release of ions, through Ca 2+ By embedding a customized COF framework, a composite material was constructed that combines strong endogenous photothermal properties with tunable calcium ion membrane disruption. However, designing a controllable delivery system that meets these requirements remains a significant research challenge. Summary of the Invention

[0005] In view of the above-mentioned prior art, the purpose of this invention is to provide an antibacterial material with calcium ion-mediated and photothermal effects, its preparation method, and its application. This invention uses cyclohexanehexanone as the basic unit, constructs a basic covalent organic framework with an aromatic tetraamine, and then performs calcium metallization to finally obtain an antibacterial material with calcium ion-mediated and photothermal effects. 2+ BQ-Ca-COF is a metal-covalent organic framework material with controlled release and synergistic photothermal irradiation. BQ-Ca-COF exhibits good photothermal conversion under 638 nm wavelength laser irradiation, and can release calcium ions under infrared (NIR) laser irradiation, increasing the permeability of bacterial membranes and synergistically enhancing the effectiveness of subsequent photothermal effects, thereby effectively killing Gram-positive and Gram-negative bacteria.

[0006] To achieve the above objectives, the present invention adopts the following technical solution:

[0007] In a first aspect, the present invention provides an antibacterial material having calcium ion-mediated and photothermal effects, wherein the antibacterial material is a calcium ion-supported porous covalent organic framework polymer; the porous covalent organic framework polymer is obtained by copolymerization of cyclohexanehexanone and aromatic tetraamine as monomers.

[0008] Preferably, the aromatic tetraamine is 2,3,5,6-tetraaminocyclohexane-2,5-diene-1,4-dione.

[0009] Preferably, the antibacterial material has the following structural formula:

[0010] .

[0011] A second aspect of the present invention provides a method for preparing the above-mentioned antibacterial material, comprising the following steps:

[0012] (1) 2,3,5,6-tetraaminocyclohexane-2,5-diene-1,4-dione and cyclohexanehexane octahydrate were added to an organic solvent for a solvothermal reaction. After the reaction was completed, the polymer was separated, washed and dried in sequence to obtain a porous covalent organic framework polymer.

[0013] (2) Under a protective atmosphere, the porous covalent organic framework polymer is dispersed in distilled water, and then an excess of calcium salt is added to obtain a dispersion. The dispersion is stirred to carry out a metallization reaction. After the reaction is completed, the polymer is centrifuged, washed, and dried in sequence to obtain an antibacterial material with calcium ion-mediated and photothermal effects.

[0014] Preferably, in step (1), the molar ratio of 2,3,5,6-tetraaminocyclohexane-2,5-diene-1,4-dione to cyclohexanehexaone octahydrate is 1.5:1; the organic solvent is a mixture of ethylene glycol and acetic acid solution; and the concentration of the acetic acid solution is 6M.

[0015] Preferably, in step (1), the temperature of the solvothermal reaction is 120°C and the time is 72h; the separation is first ultrasonically extracted with distilled water for 24h, and then Soxhlet extracted with acetone for 24h.

[0016] Preferably, in step (2), the calcium salt is calcium chloride; the concentration of the porous covalent organic framework polymer in the dispersion is 1 mg / mL and the concentration of the calcium salt is 100 mM.

[0017] Preferably, in step (2), the metallization reaction is carried out at a temperature of 25°C for 5 days.

[0018] A third aspect of the present invention provides the use of antimicrobial materials in the preparation of antimicrobial drugs.

[0019] Preferably, the antibacterial material releases Ca2+ under infrared laser irradiation. 2+ It also produces a photothermal effect, and the two work together to achieve antibacterial effect.

[0020] The beneficial effects of this invention are:

[0021] (1) The BQ-Ca-COF prepared in this invention has good photothermal conversion under 638 nm wavelength laser irradiation. At the same time, it can release calcium ions under infrared (NIR) laser irradiation, increase the permeability of bacterial membrane, and synergistically enhance the efficiency of subsequent photothermal action, thereby effectively killing Gram-positive and Gram-negative bacteria.

[0022] (2) The preparation method of the present invention is simple, the prepared BQ-Ca-COF has good biocompatibility, low hemolysis rate of red blood cells, minimal impact on the viability of 3T3 cells, and no toxic side effects on the human body, which will promote the development of multifunctional antibacterial platforms. Attached Figure Description

[0023] Figure 1 (a) Infrared spectra of CHHO, TABQ and BQ-Ca-COF; (b) Solid-state carbon spectrum of BQ-Ca-COF. 13(c) CNCMR; (d) Low-temperature N2 absorption isotherm of BQ-COF at 77 K; (e) CNCMR of BQ-Ca-COF; (f) Low-temperature N2 absorption isotherm of BQ-Ca-COF at 77 K; (g) Thermogravimetric analysis of BQ-Ca-COF; (h) X-ray diffraction pattern of BQ-Ca-COF;

[0024] Figure 2 (a) Transmission electron microscopy (TEM) image of BQ-Ca-COF at a scale of 0.5 μm; (b) Transmission electron microscopy (TEM) image of BQ-Ca-COF at a scale of 100 nm; (c) Transmission electron microscopy (TEM) image of BQ-Ca-COF at a scale of 20 nm; (d) Transmission electron microscopy (TEM) image of BQ-Ca-COF at a scale of 10 nm; (e) Scanning electron microscopy (SEM) image of BQ-Ca-COF at a scale of 2.5 μm; (f) HAADF-STEM image of BQ-Ca-COF; (g) Elemental mapping of C in BQ-Ca-COF; (h) Elemental mapping of O in BQ-Ca-COF; (i) Elemental mapping of N in BQ-Ca-COF; (j) Elemental mapping of Ca in BQ-Ca-COF.

[0025] Figure 3 EDS spectrum of BQ-Ca-COF;

[0026] Figure 4 X-ray photoelectron spectra of BQ-Ca-COF, including (a) XPS analysis spectrum, (b) C 1s spectrum, (c) N 1s spectrum, (d) Ca 2p spectrum, and (e) O 1s spectrum.

[0027] Figure 5 The photothermal properties of BQ-Ca-COF were studied, including (a) the photothermal effect of BQ-Ca-COF (100 µg / mL) at different laser powers; (b) temperature change curves of different materials at the same concentration (100 µg / mL) under laser irradiation at the same power (638 nm, 1.2 W / cm²); (c) the concentration-dependent photothermal effect of BQ-Ca-COF over 10 minutes; (d) the relationship between temperature change and BQ-Ca-COF concentration, and temperature change and power density after 10 minutes of 638 nm laser irradiation; (e) infrared thermal images of BQ-Ca-COF at different concentrations; (f) the temperature change curve of BQ-Ca-COF (100 µg / mL) in five heating-cooling cycles; (g) a comparison of the temperature change curve of BQ-Ca-COF aqueous suspension after 15 days of storage under laser irradiation; and (h) the temperature change of BQ-Ca-COF at 1.0 W / cm².2 Heating-cooling temperature curves of BQ-Ca-COF (100 µg / mL) under laser irradiation, and a graph of cooling time versus the negative natural logarithm of temperature; (i) Calcium ion release concentration curve of BQ-Ca-COF under laser irradiation (laser intensity: 1.2 W / cm²). 2 );

[0028] Figure 6 (a) Colony diagram of Staphylococcus aureus after each treatment group; (b) Survival rate of Staphylococcus aureus after each treatment group determined by plate count method; (c) Colony diagram of Escherichia coli after each treatment group; (d) Survival rate of Escherichia coli after each treatment group determined by plate count method.

[0029] Figure 7 : Concentration gradient sterilization effect diagrams, including (a) sterilization effect diagram of BQ-COF at different concentrations, (b) quantification diagram of BQ-COF by plate count method, (c) sterilization effect diagram of BQ-COF + laser at different concentrations, (d) quantification diagram of BQ-COF + laser by plate count method, (e) sterilization effect diagram of BQ-Ca-COF at different concentrations, (f) quantification diagram of BQ-Ca-COF by plate count method, (g) sterilization effect diagram of BQ-Ca-COF + laser at different concentrations, (h) quantification diagram of BQ-Ca-COF + laser by plate count method, (i) sterilization effect diagram of PT-Ca-COF + laser at different concentrations, (j) quantification diagram of PT-Ca-COF + laser by plate count method, (k) sterilization effect diagram of CE-Ca-COF + laser at different concentrations, and (l) quantification diagram of CE-Ca-COF + laser by plate count method.

[0030] Figure 8 (a) Confocal laser scanning microscopy (CLSM) images of Staphylococcus aureus biofilms after different treatments, (b) Quantitative laser fluorescence (QF) images of Staphylococcus aureus biofilms after different treatments, (c) Confocal laser scanning microscopy (CLSM) images of Escherichia coli biofilms after different treatments, (d) Quantitative laser fluorescence (QF) images of Escherichia coli biofilms after different treatments.

[0031] Figure 9 (a) Biofilm formation test against Staphylococcus aureus, (b) Biofilm formation test against Escherichia coli;

[0032] Figure 10 TEM images of bacteria, including (a) TEM images of Staphylococcus aureus after each treatment group, and (b) TEM images of Escherichia coli after each treatment group; red tips in the images represent the location of damage.

[0033] Figure 11 : Bacterial staining images, of which (a) fluorescence images of Staphylococcus aureus after each treatment incubated with SYTO-9 / PI live / dead staining agent; (b) fluorescence images of Escherichia coli after each group of treatment incubated with SYTO-9 / PI live / dead staining agent;

[0034] Figure 12 (a) Hemolysis rate of BQ-COF at different concentrations; (b) Hemolysis rate of BQ-Ca-COF at different concentrations; (c) Cytotoxicity assessment of BQ-COF and BQ-Ca-COF on 3T3 cells at different concentrations; (d) Representative images of scratch healing experiments in each group over time; (e) Quantitative analysis of cell migration rate based on relative scratch width (migration distance) for each treatment group; (f) Quantitative analysis of cell migration rate based on relative scratch area for each treatment group.

[0035] Figure 13 (a) Wound images and wound closure trajectories of mice on days 1, 3, 5, 7, and 9 after treatment in each group; (b) Changes in body weight of mice in each group during treatment; (c) Percentage of wound area (%) in each group of mice.

[0036] Figure 14 H&E and Masson staining images of wound tissues treated in each group on day 9 of the wound healing process (scale bar: 200 μm).

[0037] Figure 15 Organ staining results for each treatment group;

[0038] Figure 16 Hematological examination results of mice in each group on day 9; including (a) white blood cell count of mice on day 9; (b) red blood cell count of mice on day 9; (c) hemoglobin of mice on day 9; (d) hematocrit of mice on day 9; (e) mean corpuscular hemoglobin (MCH) of mice on day 9; (f) mean corpuscular hemoglobin concentration (MCHC) of mice on day 9; (g) mean corpuscular hematocrit of mice on day 9; (h) platelet count of mice on day 9;

[0039] Figure 17 Synthetic routes of BQ-Ca-COF, PT-Ca-COF and CE-Ca-COF, where (a) is the synthetic route of BQ-COF, PT-COF and CE-COF; and (b) is the route of metallization synthesis of BQ-Ca-COF, PT-Ca-COF and CE-Ca-COF. Detailed Implementation

[0040] It should be noted that the following detailed descriptions are illustrative and intended to provide further explanation of this application. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains.

[0041] As described in the background section, photothermal therapy can physically disrupt the integrity of bacterial membranes, inducing rapid cell death. However, the high temperature of photothermal therapy can easily damage surrounding healthy tissue. If the bacterial membrane can be pre-sensitized, its permeability can be increased, lowering the temperature threshold required for effective sterilization and thus reducing the risk of damage to surrounding healthy tissue.

[0042] Based on this, the purpose of this invention is to provide an antibacterial material with calcium ion-mediated and photothermal effects, its preparation method, and its application. This invention involves adding 2,3,5,6-tetraaminocyclohexane-2,5-diene-1,4-dione and cyclohexanehexaone octahydrate to an organic solvent for a solvothermal reaction to obtain BQ-COF. Then, under nitrogen protection, BQ-COF is uniformly dispersed in distilled water and mixed with excess calcium chloride for calcium metallization. The resulting antibacterial material, BQ-Ca-COF, with calcium ion-mediated and photothermal effects is obtained through a co-precipitation method. This material possesses Ca... 2+ The controlled release and photothermal irradiation work synergistically, exhibiting good photothermal conversion under 638 nm wavelength laser irradiation. Simultaneously, under infrared laser irradiation, the polymer generates heat, causing calcium ions to detach from the polymer and release calcium ions. 2+ It can interact with phospholipids in bacterial membranes, disrupting their structural integrity and increasing permeability, thus synergistically enhancing the effectiveness of subsequent photothermal effects and effectively killing Gram-positive and Gram-negative bacteria. Simultaneously, BQ-Ca-COF exhibits negligible hemolytic activity and minimal damage to normal cells at optimal antibacterial concentrations, indicating its significant potential in biomedicine.

[0043] To enable those skilled in the art to better understand the technical solution of this application, the technical solution of this application will be described in detail below with reference to specific embodiments.

[0044] Note: Unless otherwise specified, all PBS used in this invention is neutral PBS with a pH of 7.4;

[0045] 2,3,5,6-Tetraaminocyclohexane-2,5-diene-1,4-dione, abbreviated as TABQ, CAS: 1128-13-8;

[0046] 2,3,7,8-Tetraaminophenazine-1,4,6,9-tetraone, abbreviated as TAPT, CAS: 2857099-14-8;

[0047] 6,7,9,10,17,18,20,21-octahydrodibenzo[b,k] [1,4,7,10,13,16]hexaoxane-2,3,13,14-tetraamine, abbreviated as TACE, CAS: 122942-39-6;

[0048] Cyclohexanehexanone octahydrate, also known as cyclohexanehexanone hydrate, is abbreviated as CHHO, CAS: 527-31-1, purity > 95%, brand: McLean.

[0049] The test materials used in the embodiments of this invention are all conventional test materials in the art and can be purchased through commercial channels.

[0050] Example 1: Synthesis of BQ-Ca-COF

[0051] (1) Synthesis of BQ-COF

[0052] 55.5 mg of 2,3,5,6-tetraaminocyclohexane-2,5-diene-1,4-dione (TABQ, 0.33 mmol) and 68.6 mg of cyclohexanehexane octahydrate (CHHO, 0.22 mmol) were placed in a pyrolysis tube containing 3 mL of ethylene glycol and 3 mL of glacial acetic acid solution (6 M). The pyrolysis tube was sonicated for 15 minutes to achieve uniform dispersion, followed by degassing through three consecutive freeze-vacuum-thaw cycles. The pyrolysis tube was then transferred to a preheated oil bath and reacted at 65 °C for 12 hours. Subsequently, the reaction temperature was increased to 120 °C and the reaction was carried out for 72 hours. After the reaction, the product was cooled to room temperature and extracted with water by sonication, followed by Soxhlet extraction with acetone, each for one day. The extract was dried under vacuum at 80 °C for 12 hours to obtain a black solid product, which was BQ-COF.

[0053] (2) Metallization

[0054] Under nitrogen protection, 10 mg of synthesized BQ-COF was mixed with excess CaCl2 (110 mg) in 10 mL of deionized water and stirred at room temperature for 5 days. The resulting suspension was centrifuged at 8000 rpm for 10 minutes, washed successively with methanol (3 × 15 mL), tetrahydrofuran (3 × 15 mL), and acetone (3 × 15 mL), and then vacuum dried (80 °C, 12 hours) to obtain BQ-Ca-COF. The synthetic route is shown below. Figure 17 .

[0055] Comparative Example 1: Synthesis of PT-Ca-COF

[0056] The difference from Example 1 is that in step (1), 2,3,5,6-tetraaminocyclohexane-2,5-diene-1,4-dione was replaced with an equimolar amount of 2,3,7,8-tetraaminophenazine-1,4,6,9-tetraone (TAPT, 99.08 mg), and the solvent was replaced with 3 mL of 1,4-dioxane and 3 mL of 1,3,5-trimethylbenzene solution instead of 3 mL of ethylene glycol and 3 mL of glacial acetic acid solution. The final product was denoted as PT-Ca-COF.

[0057] Comparative Example 2: Synthesis of CE-Ca-COF

[0058] The difference from Example 1 is that in step (1), 2,3,5,6-tetraaminocyclohexane-2,5-diene-1,4-dione was replaced with an equimolar amount of 6,7,9,10,17,18,20,21-octahydrodibenzo[b,k][1,4,7,10,13,16]hexaoxane-2,3,13,14-tetraamine (TACE, 138.8 mg). The final product was denoted as CE-Ca-COF.

[0059] Example 2: Product Structure Characterization

[0060] (1) The structure was determined using infrared spectroscopy. 3 mg of BQ-Ca-COF, cyclohexanehexanone, and TABQ were thoroughly ground with dry potassium bromide powder in a mortar while keeping the powder dry. The powder was then placed in a tableting mold and pressed into a transparent, crack-free tablet. The tablet was then placed in an infrared spectroscopy scanner at 4000~400 cm⁻¹. -1 Scan the area 36 times.

[0061] Fourier transform infrared (FTIR) spectra as follows Figure 1 As shown in (a). At approximately 1245cm -1 New, distinct peaks appeared, which were attributed to the stretching vibrations of the C–C=N–C bonds within the formed aromatic nitrogen-containing heterocycle. The characteristic C=O peak (1630 cm⁻¹) -1 ) and -NH2 (3300 cm -1 The synchronous disappearance of the stretching vibration peaks further confirms that these functional groups have been consumed and that key covalent bonds were successfully formed during the skeletal polymerization process. All Ca-modified COFs exhibit significant shifts across multiple vibrational bands, especially in the low-frequency region (400-800 cm⁻¹). -1 The appearance of a distinct new vibrational peak in this region is attributed to Ca-N coordination, indicating that a stable BQ-Ca-COF covalent organic framework material was successfully formed through coupling.

[0062] (2) The carbon skeleton structure of the BQ-Ca-COF complex was further confirmed by solid-state 13C-NMR. Figure 1 (b) The solid-state 13C NMR further revealed the combination of the two compounds, with characteristic peaks at 176, 155 and 142 ppm observed simultaneously. These peaks are attributed to the remaining carbonyl carbon (C=O) in the hydroquinone unit and the imine carbon (C=N) and olefin carbon (C=C) in the newly formed quinoline unit, respectively.

[0063] (3) Figure 1 (d) and Figure 1 (f) The N2 adsorption-desorption curves and pore size distribution curves reveal the pore distribution of the BQ-COF and BQ-Ca-COF composites. The steep adsorption curve of BQ-Ca-COF at low relative pressure (P / P0 < 0.1) indicates the presence of micropores, while the significant hysteresis loop between the adsorption and desorption branches indicates the presence of numerous mesopores. Furthermore, the steep adsorption rise curve at high relative pressure (P / P0 > 0.85) indicates the presence of macropores. The Brunauer–Emmett–Teller (BET) specific surface area calculated based on the NLDFT model of BQ-COF is 17.718 m². 2 g -1 The cumulative pore volume is 0.054 cm³. 3 g -1 In contrast, the specific surface area of ​​BQ-Ca-COF measured by Brunauer-Emmett-Teller (BET) analysis was 16.262 m². 2 g -1 The cumulative pore volume is 0.050 cm³. 3 g -1 The comparison of these parameters between the two materials demonstrates that calcium was successfully loaded onto the COF. Figure 1 (c) and Figure 1 (e) The pore size distribution covers the range from micropores to macropores and is in good agreement with the results of the absorption isotherm, confirming its multi-scale porous structure.

[0064] (4) The thermal stability of BQ-Ca-COF was determined through thermogravimetric analysis. Thermogravimetric analysis is as follows: Figure 1 As shown in (g), when BQ-Ca-COF is heated from room temperature to 800°C, it undergoes a multi-stage decomposition process, exhibiting a gradual and continuous mass loss throughout the temperature range. At 100°C, it retains approximately 90% of its initial mass, and the mass loss continues steadily until 800°C, with the final residue stabilizing at approximately 38% of its original mass.

[0065] (5) The crystallinity and porosity of BQ-Ca-COF were estimated using powder-X-ray diffraction (PXRD). Figure 1As shown in (h), BQ-Ca-COF exhibits a broad peak at approximately 25°, indicating that BQ-Ca-COF possesses amorphous properties.

[0066] (6) Observe the morphology of BQ-Ca-COF using SEM and TEM and derive the elemental analysis diagram of BQ-Ca-COF and the atomic content of each element. Figure 2 (a)- Figure 2 (d) Transmission electron microscopy (TEM) images show that its bulk structure consists of stacked thin lamellae that fuse into irregular blocky aggregates with distinctly rough edges and clear outlines. Figure 2 (e) and Figure 2 (f) Scanning electron microscopy (SEM) images and HAADF-STEM images highlight the surface morphology, revealing aggregated granular or lamellar structures with rough textures and sharp edges. For example... Figure 3 As shown, elemental mapping and corresponding energy dispersive spectroscopy (EDS) confirmed that N (22.8%), O (17.7%), and Ca (2.7%) were uniformly distributed in the carbon-dominated polymer matrix; Figure 2 (g)~ Figure 2 The element distribution of (j) is consistent.

[0067] (7) XPS spectral analysis of BQ-Ca-COF. For example... Figure 4 As shown in (a), similar to the EDS results, XPS also showed the presence of C, N, O and Ca. Figure 4 (b) The C 1s spectrum shows peaks at 284.8 eV and 284.48 eV, which are attributed to C-C bonds and C=C bonds, respectively. Figure 4 (c) The N 1s spectrum shows three characteristic peaks at 401.0 eV, 399.55 eV and 400.0 eV, which correspond to NC, N-Ca and N=C groups, respectively. Figure 4 The Ca 2p spectrum of (d) shows that Ca 2p 1 / 2 and Ca 2p 3 / 2 The characteristic peaks appeared at 351.0 eV and 347.4 eV, respectively, confirming that Ca... 2+ The successful incorporation. Figure 4 In the O 1s spectrum of (e), the two fitted peaks are located at 532.7 eV and 531.5 eV, corresponding to the C=O and CO bonds, respectively. These XPS results collectively confirm the successful integration of calcium and its coordination with heteroatom sites in BQ-Ca-COF.

[0068] Example 3: Performance Testing

[0069] (1) Photothermal performance test

[0070] By varying the concentration of BQ-Ca-COF (25, 50, 75, 100, 125 μg / mL) or the laser power density (0.5, 1.0, 1.2, and 1.5 W / cm²), 2 The photothermal conversion performance of BQ-Ca-COF was studied in detail. The preparation method for different concentrations of BQ-Ca-COF was as follows: First, 1 mg of BQ-Ca-COF was weighed and thoroughly dispersed in 1 mL of distilled water using an ultrasonic apparatus to prepare a 1 mg / mL stock solution. Then, 25, 50, 75, 100, and 125 μL of the stock solution were respectively added to 975, 950, 925, 900, and 875 μL of distilled water to prepare BQ-Ca-COF aqueous dispersions of 25, 50, 75, 100, and 125 μg / mL. Pure water was used as a control, and a 638 nm laser (1.2 W / cm²) was used. 2 The temperature changes of different concentrations of BQ-Ca-COF were monitored (10 min) to preliminarily study the photophysical properties of the synthesized samples.

[0071] like Figure 5 As shown in (a), the laser power intensity is positively correlated with the dispersion temperature. Figure 5 (b) shows different products under the same concentration conditions (100 µg / mL). -1 The temperature versus time graph, comparing the photothermal effects of BQ-COF and BQ-Ca-COF, illustrates the effect of Ca... 2+ It can interact with phospholipids in bacterial membranes, disrupting their structural integrity and enhancing permeability, thus synergistically enhancing the effectiveness of subsequent photothermal effects. A comparison of the photothermal effects of six materials revealed that, under the same irradiation conditions, BQ-Ca-COF consistently maintained a higher temperature at every time point than all other samples, confirming its superior photothermal performance. Furthermore, Figure 5 Unlike the case in (c) where the temperature change of pure water is negligible, BQ-Ca-COF exhibits a concentration-dependent warming behavior, and the temperature increases rapidly with increasing concentration; for example... Figure 5 (d) shows the temperature rise versus laser power density (0.5-1.5 W / cm²). -2 ) and BQ-Ca-COF concentration (0-125 µg mL) -1 There is a near-linear correlation between them. Figure 5 (e) The excellent photothermal properties of BQ-Ca-COF can also be directly observed from the results obtained by the thermal imaging camera. All these results indicate that red light can be effectively converted into local heat by adjusting the BQ-Ca-COF concentration or the laser power. To evaluate the photostability of BQ-Ca-COF, periodic irradiation experiments were conducted, such as... Figure 5(f) shows that the BQ-Ca-COF has a stable laser switching effect, and the temperature hardly fluctuates after 5 consecutive laser on / off cycles.

[0072] The reusability of synthesized BQ-Ca-COF was further evaluated by comparing its photothermal properties after 0 days and 15 days of dispersion in water. Figure 5 As shown in (g), the heating curve collected after immersion in water for 15 days almost completely overlaps with the initial heating curve, indicating that BQ-Ca-COF has good water stability.

[0073] Furthermore, the photothermal conversion efficiency η (%) of BQ-Ca-COF was calculated to be 58%. The photothermal conversion efficiency calculation formula was obtained by referring to the photothermal conversion efficiency formula in the application No. CN115845086A entitled "A Photothermal-Fenton-like Artificial Nanoenzyme and Its Preparation Method and Application". Figure 5 (h) shows the values ​​of τS and θ in the formula for calculating photothermal conversion efficiency.

[0074] (2) Calcium ion release

[0075] The calcium ion concentration was determined using inductively coupled plasma mass spectrometry (ICP-MS). Specifically, BQ-Ca-COF was prepared into a 1 mg / mL dispersion using PBS buffer and irradiated with a 638 nm laser for 0, 2, 4, 6, and 8 minutes. Samples were taken at preset time points and the filtrate was collected through a 0.22 μm filter membrane. After treatment with nitric acid solution, the calcium ion concentration was determined using ICP-MS. Figure 5 The results in (i) show a clear time-dependent sustained-release pattern, with the entire release process being gradual and without any sudden effects. The initial time point Ca... 2+ The concentration was 188.41 μg / mL, which was attributed to the rapid dissolution of trace amounts of free calcium ions on the material surface. The concentration steadily increased with laser time, reaching 429.03 μg / mL at minute 6 and 523.48 μg / mL at minute 8.

