A nanospherical porous copper porphyrin, its preparation method and use

Abstract

The present invention belongs to the technical field of material preparation and application, and discloses a nano-spherical porous copper porphyrin, a preparation method thereof and uses thereof. The present invention uses gallium black phosphorus as a template to drive the coordination of 5,10,15,20-tetrakis(4-aminophenyl)porphyrin and terephthaloyl chloride to induce the directional formation of nano-porous copper porphyrin with gallium black phosphorus as the core; the nano-porous copper porphyrin material prepared by the present invention is a porous sphere, and has excellent photodynamic, chemical kinetic and photothermal properties. Under dark conditions, it has the characteristics of catalase and catalyzes the production of hydroxyl radicals from hydrogen peroxide; under red light irradiation, it can continuously generate singlet oxygen; under near-infrared light irradiation, it has good photothermal properties, and this material is expected to be applied to the treatment of drug-resistant bacterial infections.

Description

Technical Field

[0001] The present invention relates to a nano-spherical porous copper porphyrin and its preparation method and use, belonging to the technical field of material preparation and application. Background Art

[0002] Bacterial resistance has become a global public health safety issue of concern. Super bacteria with multiple drug resistance mechanisms continue to emerge, and there are frequent occurrences of incurable cases due to bacterial infections around the world. Photo-responsive antibacterial, as a fast, efficient, and non-resistant antibacterial method, has received extensive attention. Photo-responsive antibacterial materials generate reactive oxygen species (ROS) by absorbing light of appropriate wavelengths. ROS can penetrate the bacterial cell membrane, enter the bacteria, and damage proteins and lipids, thereby disrupting bacterial respiration and physiological activities. At the same time, there will be overexpression of hydrogen peroxide in the microenvironment of bacterial infection. Nano-catalase can catalyze hydrogen peroxide to generate hydroxyl radicals, which is also a kind of reactive oxygen species and can inhibit the growth of bacteria. At the same time, too high a concentration of hydrogen peroxide will cause irreversible damage to DNA, RNA, and proteins in normal tissues and affect intracellular signal transduction through signal proteins that control cell proliferation. Therefore, the decomposition of hydrogen peroxide by catalase can not only improve antibacterial activity, inhibit the formation of biofilms but also relieve tissue damage.

[0003] Porphyrin is a photosensitizer with high biocompatibility and has been widely used in bacterial photodynamic therapy and photothermal therapy. However, the poor water solubility of porphyrin leads to a low yield of reactive oxygen species, and the low antibacterial efficiency limits its biological applications. Black phosphorus, due to its phosphorene structure and good biocompatibility, is a promising metal-free nanomaterial in the biological field. The hydrophobicity of the P-P bond in black phosphorus itself and the hydrophilicity of the P-O bond after oxidation make it a natural surfactant, which can regulate the assembly mode of metal porphyrin. Using metal black phosphorus as a template can increase the optical activity of porphyrin derivatives, thereby increasing antibacterial activity. Therefore, studying new metal black phosphorus porphyrin antibacterial agents can improve the application potential of porphyrin derivatives in the field of bacterial infection treatment. Summary of the Invention

[0004] The present invention aims at some defects existing in the prior art and provides a nano-spherical porous copper porphyrin (denoted as BPGa@COF-Cu) and its preparation method and application.

[0005] The present invention first provides a nano-spherical porous porphyrin copper, denoted as BPGa@COF-Cu. The mass percentages of metallic gallium and copper in the nano-porous porphyrin copper are 0.7% and 4.2% respectively; the nano-porous porphyrin copper material contains black phosphorus gallium and porphyrin copper; the material components include C, N, Ga, O, P, and Cu; infrared spectra and XPS spectra show the presence of P-Ga bonds, amide bonds, and N-Cu bonds. BPGa@COF-Cu is a spherical nanoparticle with a specific surface area of 17.40 m 2 / g, average pore volume 0.05m 2 / g, the average pore size is 18.36nm, and the average particle size of nanoparticles is 344.8nm. Infrared spectrum shows that BPGa@COF-Cu has a peak at 1667cm -1 A new characteristic absorption peak is generated, corresponding to the stretching vibration of the carbon group of the conjugated amide bond, 1000-1500 cm -1 The multiple absorption peaks are characteristic peaks of porphyrin; the XPS spectrum shows that the constituent elements include C, N, Ga, O, P, and Cu, including P-Ga bonds, amide bonds, and N-Cu bonds.

[0006] The present invention also provides a preparation method of BPGa@COF-Cu. In the present invention, black phosphorus crystals are first obtained by grinding blocky black phosphorus; black phosphorus gallium is prepared in one step by a liquid phase exfoliation ultrasonic method, wherein gallium and black phosphorus are connected by a P-Ga bond; then, black phosphorus gallium is used as a template to induce the directional formation of 5,10,15,20-tetrakis(4-aminophenyl)porphyrin and terephthaloyl chloride to generate an amide bond and wrap the black phosphorus gallium; finally, porous spherical BPGa@COF-Cu is obtained by coordination-induced assembly between the N atoms of the porphyrin and copper ions, wherein the copper ions and the porphyrin are connected by an N-Cu bond.

[0007] The specific steps are as follows:

[0008] (1) Disperse the black phosphorus gallium compound, 5,10,15,20-tetrakis(4-aminophenyl)porphyrin, and terephthaloyl chloride in tetrahydrofuran solution, respectively. Slowly add the black phosphorus gallium tetrahydrofuran solution to the 5,10,15,20-tetrakis(4-aminophenyl)porphyrin tetrahydrofuran solution, then add the terephthaloyl chloride tetrahydrofuran solution, and finally add triethylamine. Mix well and stir to react. After the reaction is completed, centrifuge, wash with tetrahydrofuran, and vacuum dry to obtain the black phosphorus gallium porphyrin compound.

[0009] Among them, the dosage ratio of gallium black phosphorus, 5,10,15,20-tetra(4-aminophenyl)porphyrin, terephthaloyl chloride and triethylamine is 1-10 mg: 10-200 mg: 10-100 mg: 10-50 μL; preferably, the dosage ratio of gallium black phosphorus, 5,10,15,20-tetra(4-aminophenyl)porphyrin, terephthaloyl chloride and triethylamine is 5 mg: 101 mg: 60 mg: 20 μL;

[0010] The reaction temperature is 10-50 °C, and the reaction time is 1-24 hours; preferably, the reaction temperature is 25 °C and the reaction time is 12 hours.

[0011] (2) Disperse the gallium black phosphorus porphyrin compound and copper chloride in N,N-dimethylformamide (DMF) solution respectively, drop the copper chloride DMF solution into the gallium black phosphorus porphyrin DMF solution, and stir for reaction. After the reaction is completed, centrifuge, wash with DMF, and dry in vacuum to obtain the gallium black phosphorus porphyrin copper compound.

[0012] Among them, the mass ratio of the gallium black phosphorus porphyrin to copper chloride is 5:1-1:10; preferably, the mass ratio of the gallium black phosphorus porphyrin to copper chloride is 1:5;

[0013] The reaction temperature is 10-150 °C, and the reaction time is 1-20 hours; preferably, the reaction temperature is 100 °C and the reaction time is 10 hours.

[0014] The nano-spherical porous porphyrin copper material prepared in the present invention has the ability to absorb near-infrared light and convert it into heat, and can generate singlet oxygen under red light irradiation. In addition, it can also catalyze hydrogen peroxide to generate hydroxyl radicals.

[0015] The present invention also provides the use of nano-spherical porous porphyrin copper (BPGa@COF-Cu), and the use is for the inhibition of Escherichia coli or Staphylococcus aureus.

[0016] The bacteriostatic method of BPGa@COF-Cu is as follows:

[0017] Mix 10 5 CFU / mL of the bacterial solution with the corresponding concentration of the compound, and then judge the bacteriostatic situation through plate pictures after incubation. The dark group is the compound incubated with the bacteria for 2 hours, and the light group is the compound incubated with the bacteria for 2 hours under red light irradiation. Finally, take 200 μL and spread it evenly on the agar plate. After incubating for 12 hours, take pictures to record the plate pictures.

[0018] The beneficial effects of the present invention:

[0019] The present invention discloses a preparation method of a nano-spherical porous copper porphyrin material. In the prior art, metal porphyrins have a single-molecule or metal MOF structure. For the first time, the present invention uses a gallium phosphide template to induce the directional coupling of porphyrins through coordination and dipole interactions to form porous nano-copper porphyrin with gallium phosphide as the core, and this structure is reported for the first time. In addition, the one-step directional synthesis reaction method using gallium phosphide as the template is significantly different from the method of metal coordination polymerization synthesis in the literature, belonging to an innovation in the synthesis method. The prepared BPGa@COF-Cu has high photoactivity. Using 1,3-diphenylisobenzofuran (DPBF) as a singlet oxygen scavenger, it is found that singlet oxygen can be continuously generated under red light irradiation, and the activity is significantly higher than that of metal porphyrins. At the same time, using methylene blue (MB) as a hydroxyl radical scavenger, it is found that this material can catalyze hydrogen peroxide to generate hydroxyl radicals as catalase. In addition, the ability of this material to continuously generate heat under near-infrared light (808 nm) irradiation is further verified. At the same time, the antibacterial experiment shows that the spherical porous structure enables BPGa@COF-Cu to have high antibacterial activity in the dark, which is significantly different from the current use of porphyrins mostly for photo-driven singlet oxygen photodynamic antibacterial therapy, indicating that BPGa@COF-Cu is a new type of antibacterial material. The present invention has double innovations in the synthesis method and new material design. Description of the Drawings

[0020] