Organic polymer composite material based on BODIPY photo-controlled CO release and its preparation method and application

Abstract

The present invention discloses an organic polymer composite material based on BODIPY light-controlled CO release, as well as its preparation method and application, belonging to the field of biomedicine technology. Tetraaldehyde BDP and 2,2'-bipyridine-5,5'-diamine are dissolved in an organic solvent, ultrasonically dispersed to form a suspension, and after cyclic deoxygenation, heated in a sealed environment for reaction to obtain a powdered black solid BB-COF. BB-COF and pentacarbonyl manganese bromide are suspended in anhydrous ether under a protective atmosphere, and stirred at room temperature for reaction to obtain an organic polymer composite material based on BODIPY light-controlled CO release. The composite material prepared by the present invention has an eggshell-like spherical structure and the ability to release CO under light-controlled CO. It can achieve antibacterial effects through a quadruple synergistic mechanism of photothermal therapy, photodynamic therapy, Fenton reaction, and carbon monoxide (CO) release, making it safer for use in organisms.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine technology, and in particular to an organic polymer composite material based on BODIPY light-controlled CO release, and a preparation method and application thereof. Background Art

[0002] Over the past few decades, numerous antibiotic-free antimicrobial approaches that do not induce resistance have been developed as alternatives to antibiotics, including thermal damage-based photothermal therapy (PTT), reactive oxygen species (ROS)-based photodynamic therapy (PDT), enzyme therapy, and sonodynamic therapy (SDT). However, these technologies face various challenges in practical application. For example, limited oxygen concentration significantly hinders the efficiency of PDT, and PTT faces challenges in therapeutic specificity, all of which inevitably cause damage to normal tissue. To simultaneously address these issues, achieve satisfactory therapeutic effects, and minimize side effects, complex therapeutic strategies that not only exert targeted therapeutic effects but also effectively reshape the pathological wound microenvironment offer unique advantages in the treatment of infectious wounds. However, these complex therapeutics still face significant challenges, including low active site density, poor selectivity, and low specificity. Therefore, the development of intelligent and efficient complex therapeutics remains urgent.

[0003] Phototherapy primarily consists of PDT and PTT, both non-invasive and highly effective. PDT involves laser irradiation in the presence of a photosensitizer (PS) to generate reactive oxygen species (ROS), which react with various bacterial biomolecules (such as lipids in cell membranes, proteins within cells, and nucleic acids). PTT also occurs through laser irradiation in the presence of a photosensitizer (or, in some cases, materials with photothermal conversion properties). However, it primarily converts light energy into heat, raising the local temperature. When the temperature rises to a certain level, it can cause irreversible damage to bacteria, such as denaturing bacterial proteins. The Fenton reaction is an inorganic chemical reaction in which hydrogen peroxide (H2O2) reacts with a metal under acidic conditions (typically pH 2-4) to generate highly reactive hydroxyl radicals (·OH). These hydroxyl radicals can damage bacterial cell structures, providing a potent disinfectant and sterilizer. Some studies have shown that CO gas can change the fluidity and permeability of bacterial cell membranes; it may cause structural changes in bacterial cell membranes by interacting with certain components on the bacterial cell membrane, such as lipids or membrane proteins. Moreover, compared with some chemical disinfectants, the decomposition products of CO gas after sterilization are relatively simple, mainly harmless substances such as carbon dioxide.

[0004] Boron dipyrrolidone (BODIPY) compounds are an important class of fluorescent dyes. Due to their excellent optical properties, they have been reported to be used in photothermal therapy (PTT) and photodynamic therapy (PDT) for tumors and antibacterial applications. However, many challenges remain in their application, such as the inability of a single substance to simultaneously possess multiple functions, such as PTT and PDT, the inability to simultaneously excite multiple photosensitizers with a single wavelength laser, and the lack of a clear understanding of the structure-activity relationship between BODIPY and PTT / PDT. These issues hinder further development of BODIPY in the field of phototherapy. Patent application number CN116785433A discloses a photothermal-photodynamically driven NO-releasing synergistic cationic bactericidal material and its application. The porous polymer BG-POP is obtained by dehydration condensation reaction of a boron dipyrrolidone (Bodipy) compound and triaminoguanidine hydrochloride. The porous polymer is then added to water and mixed with sodium nitroprusside to produce the photothermal-photodynamically driven NO-releasing synergistic cationic bactericidal material BG-SNP. However, there is currently no composite material based on BODIPY's light-controlled release of CO for antibacterial purposes. If it can simultaneously possess the four antibacterial modes of photothermal, photodynamic, CO and Fenton, it is expected to obtain a more efficient and controllable broad-spectrum fungicide. Summary of the Invention

[0005] In response to the above-mentioned prior art, the present invention aims to provide an organic polymer composite material based on BODIPY-based photocontrolled CO release, as well as its preparation method and application. The present invention utilizes tetraaldehyde BDP and 2,2'-bipyridine-5,5'-diamine to polymerize, and utilizes bipyridine's multidentate ligand to load pentacarbonyl manganese bromide. This organic polymer composite material exhibits an eggshell-like spherical structure and photocontrolled CO release. It exhibits antibacterial properties through a four-fold synergistic mechanism involving photothermal therapy, photodynamic therapy, the Fenton reaction, and carbon monoxide (CO) release, resulting in improved safety for biological applications.

[0006] To achieve the above object, the present invention adopts the following technical solutions:

[0007] In a first aspect of the present invention, a method for preparing an organic polymer composite material based on BODIPY light-controlled CO release is provided, comprising the following steps:

[0008] (1) BODIPY undergoes iodination reaction with an iodine-containing compound to obtain iodinated BODIPY;

[0009] (2) BODIPY iodide, 3,5-diformylphenylboronic acid and potassium carbonate are dissolved in an organic solvent, a catalyst is added under a protective atmosphere and the reaction is carried out to obtain tetraaldehyde BDP;

[0010] (3) Tetraaldehyde BDP and polyamino bipyridine are dissolved in an organic solvent, ultrasonically dispersed to form a suspension, and then subjected to cyclic deoxygenation and heated in a sealed environment to react to obtain a powdery black solid BB-COF;

[0011] (4) Under a protective atmosphere, BB-COF and pentacarbonyl manganese bromide were suspended in anhydrous ether and stirred at room temperature to obtain an organic polymer composite material based on BODIPY photocontrolled CO release.

[0012] Preferably, in step (1), the iodine-containing compound is N-iodosuccinimide; the molar ratio of BODIPY to N-iodosuccinimide is 1:2; and the iodination reaction is performed by dissolving BODIPY and N-iodosuccinimide in anhydrous dichloromethane and stirring the mixture at room temperature.

[0013] Preferably, in step (2), the molar ratio of the iodinated BODIPY to 3,5-diformylphenylboronic acid is 1:2; the organic solvent is 1,4-dioxane; and the catalyst is tetrakis(triphenylphosphine)palladium.

[0014] Preferably, in step (2), the reaction temperature is 120° C. and the reaction time is 12 h.

[0015] Preferably, in step (3), the polyamino bipyridine is 2,2'-bipyridine-5,5'-diamine; the organic solvent is a mixture of toluene and acetic acid solution in a volume ratio of 20:1; the concentration of the acetic acid solution is 3M; the temperature of the heating reaction is 120°C, and the reaction time is 72h.

[0016] Preferably, the molar ratio of the tetraaldehyde BDP to 2,2'-bipyridine-5,5'-diamine is 1:2.

[0017] Preferably, in step (4), the molar ratio of BB-COF to manganese pentacarbonyl bromide (Mn(CO)5Br) is 1:4; and the reaction time at room temperature is 4 hours.

[0018] In a second aspect, the present invention provides an organic polymer composite material based on BODIPY photo-controlled CO release obtained by the above-mentioned preparation method. The organic polymer composite material based on BODIPY photo-controlled CO release is obtained by polymerizing polyaldehyde BDP and polyamino bipyridine as monomers, and using bipyridine's multidentate ligand to load pentacarbonyl manganese bromide; the organic polymer composite material based on BODIPY photo-controlled CO release has an egg-shell spherical structure and releases CO under laser irradiation.

[0019] Preferably, the polyaldehyde BDP is tetraaldehyde BDP; and the polyamino bipyridine is 2,2'-bipyridine-5,5'-diamine.

[0020] The third aspect of the present invention provides the use of an organic polymer composite material based on BODIPY light-controlled CO release in the preparation of antibacterial drugs.

[0021] Organic polymer composites based on BODIPY photo-controlled CO release have synergistic antibacterial effects through four therapies: photothermal therapy, photodynamic therapy, Fenton reaction, and carbon monoxide (CO) release.

[0022] Beneficial effects of the present invention:

[0023] (1) The organic polymer composite material based on BODIPY photo-controlled CO release (hereinafter referred to as BB-COF@Mn) prepared in the present invention can induce good photothermal conversion by 638 nm wavelength laser irradiation, and can also generate singlet oxygen, hydroxyl radicals and superoxide anions through the electron transfer induced by it. In addition, pentacarbonyl manganese bromide loaded on the material can release carbon monoxide by photo-control, so that BB-COF@Mn has a quadruple synergistic mechanism of photothermal therapy, photodynamic therapy, Fenton reaction and carbon monoxide (CO) release, realizing combined broad-spectrum antibacterial effect.

[0024] (2) The preparation method of the present invention is simple, and the prepared BB-COF@Mn has good biocompatibility, stable structure and performance. The hemolysis rate of red blood cells is less than 1.5%, the effect on the viability of HEK293 cells and L929 cells is minimal, and it has no toxic side effects on the human body. This will promote the development of multifunctional antibacterial platforms. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 : Characterization analysis diagram of BB-COF@Mn; (a) is the infrared spectrum of BB-COF@Mn; (b) is the solid carbon spectrum of BB-COF@Mn 13 C NMR; (c) is the pore size distribution curve of BB-COF@Mn; (d) is the low-temperature N2 absorption isotherm of BB-COF@Mn at 77 K; (e) is the thermogravimetric curve of BB-COF@Mn; (f) is the particle size distribution diagram of BB-COF@Mn;

[0026] Figure 2 : SEM and TEM images of BB-COF@Mn; (a) is the SEM of BB-COF at a scale of 5µm; (b) is the SEM of BB-COF at a scale of 2µm; (c) is the SEM of BB-COF@Mn at a scale of 5µm; (d) is the SEM of BB-COF@Mn at a scale of 2µm; (e) is the TEM of BB-COF at a scale of 1µm; (f) is the TEM of BB-COF at a scale of 500nm; (g) is the TEM of BB-COF at a scale of 50nm; (h) is the TEM of BB-COF at a scale of 10nm; (i) is the TEM of BB-COF@Mn at a scale of 1µm; (j) is the TEM of BB-COF at a scale of 500nm @Mn; (k) is the TEM of BB-COF@Mn at a scale of 50 nm; (l) is the TEM of BB-COF@Mn at a scale of 10 nm; (m) is the mapping diagram of BB-COF@Mn; (n) is the distribution of the B element in BB-COF@Mn; (o) is the distribution of the C element in BB-COF@Mn; (p) is the distribution of the N element in BB-COF@Mn; (q) is the distribution of the O element in BB-COF@Mn; (r) is the distribution of the F element in BB-COF@Mn; (s) is the distribution of the Mn element in BB-COF@Mn; (t) is the distribution of the Br element in BB-COF@Mn;

[0027] Figure 3 : EDS pattern of BB-COF@Mn;

[0028] Figure 4 : Photothermal performance of BB-COF@Mn; (a) is at 1.5W / cm2 Concentration-dependent photothermal effect of BB-COF@Mn under 638 nm laser irradiation; (b) is the concentration-dependent photothermal effect of BB-COF@Mn under 0.3, 0.7, 1.0, 1.3 and 1.5 W / cm 2 (c) Thermal imaging of the heating process of BB-COF@Mn with different concentrations within 15 minutes.

[0029] Figure 5 :Photothermal stability of BB-COF@Mn; (a) is 638nm laser at 1.5W / cm 2 Temperature variation curve of BB-COF@Mn (200 μg / mL) after three times of photo-cooling; (b) is the temperature variation curve of BB-COF@Mn (200 μg / mL) after three times of photo-cooling; (c) is the temperature variation curve of BB-COF@Mn (200 μg / mL) after three times of photo-cooling; (d) is the temperature variation curve of BB-COF@Mn (200 μg / mL) after three times of photo-cooling; (e) is the temperature variation curve of BB-COF@Mn (200 μg / mL) after three times of photo-cooling; (f ... 2 ) switch, the temperature change curve of BB-COF@Mn (200 μg / mL) aqueous dispersion during a continuous heating-cooling process; (c) is a curve showing the negative natural logarithm of the ratio of the difference between the corresponding temperature and the ambient temperature and the difference between the highest temperature and the ambient temperature at each moment in the cooling cycle of BB-COF@Mn;

[0030] Figure 6 : Electron paramagnetic resonance spectrum of BB-COF@Mn;

[0031] Figure 7 : Photodynamic performance of BB-COF@Mn, where (a) is the photodynamic performance of DPBF under red light (638 nm, 1.5 W / cm 2 ) UV-visible spectrum under red light (638 nm, 1.5 W / cm 2 ) UV-visible spectrum under irradiation;

[0032] Figure 8 : Photocontrolled CO release ability and Fenton reaction of BB-COF@Mn; (a) is the UV-visible spectra of Deoxy-Mb and BB-COF@Mn; (b) is the UV-visible spectra of Deoxy-Mb and BB-COF@Mn under 638 nm laser irradiation; (c) is the UV-visible spectra of four groups (A: TMB, B: TMB+H2O2, C: TMB+ H2O2+BB-COF, D: TMB+ H2O2+BB-COF@Mn); (d) is the UV-visible spectra of different concentrations of BB-COF@Mn (50, 100, 125, 150 and 200 μg / mL) + TMB+H2O2, experimental conditions: 1.5 mmol / L TMB, 20 mmol / L hydrogen peroxide, triggered in a water environment with pH = 4.5;

[0033] Figure 9 : Antibacterial activity of BB-COF@Mn at different concentrations; (a) is the culture image of Staphylococcus aureus and Escherichia coli after treatment with different concentrations of BB-COF@Mn; (b) is the bacterial survival rate statistics of Staphylococcus aureus and Escherichia coli determined by the plate count method;

[0034] Figure 10 : Antibacterial ability under different treatment methods; (a) is the culture diagram of Staphylococcus aureus and Escherichia coli after treatment with different treatment methods; (b) is the bacterial survival rate statistics of Staphylococcus aureus and Escherichia coli determined by plate count method;

[0035] Figure 11 : Transmission electron microscopy images of Staphylococcus aureus and Escherichia coli under different treatments, scale bar is 2.0 μm;

[0036] Figure 12 : Biocompatibility experiment of BB-COF@Mn; (a) is the hemolysis rate of BB-COF@Mn at different concentrations; (b) is the cell viability after incubation of L929 cells or HEK293 cells with different concentrations of BB-COF@Mn. DETAILED DESCRIPTION

[0037] It should be noted that the following detailed descriptions are illustrative and intended to provide further explanation of the present application. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the present application belongs.

[0038] As mentioned in the background section, there are reports of BODIPY being used for photothermal therapy (PTT) and photodynamic therapy (PDT) for tumors and antibacterial applications. However, numerous challenges remain in its application, including the inability of a single material to simultaneously possess multiple functions, such as PTT and PDT, the inability to simultaneously excite multiple photosensitizers with a single wavelength laser, and the lack of a clear understanding of the structure-activity relationship between BODIPY and PTT / PDT. These challenges hinder the further development of BODIPY in the field of phototherapy. The development of a multimodal, synergistic antibacterial material based on BODIPY is needed.

[0039] Based on this, the present invention aims to provide an organic polymer composite material based on BODIPY's photocontrolled CO release, as well as its preparation method and application. The present invention first prepares polyaldehyde BODIPY (tetraaldehyde BDP) through iodination and Suzuki coupling. The tetraaldehyde BDP is then polymerized with polyamino bipyridine (2,2'-bipyridine-5,5'-diamine) to produce BB-COF. BB-COF exhibits only two antibacterial modes: photothermal and photodynamic. Therefore, the multidentate bipyridine ligand on BB-COF is then combined with Mn(CO)5Br to produce BB-COF@Mn. The Mn(CO)5Br in BB-COF@Mn releases CO under infrared light, and Mn exhibits a Fenton effect, resulting in BB-COF@Mn exhibiting four antibacterial modes: photothermal therapy, photodynamic therapy, Fenton reaction, and carbon monoxide (CO) release. In addition, the BB-COF@Mn prepared in the present invention has an egg-shell spherical structure, which is conducive to drug loading and can provide an excellent carrier for the future development of PTT / PDT anti-tumor drugs.

[0040] In order to enable those skilled in the art to more clearly understand the technical solution of the present application, the technical solution of the present application will be described in detail below with reference to specific embodiments.

[0041] The test materials used in the examples of the present invention are all conventional test materials in the art and can be purchased through commercial channels.

[0042] Example 1 Preparation of BB-COF@Mn

[0043] (1) Preparation of BODIPY: Under nitrogen protection, freshly distilled benzaldehyde (2.1 mL, 20 mmol) and 2,4-dimethylpyrrole (5 mL, 48 mmol) were dissolved in dry dichloromethane (80 mL), and trifluoroacetic acid (TFA, 3 drops) was added dropwise. Subsequently, 2,3-dichloro-5,6-dicyano-p-benzoquinone (DDQ, 5.54 g, 20 mmol) was added. After stirring at room temperature for 10 minutes, triethylamine (20 mL) and boron trifluoride-diethyl ether complex (BF3·OEt2, 20 mL) were slowly added in sequence and stirring was continued for 1 hour. The reaction solution was quenched with water and extracted three times with dichloromethane. The organic phases were combined and washed with saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified by column chromatography (silica gel, volume ratio of petroleum ether to ethyl acetate = 10:1) to obtain a dark red solid BODIPY.

[0044] .

[0045] (2) Preparation of iodinated BODIPY (Diiodo-BDP): BODIPY (200 mg, 0.31 mmol) and excess N-iodosuccinimide (140 mg, 0.62 mmol) were dissolved in anhydrous dichloromethane (15 mL) and stirred at room temperature for 30 minutes. After the reaction, the reaction solution was extracted three times with dichloromethane, and the organic phases were combined. After concentration under reduced pressure, the solution was purified by column chromatography (silica gel, volume ratio of dichloromethane to petroleum ether = 2:1) to obtain a dark red solid Diiodo-BDP. The synthetic route is:

[0046] .

[0047] (3) Preparation of tetraaldehyde BDP: Diiodo-BDP (85.0 mg, 0.148 mmol), 3,5-diformylphenylboronic acid (52.7 mg, 0.296 mmol) and potassium carbonate (40.9 mg, 0.296 mmol) were dissolved in anhydrous 1,4-dioxane (15 mL). Tetrakis(triphenylphosphine)palladium (12.0 mg, 0.296 mmol) was added under argon protection. After stirring at 120°C for 12 hours, the mixture was extracted with dichloromethane, concentrated under reduced pressure, and purified by column chromatography (silica gel, volume ratio of dichloromethane to ethyl acetate = 10:1) to obtain red solid tetraaldehyde BDP. The synthetic route is:

[0048] .

[0049] (4) Preparation of BB-COF: Tetraaldehyde BDP (29.4 mg, 0.05 mmol) and 2,2'-bipyridine-5,5'-diamine (18.6 mg, 0.1 mmol) were dissolved in toluene (4 mL) and 3 M acetic acid solution (0.2 mL), and dispersed by ultrasonication (power 100w) for 10 minutes to form a uniform suspension. After three freeze-evacuation-thaw cycles of deoxygenation, the mixture was sealed in a pressure-resistant glass tube and reacted at 120°C for 72 hours to produce a brown precipitate. The product was washed three times with deionized water, methanol, ethanol, tetrahydrofuran, acetone and dichloromethane, respectively, and vacuum dried at 60°C for 8 hours to obtain a powdery black solid BB-COF. The synthetic route is as follows:

[0050] .

[0051] (5) Preparation of BB-COF@Mn: Under argon protection, BB-COF (34.72 mg, 0.05 mmol) and pentacarbonyl manganese bromide (Mn(CO)5Br, 55.2 mg, 0.2 mmol) were suspended in anhydrous ether (20 mL) and stirred at room temperature for 4 hours. The precipitate was collected by filtration, washed with anhydrous ether, and dried in vacuo at 60°C for 8 hours to obtain the pentacarbonyl manganese bromide-loaded material BB-COF@Mn. The synthetic route is:

[0052] .

[0053] Example 2: Characterization

[0054] (1) Figure 1 (a) is the infrared spectrum of BB-COF@Mn, which verifies the successful construction of BB-COF@Mn. From the Fourier transform infrared spectrum of BB-COF@Mn, it can be observed that the newly formed C=N bond (1603 cm -1 ). In addition, at 3205 cm -1 and 3332 cm -1 The NH stretching vibration peak of 2,2'-bipyridine-5,5'-diamine disappears, and the characteristic stretching band of C=O of 3,5-diformylphenylboronic acid (1704 cm -1 ) was greatly weakened, indicating that the free aldehyde was completely consumed after polymerization. All these results imply the successful synthesis of the COF skeleton. In addition, the characteristic vibration of the keto carbonyl group of Mn(CO)5Br was found at 2065 cm -1 and 1988 cm -1 There are two prominent strong infrared absorption peaks at the 400 nm region, showing obvious typical carbonyl characteristic peaks, which also proves that BB-COF@Mn has successfully introduced Mn(CO)5Br.

[0055] (2) Figure 1 (b) is the solid-state carbon spectrum of BB-COF@Mn, 13 C - NMR analysis further confirmed the skeletal structure of BB-COF@Mn. A pyrrole ring methyl signal at 12.52 ppm, aromatic carbon peaks at 129.64-135.49 ppm, and a peak characteristic of an imine bond at 151.9 ppm were observed simultaneously, forming the core framework signal. New peaks at 57.57 ppm and 65.48 ppm are attributed to the coordination of Mn(CO)5Br. These results demonstrate the successful synthesis of BB-COF@Mn.

[0056] (3) Figure 1(c) shows the pore size distribution curve drawn according to the nonlocal density functional theory (NLDFT) model, which can be used to visually observe the porosity. It can be seen that BB-COF@Mn shows a wide PSD from 0 nm to 40 nm, with the main pore size distribution at 1.5454 nm. The BET surface area of ​​BB-COF@Mn is calculated to be 195.51 m 2 g -1 , the total pore volume is 0.1459 cm 3 g -1 . Figure 1 (d) is the low-temperature N2 adsorption isotherm of BB-COF@Mn at 77 K. The porosity of the synthesized material BB-COF@Mn was determined using low-temperature nitrogen adsorption. BB-COF@Mn exhibits a typical curve that conforms to a Type III adsorption isotherm, where the adsorption capacity increases slowly at low pressures and gradually increases with increasing pressure. However, the slope of the curve remains positive, indicating that its pore structure is primarily composed of micropores.

[0057] (4) Figure 1 (e) shows the thermogravimetric curves of BB-COF and BB-COF@Mn. Thermogravimetric analysis demonstrates the thermal stability of BB-COF@Mn. Comparing the thermogravimetric analyses of BB-COF and BB-COF@Mn reveals that before reaching 100°C, the material loss rate of BB-COF@Mn is significantly higher due to the volatilization of strongly adsorbed water within the porous framework and the onset of CO release from the loaded Mn(CO)5Br. However, after CO release, the structural stability of BB-COF@Mn improves compared to BB-COF, and the material loss rate decreases.

[0058] (5) Figure 1 (f) is a particle size analysis of BB-COF@Mn. The results show that the particle size of BB-COF@Mn is concentrated in the range of 900 nm to 1500 nm (accounting for 90% of the total), showing an overall normal distribution, indicating that the particle size of BB-COF@Mn is relatively stable.

[0059] (6) From Figure 2 (a) ~ Figure 2 (l) It can be seen that the BB-COF@Mn prepared in the example has an egg-shell spherical structure, and the lattice fringes are clearly discernible. The difference in light and dark contrast confirms its highly ordered crystal arrangement. In addition, there are abundant microporous structures inside the BB-COF@Mn. TEM comparative analysis shows that the loading of Mn(CO)5Br significantly changes the surface morphology of the material. Figure 2 (e)~ Figure 2 (h) It can be seen that the surface of BB-COF without Mn(CO)5Br is smooth, while Figure 2 (i) ~ Figure 2(l) shows that the surface of BB-COF@Mn presents a uniformly distributed rough texture, indicating the successful anchoring of pentacarbonyl manganese bromide.

[0060] (7) Figure 2 (m)- Figure 2 (t) is the transmission electron microscopy mapping of BB-COF@Mn and the distribution of elements such as C, N, and O in BB-COF@Mn. It can be seen that C (56.78%), N (8.39%), O (8.82%), Mn (10.36%), Br (10.37%), and F (1.5%) are evenly distributed in BB-COF@Mn, which indicates that the structural composition of the material has a certain uniformity. Figure 3 This is the EDS diagram of BB-COF@Mn. From the X-ray energy spectrum (EDX), it can be found that C, N, O, Mn, Br, and F elements are widely present.

[0061] (8) BB-COF@Mn was irradiated with a 638 nm laser for 900 seconds to measure its photothermal effect. BB-COF@Mn aqueous dispersions of different concentrations (0, 50, 100, and 200 μg / mL) were prepared in 2 mL centrifuge tubes using deionized water and BB-COF@Mn. The total volume of the BB-COF@Mn aqueous dispersions at each concentration was 1 mL. Figure 4 (a) is 1.5W / cm 2 The concentration-dependent photothermal effect of BB-COF@Mn under 638 nm laser irradiation is shown in Figure 2. 2 When the concentration of BB-COF@Mn increases, the temperature change caused by the heating of the suspension becomes larger and larger. Figure 4 (b) is at 0.4, 0.7, 1.0, 1.3 and 1.5 W / cm 2 Laser power-dependent photothermal effect of BB-COF@Mn under 638 nm laser irradiation. Figure 4 (c) is a thermal imaging image of the heating process of BB-COF@Mn (0, 50, 100, and 200 μg / mL) within 15 minutes. These results indicate that BB-COF@Mn has high photothermal conversion efficiency and concentration-dependent photothermal performance.

[0062] (9) Figure 5 (a) is 638nm laser at 1.5 W / cm 2 The temperature change curve of BB-COF@Mn (200μg / mL) was obtained by three cycles of light cooling. The temperature change of BB-COF@Mn was recorded by ON / OFF cycle irradiation experiment. The results showed that after three cycles (638nm, 1.5W / cm 2) showed good photothermal stability. The highest temperature of the three cycles could reach about 62°C, and there was no obvious change in the temperature rise and cooling trend. The above results show that BB-COF@Mn has good photothermal effect and photothermal stability, making it a potential photothermal antibacterial agent. In addition, the photothermal conversion efficiency of BB-COF@Mn was also investigated. Figure 5 (b) BB-COF@Mn (200 μg / mL) aqueous dispersion under 638 nm laser irradiation (1.5 W / cm 2 ) photothermal effect. When BB-COF@Mn (200µg / mL) was exposed to 638nm (1.5W / cm 2 ) when irradiated with a laser. The temperature increased to 62.8°C. After stopping the laser irradiation, the temperature dropped to room temperature. Therefore, the calculated photothermal conversion efficiency was 59.8% (for the specific calculation formula, see the photothermal conversion efficiency formula disclosed in patent application number CN116333336A, "A Ferrocene-Based Metal-Organic Framework Material, Its Preparation Method, and Applications").

[0063] (10) Photodynamic performance testing of BB-COF@Mn. Figure 6 The electron paramagnetic resonance spectrum shows that BB-COF@Mn can catalyze the generation of singlet oxygen. Figure 7 (a) ~ Figure 7 (b) Demonstrates the ability of BB-COF@Mn to produce singlet oxygen. Using DPBF as a probe and 808nm laser as a light source, the photodynamic properties of BB-COF@Mn were detected by UV-visible spectroscopy. By controlling the laser irradiation time and monitoring the absorption intensity of DPBF (1 mg / mL, 30μL, DMSO as solvent) in the mixture (DPBF+BB-COF@Mn+light) at a wavelength of 418 nm, the ability of BB-COF@Mn to produce singlet oxygen was evaluated. The results showed that compared with the blank control (DPBF+light), BB-COF@Mn+DPBF+light (200μg / ml, 638nm laser 1.5 W / cm 2 ) A large amount of singlet oxygen can be produced in the first minute, and the peak value of DPBF drops significantly and rapidly, and the subsequent peak value decrease slows down. After 15 minutes of illumination, 99% of DPBF is consumed, which means that BB-COF@Mn produces a large amount of singlet oxygen under illumination.

[0064] (11) Photocontrolled CO release ability and Fenton reaction detection of BB-COF@Mn. In order to study the CO release characteristics of BB-COF@Mn, myoglobin (Mb) experiments were carried out.

[0065] Mb in horse skeletal muscle was first reduced to deoxymyoglobin (Deoxy-Mb) using sodium dithionite, followed by the addition of BB-COF@Mn. Deoxy-Mb reacted with CO to form monomyoglobin (MbCO). CO release was measured as the difference in UV-visible (UV-vis) absorbance between Deoxy-Mb (557 nm) and MbCO (540 and 577 nm).

[0066] from Figure 8 (a) and Figure 8 As can be clearly observed in (b), the absorbance of Deoxy-Mb barely changes in the absence of BB-COF@Mn and light. This indicates that no significant reaction occurs under these conditions. However, upon addition of BB-COF@Mn and laser irradiation, the position and intensity of the absorption peak shift. This result fully demonstrates that BB-COF@Mn can effectively control CO release under laser irradiation.

[0067] In addition, the presence of Mn(CO)5Br endows BB-COF@Mn with peroxidase-like activity. Therefore, a dual substrate system was used, namely H2O2 and 3,3',5,5'-tetramethylbenzidine (TMB), in which TMB was used as an indicator of the decomposition of H2O2 to generate •OH under the catalysis of BB-COF@Mn. The experiment was divided into four groups: TMB, TMB+H2O2, TMB+H2O2+BB-COF, and TMB+H2O2+BB-COF@Mn. Figure 8 As can be seen in (c), neither the TMB group nor the H2O2+TMB group produced a visible blue change and detectable UV-visible absorption at a wavelength of 650 nm. The BB-COF@Mn + H2O2+TMB group had obvious UV-visible absorption at 650 nm. In contrast, even in the presence of H2O2 and TMB, the BB-COF solution (i.e., TMB+H2O2+BB-COF group) did not produce a blue change or detectable UV-visible absorption at a wavelength of 650 nm. This result indicates that the peroxidase-like catalytic activity of BB-COF@Mn is attributed to the presence of Mn(CO)5Br. Figure 8 As shown in (d), the experimental conditions are: 1.5 mmol / L TMB, 20 mmol / L H₂O₂, and triggering in an aqueous environment with a pH of 4.5. As the concentration of BB-COF@Mn increases (50, 100, 125, 150, and 200 μg / mL), the system produces significantly more •OH, and its UV absorption at 650 nm also increases. These results demonstrate that BB-COF@Mn has the ability to generate hydroxyl radicals.

[0068] Test Example 1: In vitro bacterial growth inhibition test

[0069] Take 5 mL of LB liquid medium and add it to the shaking tube. Use a bacterial pick stick to pick a colony from the evenly spread LB solid medium containing Staphylococcus aureus or Escherichia coli and place it in the above 5 mL of LB liquid medium. Place the shaking tube in a shaker and culture at 37°C for 8-12 hours until the final OD value of the bacterial solution at 600 nm is 0.1 (approximately equivalent to a bacterial concentration of 1×10 8 CPU / mL), and Staphylococcus aureus and Escherichia coli bacterial liquids were obtained respectively.

[0070] (1) In order to verify the effect of BB-COF@Mn on bacterial activity, BB-COF@Mn dispersions of different concentrations (0, 50, 100, and 200 μg / mL) were prepared with PBS at pH 7 and 20 μL of H2O2 (20 mmol) was added and incubated with 100 μL of Staphylococcus aureus solution (1×10 8 CPU / mL) or Escherichia coli solution (1×10 8 CPU / mL) was mixed and then made up to 1mL with pH7 PBS. After 15min of 638nm laser irradiation, the cells were cultured in a shaker at 37℃ for 8h. Different numbers of colonies were shown in the bacterial culture dishes. Figure 9 (a) is a picture of Staphylococcus aureus and Escherichia coli culture after treatment with different concentrations of BB-COF@Mn; Figure 9 (b) Plate count assay for the survival of Staphylococcus aureus and Escherichia coli. The colony counts of S. aureus and E. coli decreased with increasing BB-COF@Mn concentration. When the BB-COF@Mn concentration reached 200 µg / mL, the survival rates of S. aureus and E. coli were reduced to 0.41% and 0.84%, respectively, after 15 minutes of irradiation.

[0071] (2) To compare the antibacterial activity of the four antibacterial modes of thermal therapy, photodynamic therapy, Fenton reaction and CO release, Staphylococcus aureus and Escherichia coli were treated with different groups and divided into 8 groups: PBS group (Group I), H2O2 (Group II), PBS+laser group (Group III), BB-COF group (Group IV), BB-COF+laser group (Group V), BB-COF@Mn group (Group VI), BB-COF@Mn+laser group (Group VII), and BB-COF@Mn+H2O2+laser group (Group VIII).

[0072] The specific treatment process of group I and group III was as follows: 80 μL of PBS with pH 7 was mixed with 100 μL of Staphylococcus aureus liquid (1×10 8CPU / mL) or Escherichia coli solution (1×10 8 CPU / mL) was mixed, and then supplemented to 1 mL with pH 7 PBS to obtain group I; after 15 minutes of 638 nm laser irradiation, group V was obtained.

[0073] The specific treatment process of group II was as follows: 80 μL of PBS with pH 7 was mixed with 100 μL of Staphylococcus aureus liquid (1×10 8 CPU / mL) or Escherichia coli solution (1×10 8 CPU / mL) were mixed, 20 μL of H2O2 (20 mmol) was added, and then the volume was made up to 1 mL with PBS at pH 7 to obtain Group II.

[0074] The specific treatment process of Groups IV and V was as follows: BB-COF prepared in step (4) of Example 1 was mixed with PBS at pH 7 to obtain a BB-COF solution; 200 μL of BB-COF solution (1 mg / mL) was mixed with 100 μL of Staphylococcus aureus solution (1×10 8 CPU / mL) or Escherichia coli solution (1×10 8 CPU / mL) was mixed, and then supplemented to 1 mL with pH 7 PBS to obtain group IV; after 15 minutes of 638 nm laser irradiation, group V was obtained.

[0075] The specific treatment process of Groups VI, VII, and VIII was as follows: BB-COF@Mn prepared in Example 1 was mixed with PBS at pH 7 to obtain BB-COF@Mn solution; 200 μL BB-COF@Mn solution (1 mg / mL) was mixed with 100 μL Staphylococcus aureus solution (1×10 8 CPU / mL) or Escherichia coli solution (1×10 8 CPU / mL) was mixed, and then supplemented to 1 mL with pH 7 PBS to obtain group VI; after 15 minutes of 638 nm laser irradiation, group VII was obtained; and 20 μL of H2O2 (20 mmol) was added to obtain group VIII.

[0076] The above 8 groups were treated and cultured in a shaker at 37℃ for 8h. Different numbers of colonies appeared in the bacterial culture dishes. Then the number of bacteria in each group was estimated by plate counting method. Figure 10As shown, using the PBS group (Group I) as the control group, a considerable number of bacterial colonies were observed in Groups II, III, IV, and VI. In other words, there was no significant difference in bacterial colony counts among the PBS, H2O2, BB-COF, and BB-COF@Mn groups, regardless of whether laser irradiation was performed. The BB-COF + laser group (Group V) exhibited a moderate bactericidal effect, reducing the number of Staphylococcus aureus to 54.6% and Escherichia coli to 65.8%, indicating that BB-COF caused a certain number of bacterial deaths through photothermal and photodynamic methods. The BB-COF@Mn + laser group (Group VII) also exhibited a similar bactericidal effect, reducing the number of Staphylococcus aureus to 32.1% and Escherichia coli to 39.5%. After treatment with the BB-COF@Mn+H2O2+laser group (Group VIII), the numbers of Staphylococcus aureus and Escherichia coli decreased to 0.41% and 0.83%, respectively. This indicates that the photothermal, photodynamic, and CO2-releasing properties of BB-COF@Mn exerted significant bactericidal effects. This demonstrates the remarkable four-strategy combined broad-spectrum antibacterial properties of BB-COF@Mn.

[0077] Test Example 2: Transmission electron microscopy observation of bacterial morphology

[0078] To further understand the above antibacterial effects, TEM was used to study the changes in the morphological structure of Escherichia coli and Staphylococcus aureus ( Figure 11), the grouping was the same as that of Experimental Example 1 (2), and was divided into 8 groups: PBS group (Group I), H2O2 (Group II), PBS + laser group (Group III), BB-COF group (Group IV), BB-COF + laser group (Group V), BB-COF@Mn group (Group VI), BB-COF@Mn + laser group (Group VII), and BB-COF@Mn + H2O2 + laser group (Group VIII). The bacterial cell walls of the PBS group (I), H2O2 group (II) and PBS+laser group (III) were intact and smooth, and the flagellar structure of Escherichia coli was clearly visible, indicating that PBS, light and H2O2 treatment had no antibacterial effect on Staphylococcus aureus and Escherichia coli; the BB-COF group (IV) only caused local membrane wrinkling, while the BB-COF+laser group (V) led to increased membrane permeability; BB-COF@Mn (VI) also only caused local membrane wrinkling, indicating that it had only limited antibacterial effect in the absence of laser irradiation and CO release; and in the BB-COF@Mn+laser group (VII), the bacterial membrane structure and cell matrix were destroyed under the synergistic effect of photothermal effect and CO release; in the BB-COF@Mn+H2O2+laser group (VIII), the hydroxyl radicals (•OH) produced by the Fenton reaction acted together with CO, resulting in the destruction of the bacterial membrane structure and the denaturation and leakage of the cell matrix. Therefore, it can be concluded that BB-COF@Mn can achieve rapid and efficient killing of Staphylococcus aureus and Escherichia coli through the combined application of four strategies induced by 638 nm laser: photothermal, photodynamic, Fenton reaction, and CO release.

[0079] Test Example 3: Hemolysis Test

[0080] Fresh blood was collected from female BALB / c mice (purchased from Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd.). Red blood cells (RBCs) were collected by centrifugation at 1500 rpm for 20 minutes and washed three times with PBS. RBCs (4% w / w) were mixed with various concentrations of BB-COF@Mn in PBS (50, 100, 150, 200, and 250 µg / mL) at a 1:9 (v / v) ratio and incubated at 37°C on a shaker for 3 hours. The mixture was then centrifuged at 12,000 rpm for 20 minutes. The hemolytic effect of BB-COF@Mn was determined by UV-visible spectroscopy, measuring the absorbance at 540 nm of the supernatant after centrifugation of the RBC solutions incubated with various concentrations of BB-COF@Mn. Distilled water served as a positive control (+), and PBS served as a negative control (-). Figure 12 (a) is the hemolysis rate of BB-COF@Mn at different concentrations on red blood cells. It was found that when the concentration of BB-COF@Mn increased from 50µg / mL to 250µg / mL, the hemolysis rate remained below 1.5%, which indicates that the material has good biocompatibility.

[0081] Test Example 4: Cytotoxicity Test

[0082] Cytotoxicity is also an important indicator for evaluating the biocompatibility of materials. Human embryonic kidney cells (HEK293 cells) and mouse fibroblasts (L929 cells) were used for testing.

[0083] HEK293 cells or L929 cells were plated at 8 × 10 cells per well in a 96-well plate. 3 Cells were seeded at a density of 100 μL per well, with 100 μL of cell suspension added to each well. Surrounding wells were sealed with 100 μL of PBS to prevent excessive evaporation. After a 24-hour incubation, cells were treated with PBS (control) and various concentrations of BB-COF@Mn in PBS (50, 100, 150, and 200 μg / mL). After 24 hours, the supernatant was discarded, and an equal volume of culture medium containing 10 μL of MTT (5 mg / mL) in PBS (pH 7.4) was added to each well. After 4 hours, the supernatant was aspirated, and 100 μL of DMSO was added to each well to dissolve the MTT-formamide crystals. After 5 minutes, the absorbance was measured at 490 nm in a microplate reader. Figure 12 (b) shows the changes in cell viability after incubation of HEK293 cells or L929 cells with different concentrations of BB-COF@Mn. It can be seen that with the increase in the concentration of BB-COF@Mn, the survival rate of HEK293 cells and L929 cells slowly decreased after 24 hours. At a concentration of 200 μg / mL, the survival rate of HEK293 cells and L929 cells still remained above 85%, which once again shows that BB-COF@Mn has good biocompatibility.

[0084] The above description is merely a preferred embodiment of the present application and is not intended to limit the present application. Various modifications and variations are possible for those skilled in the art. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present application shall be included within the scope of protection of the present application.

Claims

1. A method for preparing an organic polymer composite material based on BODIPY light-controlled CO release, characterized in that: The following steps are involved: (1) BODIPY undergoes iodination reaction with an iodine-containing compound to obtain iodinated BODIPY; (2) BODIPY iodide, 3,5-diformylphenylboronic acid and potassium carbonate are dissolved in an organic solvent, a catalyst is added under a protective atmosphere and the reaction is carried out to obtain tetraaldehyde BDP; (3) dissolving tetraaldehyde BDP and polyamino bipyridine in an organic solvent, ultrasonically dispersing to form a suspension, cyclically deoxidizing, and heating in a sealed environment to react to obtain a powdery black solid BB-COF; the polyamino bipyridine is 2,2'-bipyridine-5,5'-diamine; (4) Under a protective atmosphere, BB-COF and pentacarbonyl manganese bromide were suspended in anhydrous ether and stirred at room temperature to obtain an organic polymer composite material based on BODIPY photocontrolled CO release.

2. The preparation method according to claim 1, characterized in that In step (1), the iodine-containing compound is N-iodosuccinimide; the molar ratio of BODIPY to N-iodosuccinimide is 1:2; and the iodination reaction is performed by dissolving BODIPY and N-iodosuccinimide in anhydrous dichloromethane and stirring the mixture at room temperature.

3. The preparation method according to claim 1, characterized in that In step (2), the molar ratio of the iodinated BODIPY to 3,5-diformylphenylboronic acid is 1:2; the organic solvent is 1,4-dioxane; and the catalyst is tetrakis(triphenylphosphine)palladium.

4. The preparation method according to claim 1, characterized in that In step (2), the reaction temperature is 120° C. and the reaction time is 12 h.

5. The preparation method according to claim 1, characterized in that In step (3), the organic solvent is a mixture of toluene and acetic acid solution in a volume ratio of 20:1; the concentration of the acetic acid solution is 3M; the temperature of the heating reaction is 120°C, and the reaction time is 72h.

6. The preparation method according to claim 1, characterized in that The molar ratio of the tetraaldehyde BDP to 2,2'-bipyridine-5,5'-diamine is 1:

2.

7. The preparation method according to claim 1, characterized in that In step (4), the molar ratio of BB-COF to pentacarbonyl manganese bromide is 1:4; and the reaction time under room temperature stirring is 4 hours.

8. The organic polymer composite material based on BODIPY light-controlled CO release obtained by the preparation method according to any one of claims 1 to 7, characterized in that: The organic polymer composite material based on BODIPY light-controlled CO release is obtained by polymerizing polyaldehyde BDP and polyamino bipyridine as monomers, and using bipyridine's multidentate ligand to load pentacarbonyl manganese bromide. The organic polymer composite material based on BODIPY light-controlled CO release has an eggshell-like spherical structure and releases CO under laser irradiation.

9. The organic polymer composite material based on BODIPY light-controlled CO release according to claim 8, characterized in that: The polyaldehyde BDP is a tetraaldehyde BDP; and the polyamino bipyridine is 2,2'-bipyridine-5,5'-diamine.

10. Use of the organic polymer composite material based on BODIPY light-controlled CO release according to claim 8 or 9 in the preparation of antibacterial drugs.

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