[0076] Test Example 1: In vitro antibacterial test

[0077] (1) Bacterial culture

[0078] This experiment used two bacteria, *Staphylococcus aureus* and *Escherichia coli*, and utilized second-generation bacteria to complete the following experiments. The specific culture method for the second-generation bacteria was as follows: First, the frozen bacteria were thawed at 37°C. 100 µL of the bacterial culture was transferred to a shaker tube containing 5 mL of liquid culture medium and incubated on a shaker (110 rpm, 37°C) for 12 hours. Then, 100 µL of the cultured bacterial culture was transferred to a 2 mL EP tube containing 900 µL of the culture medium, and further diluted using a serial dilution method at 10⁻⁶ increments. -2 Dilute 5-10 tubes, take 100 µL of bacterial suspension from each tube, and spread it evenly on a petri dish containing solid culture medium using a spreader. Incubate at 37°C for 24 hours, observe clonal morphology and colony count. The petri dish with approximately 1000 colonies is considered the first generation of bacteria. Use a pick to pick one colony from the first generation and add it to a shaker tube containing 5 mL of liquid culture medium. Culture the second generation of bacteria using the same method. The specific preparation method for the liquid culture medium is as follows: Disperse 5g of LB broth in 200 mL of distilled water, and then autoclave to obtain the bacterial liquid culture medium. The specific preparation method for the solid culture medium is as follows: Disperse 5g of LB broth and 3g of agar in 200 mL of distilled water, and then autoclave to obtain the solid culture medium.

[0079] (2) Determination of the antibacterial activity of BQ-Ca-COF by plate count method

[0080] The BQ-COF, BQ-Ca-COF, PT-Ca-COF, and CE-Ca-COF prepared in Examples 1 and Comparative Examples 1-2 were prepared into solutions: 2 mg of BQ-COF, BQ-Ca-COF, PT-Ca-COF, or CE-Ca-COF powder was fully dispersed in 2 mL of PBS to prepare a 1 mg / mL BQ-COF or BQ-Ca-COF stock solution. 10 µL of each solution was added to six 2 mL EP tubes. 8 CFU mL -1 Bacterial solutions (S. aureus or E. coli) were prepared by adding 1000, 975, 950, 925, 900, and 875 μL of PBS, respectively, followed by adding 0, 25, 50, 75, 100, and 125 μL of 1 mg / mL BQ-COF, BQ-Ca-COF, PT-Ca-COF, or CE-Ca-COF stock solutions, respectively, to obtain BQ-COF, BQ-Ca-COF, PT-Ca-COF, or CE-Ca-COF solutions with concentrations of 0, 25, 50, 75, 100, and 125 μg / mL.

[0081] The experiment was divided into six batches:

[0082] The first batch consisted of BQ-COF solutions with concentrations of 0, 25, 50, 75, 100, and 125 μg / mL. Each concentration of BQ-COF solution was incubated for 12 hours in a constant-temperature shaker (110 rpm, 37℃). The resulting bacterial cultures were then serially diluted 10⁻⁶ times using the same methods as bacterial cultures. 5 The bacterial culture was doubled, and 100 μL of the well-spread bacterial solution was transferred to a solid culture medium, spread evenly, and incubated at 37°C for 24 h. Colonies were counted and bacterial activity was compared between groups. Figure 7 (a) ~ Figure 7 As shown in (b), the antibacterial activity of BQ-COF solution against Staphylococcus aureus and Escherichia coli at 125 μg / mL is almost negligible.

[0083] The second batch consisted of BQ-COF solutions with concentrations of 0, 25, 50, 75, 100, and 125 μg / mL. Different concentrations of BQ-COF solutions were irradiated with a laser (laser parameters: λ = 638 nm, 1.2 W / cm²). 2 Different concentrations of BQ-COF solution were incubated in a constant temperature shaker (110 rpm, 37℃) for 12 h, and then the cultured bacterial solutions were serially diluted 10 times according to the bacterial culture method. 5 The bacterial culture was doubled, and 100 μL of the well-spread bacterial solution was transferred to a solid culture medium, spread evenly, and incubated at 37°C for 24 h. Colonies were counted and bacterial activity was compared between groups. Figure 7 (c)~ Figure 7 As shown in (d), the antibacterial rates of the BQ-COF+ laser group at 100 μg / mL can reach 86.80 ±1.96% and 87.44 ±1.86%, respectively.

[0084] The third batch consisted of BQ-Ca-COF solutions with concentrations of 0, 25, 50, 75, 100, and 125 μg / mL. The different concentrations of BQ-Ca-COF solutions were incubated for 12 hours on a constant-temperature shaker (110 rpm, 37℃). The resulting bacterial cultures were then serially diluted 10⁻⁶ times according to bacterial culture methods. 5 The bacterial culture was doubled, and 100 μL of the well-spread bacterial solution was transferred to a solid culture medium, spread evenly, and incubated at 37°C for 24 h. Colonies were counted and bacterial activity was compared between groups. Figure 7 (e)~ Figure 7 As shown in (f), the BQ-Ca-COF group showed weak antibacterial effect and almost no antibacterial activity at 125 μg / mL.

[0085] The fourth batch consisted of BQ-Ca-COF solutions with concentrations of 0, 25, 50, 75, 100, and 125 μg / mL. Different concentrations of BQ-Ca-COF solutions were irradiated with a laser (laser parameters: λ = 638 nm, 1.2 W / cm²). 2 Different concentrations of BQ-Ca-COF solutions were incubated in a constant temperature shaker (110 rpm, 37℃) for 12 h, and then the cultured bacterial solutions were serially diluted 10 times according to the bacterial culture method. 5 The bacterial culture was then transferred to a solid culture medium at a ratio of 1:1. The culture was spread evenly and incubated at 37°C for 24 hours. Colonies were counted and bacterial activity was compared among the groups. Figure 7 (g)~ Figure 7 As shown in (h), the BQ-Ca-COF + laser group achieved a significant antibacterial effect at a concentration of 100 μg / mL. The BQ-Ca-COF + laser group achieved a significant antibacterial effect at a concentration of 100 μg / mL, with antibacterial rates of 99.91 ± 0.09% and 99.97 ± 0.06%.

[0086] The fifth batch consisted of PT-Ca-COF solutions with concentrations of 0, 50, 100, 200, 300, and 400 μg / mL. These solutions were irradiated with a laser (laser parameters: λ = 638 nm, 1.2 W / cm²). 2 Different concentrations of PT-Ca-COF solutions were incubated in a constant temperature shaker (110 rpm, 37℃) for 12 h, and then the cultured bacterial solutions were serially diluted 10 times according to the bacterial culture method. 5 The bacterial culture was then transferred to a solid culture medium at a ratio of 1:1. The culture was spread evenly and incubated at 37°C for 24 hours. Colonies were counted and bacterial activity was compared among the groups. Figure 7 (i)~ Figure 7 As shown in (j), the PT-Ca-COF + laser combination can achieve a certain antibacterial effect at 400 μg / mL, with antibacterial rates of 88.74 ± 2.17% and 90.52 ± 0.97%, respectively.

[0087] The sixth batch consisted of CE-Ca-COF solutions with concentrations of 0, 50, 100, 200, 300, and 400 μg / mL. These solutions were irradiated with a laser (laser parameters: λ = 638 nm, 1.2 W / cm²). 2 Different concentrations of CE-Ca-COF solutions were incubated in a constant temperature shaker (110 rpm, 37℃) for 12 h, and then the cultured bacterial solutions were serially diluted 10 times according to the bacterial culture method. 5The bacterial culture was then transferred to a solid culture medium at a ratio of 1:1. The culture was spread evenly and incubated at 37°C for 24 hours. Colonies were counted and bacterial activity was compared among the groups. Figure 7 (k)~ Figure 7 As shown in (l), the CE-Ca-COF + laser group still had no antibacterial effect at 400 μg / mL, indicating that the material basically does not have antibacterial ability.

[0088] like Figure 7 As shown, in the dose-response study, the pure materials without laser treatment showed almost no antibacterial effect, while the materials + laser treatment groups all exhibited concentration-dependent antibacterial effects. However, at equivalent concentrations, the calcium-containing platform consistently achieved superior bactericidal effects. Comparing the antibacterial effects of the three calcium-containing materials under laser treatment, it can be seen that BQ-Ca-COF has a better antibacterial effect than PT-Ca-COF and CE-Ca-COF. The above experiments demonstrate that with increasing BQ-Ca-COF solution concentration, its bactericidal performance against Staphylococcus aureus and Escherichia coli is significantly enhanced. Through comparison, it can be found that under laser treatment, BQ-Ca-COF exhibits superior antibacterial performance compared to other materials.

[0089] (3) In vitro antibacterial activity of BQ-Ca-COF under different treatments

[0090] The antibacterial activity under different treatments was studied using the plate count method, with bacterial culture as in (1). The experiment was divided into six groups: (I) control group (PBS), (II) BQ-COF, (III) BQ-Ca-COF, (IV) laser group, (V) BQ-COF + laser, and (VI) BQ-Ca-COF + laser. All of the above groups were treated with bacterial suspension (10 µL 10 8 CFU mL -1 Groups containing bacterial solutions (S. aureus or E. coli). In groups containing BQ-Ca-COF or BQ-COF, the concentration of either BQ-Ca-COF or BQ-COF was 100 μg / mL. The preparation method was as follows: add 10 µL of BQ-Ca-COF to each 2 mL EP tube. 8 CFU mL -1 Bacterial solutions (S. aureus or E. coli) were mixed with 900 μL of PBS and 100 μL of 1 mg / mL BQ-Ca-COF or BQ-COF to obtain a BQ-Ca-COF dispersion or BQ-COF dispersion with a concentration of 100 μg / mL. In the group containing laser, the laser parameters were: λ = 638 nm, 1.2 W / cm². 2(IV) The laser group consisted of PBS and laser. Then, each group was placed in a constant temperature shaker (110 rpm, 37°C) and incubated for 12 h. The cultured bacterial solutions were then serially diluted 10⁻⁶ times according to bacterial culture methods. 5 The bacterial culture was then transferred to a solid culture medium at a ratio of 1:1. The culture was spread evenly and incubated at 37°C for 24 hours. Colonies were counted and bacterial activity was compared with that of each group.

[0091] like Figure 6 As shown, when treated with BQ-COF or BQ-Ca-COF alone, the bacterial survival rate exceeded 80%, indicating limited inherent antibacterial activity. In stark contrast, under laser irradiation, BQ-Ca-COF exhibited a significantly enhanced bactericidal effect, with survival rates for both strains approaching 0%. Specifically, the survival rate of Staphylococcus aureus plummeted to 0.18% ± 0.20%, and the survival rate of Escherichia coli decreased to 0.22% ± 0.13%, far lower than the levels in the corresponding unirradiated groups (treated with BQ-Ca-COF alone) (87.51% ± 3.53% for Staphylococcus aureus and 79.41% ± 11.79% for Escherichia coli).

[0092] (4) To study the antibacterial properties of BQ-Ca-COF, an anti-biofilm formation experiment was conducted. The analysis was performed using a confocal laser scanning microscope (CLSM), with the same groupings as in experiment (3). After 48 hours of biofilm growth, the samples were washed, stained with propidium iodide (PI), and observed using CLSM. Figure 8 As shown, consistent with the in vitro antibacterial results, confocal laser scanning microscopy images revealed differences in the degree of structural damage among the treatment groups, with the BQ-Ca-COF+ laser group exhibiting the most severe damage. Quantitative analysis showed that the biofilm disruption rate (biofilm disruption rate = 100% - biofilm rate) of Staphylococcus aureus in the BQ-Ca-COF+ laser group was 97.84±0.28%, compared to 1.27±2.19% in the BQ-COF group, 54.48±2.14% in the BQ-COF+ laser group, and 3.62±3.86% in the BQ-Ca-COF group. The biofilm disruption rate in the BQ-Ca-COF+ laser group was significantly higher than that in the other groups. Similarly, the biofilm disruption rate of Escherichia coli in the BQ-Ca-COF+ laser group was 96.71±0.60%.

[0093] Simultaneously, crystal violet (CV) staining was used to quantify the biofilm inhibition effect, with phosphate-buffered saline (PBS) as a negative control. The experiment was divided into five groups: control group (PBS), BQ-COF, BQ-Ca-COF, BQ-COF + laser, and BQ-Ca-COF + laser group. Figure 9As shown, when PBS is used alone, its anti-biofilm activity is negligible. Figure 9 As shown, compared to BQ-COF, the BQ-Ca-COF group (with laser treatment) and the untreated BQ-Ca-COF group exhibited a more significant biofilm disruption effect. This comparison demonstrates that Ca... 2+ The presence of [a substance] can disrupt the integrity of biofilms and enhance permeability. The BQ-Ca-COF+laser group showed particularly outstanding performance, with an inhibition rate of 96.27±1.07% for Staphylococcus aureus biofilm (inhibition rate = 100% - biofilm rate) and an inhibition rate of 91.29±1.04% for Escherichia coli biofilm, significantly better than all other treatment groups.

[0094] Experimental Example 2: Transmission Electron Microscopy of Bacteria

[0095] Bacterial suspensions were prepared according to the method in Experimental Example 1 (3): (I) control group, (II) BQ-COF, (III) BQ-Ca-COF, (IV) laser, (V) BQ-COF + laser, and (VI) BQ-Ca-COF + laser. Then, 100 μL of the bacterial suspension was fixed in 2.5 wt% glutaraldehyde solution (4°C, 2 h), washed three times with PBS, embedded in agar, and blocked. The bacteria were then dehydrated by continuous treatment with ethanol solutions (30 wt%, 50 wt%, 70 wt%, 90 wt%, 95 wt%, and 100 wt%) at room temperature for 10 min, followed by treatment with acetone at room temperature for 3 h, and then embedded in a gradient infiltration medium (epoxy resin) (impregnated with acetone and epoxy resin at mass ratios of 3:1, 1:1, and 1:3 for 1 hour each, and finally impregnated with pure epoxy resin overnight). Negative staining was performed, and sections were prepared on nickel grids. A nickel grid was placed under a TEM to capture bacterial morphology. The integrity of the bacterial membranes from different treatment groups was then observed using the TEM.

[0096] like Figure 10 As shown, Staphylococcus aureus and Escherichia coli in the control group, BQ-COF group, and BQ-Ca-COF alone maintained intact cell membranes and typical morphology. In contrast, bacteria treated with BQ-COF + laser or BQ-Ca-COF + laser showed severe membrane damage, content leakage, and structural distortion. Most notably, the BQ-Ca-COF + laser group resulted in the most severe morphological disruption, including complete membrane rupture, confirming the high bactericidal efficacy observed in the viability assay.

[0097] Experimental Example 3: Bacterial Viability / Deadness Staining Test

[0098] SYTO-9 and PI are used to distinguish between live and dead microbial cells. SYTO-9 can penetrate all bacterial membranes (intact and damaged), thus marking the bacteria as green; PI only penetrates damaged bacterial membranes, marking the bacteria as red, while reducing the green color of SYTO-9.

[0099] Bacterial suspensions were prepared according to the method in Experimental Example 1 (3): (I) control group, (II) BQ-COF, (III) BQ-Ca-COF, (IV) laser, (V) BQ-COF + laser, and (VI) BQ-Ca-COF + laser group. Then, 100 µL of bacterial suspension from each group was mixed with 20 µL of SYTO-9 (1.0 × 10⁻⁶). -3 M) and 20 µL PI (1.5×10 -3 M) Incubate in the dark at 37°C for 15 min. After staining, centrifuge each group in PBS to remove excess SYTO-9 and PI. Then resuspend the bacteria in 50 µL PBS and place them on a glass slide. Images of Escherichia coli or Staphylococcus aureus are then captured using a fluorescence inverted microscope.

[0100] from Figure 11 The significant PTT synergistic antibacterial effect of BQ-Ca-COF can also be directly observed in the results of live and dead bacterial staining. (I) Control group, (II) BQ-COF, (III) BQ-Ca-COF treated bacteria showed strong green fluorescence, consistent with the results of the plate counting method. In contrast, (V) BQ-COF + laser, (VI) BQ-Ca-COF + laser treated bacteria showed obvious red fluorescence. The BQ-Ca-COF + laser group showed almost all bacteria dead, exhibiting the highest bactericidal efficiency. All bacteria were stained with red fluorescence, indicating a large number of dead bacteria.

[0101] Experimental Example 4: In vitro biocompatibility experiment

[0102] (1) Hemolysis test

[0103] Fresh blood was collected from 5-week-old female BALB / c mice (purchased from Jinan Pengyue Experimental Animal Breeding Co., Ltd.). After centrifugation at 10,000 rpm for 10 minutes, red blood cells were collected and washed with the same volume of PBS until colorless, then the supernatant was discarded. Red blood cells were diluted with PBS at a volume ratio of 3:11, and different concentration gradients (50, 100, 150, 200 μg / mL) of BQ-Ca-COF or BQ-COF solution (red blood cell solution: BQ-Ca-COF solution volume ratio = 1:9) were added. The mixture was incubated at 37℃ for 3 hours and centrifuged at 10,000 rpm for 10 minutes. Then, 100 µL of supernatant from each group was placed in a 96-well plate, and the absorbance of each group was measured at 570 nm using an enzyme-labeled immunosorbent assay (ELISA). Distilled water was used as a positive control, and PBS as a negative control. The formula for calculating hemolysis volume is as follows:

[0104] Hemolysis volume (%) = (A-An) / (Ap-An) × 100%;

[0105] Where A is the absorbance obtained by taking the supernatant after adding BQ-Ca-COF to red blood cells;

[0106] An is the absorbance obtained by taking the supernatant after adding PBS to red blood cells (negative control).

[0107] Ap is the absorbance obtained by adding distilled water to red blood cells and taking the supernatant (positive control).

[0108] like Figure 12 As shown in (a) and 12(b), the hemolysis rate of both BQ-COF and BQ-Ca-COF remained below 3% as the concentration increased from 50 to 200 μg / mL. This indicates that BQ-Ca-COF and BQ-COF have good blood compatibility and do not damage the erythrocyte membrane.

[0109] (2) Cytotoxicity test

[0110] In 96-well plates, mouse 3T3 fibroblasts (from the Cell Bank of the Chinese Academy of Sciences) were cultured at a density of 5 × 10⁶ cells per well. 3 Seed cells at a density of 180 µL per well, with 200 µL of PBS added to surrounding replicate wells for liquid sealing to prevent excessive evaporation. After 24 h of incubation, 20 µL of BQ-Ca-COF or BQ-COF at different concentrations (25-125 μg / mL) were added and incubated for 72 h. Then, 20 µL of MTT (4 mg / mL) solution was added to each well, and the cells were incubated for 4 h. The supernatant was then aspirated, and 150 µL of dimethyl sulfoxide was added to dissolve the MTT (tetramethylazazole blue). After dissolving on a shaker for 10 min, the absorbance of the 96-well plate was measured at 570 nm using a microplate reader. Each experiment was repeated three times. Figure 12As shown in (c), after 72 h of culture, even at high concentrations (125 μg / mL), the survival rate of 3T3 cells treated with BQ-Ca-COF and BQ-COF was above 80%.

[0111] (3) Cell scratch test

[0112] Mouse 3T3 fibroblasts (from the Cell Bank of the Chinese Academy of Sciences) were seeded in 6-well plates and cultured for 24 hours. Cells were then scraped from each well using the tip of a sterile pipette and washed twice with PBS. 1 mL of mitomycin C (purchased from Shanghai Aladdin Biochemical Technology Co., Ltd.) was added to each well, and the cells were cultured for 1 hour. Cells were then treated with mitomycin C using BQ-Ca-COF, BQ-COF, and PBS (blank group), respectively. Cell healing was observed under a microscope at 0, 12, 24, and 36 hours. The distance of cell migration was measured using ImageJ software, and the percentage of wound healing was calculated. Figure 12 As shown in (d), the scratch closure rate of the BQ-Ca-COF group was significantly faster than that of the PBS group. Figure 12 (e) and Figure 12 Quantitative analysis in (f) showed that, based on scratch area measurement, the wound healing rate of the BQ-Ca-COF group reached 22.48±0.115% after 36 hours, significantly higher than that of the PBS group (19.48±0.13%) and the BQ-COF group (20.04±0.04%). These results clearly demonstrate that BQ-Ca-COF can effectively promote cell migration and enhance wound healing.

[0113] Experimental Example 5:

[0114] (1) In vivo wound healing experiment

[0115] Five-week-old BALB / c mice (purchased from Jinan Pengyue Experimental Animal Breeding Co., Ltd.) were randomly divided into six groups: blank control group, control group (I), BQ-COF group (II), BQ-Ca-COF group (III), BQ-COF + laser group (IV), and BQ-Ca-COF + laser group (V), with six mice in each group. Except for the blank control group, which received no treatment, the hind hair of each mouse in all other groups was shaved before surgery to create a 5 mm wound, which was then infected with Staphylococcus aureus (1×10⁻⁶). 6 A wound healing model was established by administering CFU / mL for 24 hours. Subsequently, animals were housed individually and treated according to designated groups (administration concentration 100 μg / mL, dose 20 μL). Wound appearance was recorded on days 1, 3, 5, 7, and 9, while body weight was monitored. Changes in wound size were measured using an image analysis program (Image.J).

[0116] The wound condition was recorded on days 1, 3, 5, 7, and 9 post-treatment. Figure 13 As shown in (a), the wound area gradually decreased over time in all groups. However, by day 9, significant differences in healing outcomes were observed between the groups. Quantitative analysis Figure 13 (c) shows that the BQ-Ca-COF + laser group, which combines this material with PTT, achieved the most significant healing effect, with a wound closure rate exceeding 90%. The wound closure rate was 67.73% in the control group, 70.63% in the BQ-COF group, 74.51% in the BQ-Ca-COF group, and 79.84% in the BQ-COF + laser group; the healing rate of the BQ-Ca-COF + laser group was higher than that of the other groups. Meanwhile... Figure 13 (b) It can be seen that, compared with the body weight of normal mice (blank group), no significant weight change was detected in any group during the entire treatment process, indicating that BQ-Ca-COF has good biocompatibility.

[0117] (2) Tissue staining experiment

[0118] Histological analysis was performed using hematoxylin and eosin (H&E) and Masson staining to directly assess the healing status of skin tissue 9 days after treatment. Figure 14 As shown, the control groups all exhibited significant inflammatory cells, and the epidermis remained incomplete regardless of laser irradiation levels. In contrast, the treatment groups showed varying degrees of skin structure regeneration. The BQ-Ca-COF+ laser group demonstrated the most significant effect in promoting wound healing, with visible blood vessels, collagen fibers, and a naturally matured epidermal layer, indicating complete and excellent wound healing.

[0119] In addition, histological sections were collected, and H&E staining was used to assess the damage to internal organs (heart, liver, spleen, lungs, and kidneys) in different groups. Figure 15 As shown, no abnormal lesions or inflammation were observed in the major organs, and no histological changes were found. These results confirm that BQ-Ca-COF has good biocompatibility in vivo.

[0120] (3) Complete blood count test

[0121] On day 9, approximately 2 mL of blood was collected from the ophthalmic artery of the mice, and each sample underwent routine blood testing. Figure 16 As can be seen, all blood parameters were within the normal range, indicating that BQ-Ca-COF has good biocompatibility.

[0122] The above description is merely a preferred embodiment of this application and is not intended to limit this application. Various modifications and variations can be made to this application by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the protection scope of this application.

Claims

1. An antibacterial material with calcium ion-mediated and photothermal effects, characterized in that, The antibacterial material is a calcium ion-supported porous covalent organic framework polymer; the porous covalent organic framework polymer is obtained by copolymerization of cyclohexanehexanone and aromatic tetraamine as monomers. The aromatic tetraamine is 2,3,5,6-tetraaminocyclohexane-2,5-diene-1,4-dione; The structural formula of the antibacterial material is: 。 2. The method for preparing the antibacterial material according to claim 1, characterized in that, Includes the following steps: (1) 2,3,5,6-tetraaminocyclohexane-2,5-diene-1,4-dione and cyclohexanehexane octahydrate were added to an organic solvent for a solvothermal reaction. After the reaction was completed, the polymer was separated, washed and dried in sequence to obtain a porous covalent organic framework polymer. (2) Under a protective atmosphere, the porous covalent organic framework polymer is dispersed in distilled water, and then an excess of calcium salt is added to obtain a dispersion. The dispersion is stirred to carry out a metallization reaction. After the reaction is completed, the polymer is centrifuged, washed, and dried in sequence to obtain an antibacterial material with calcium ion-mediated and photothermal effects.

3. The preparation method according to claim 2, characterized in that, In step (1), the molar ratio of 2,3,5,6-tetraaminocyclohexane-2,5-diene-1,4-dione to cyclohexanehexane octahydrate is 1.5:1; the organic solvent is a mixture of ethylene glycol and acetic acid solution; and the concentration of the acetic acid solution is 6M.

4. The preparation method according to claim 2, characterized in that, In step (1), the temperature of the solvothermal reaction is 120°C and the time is 72h; the separation is first ultrasonically extracted with distilled water for 24h, and then Soxhlet extracted with acetone for 24h.

5. The preparation method according to claim 2, characterized in that, In step (2), the calcium salt is calcium chloride; the concentration of the porous covalent organic framework polymer in the dispersion is 1 mg / mL and the concentration of the calcium salt is 100 mM.

6. The preparation method according to claim 2, characterized in that, In step (2), the metallization reaction is carried out at a temperature of 25°C for 5 days.

7. The use of the antibacterial material according to claim 1 in the preparation of antibacterial drugs.

8. The application according to claim 7, characterized in that, The antibacterial material releases Ca under infrared laser irradiation. 2+ It also produces a photothermal effect, and the two work together to achieve antibacterial effect.

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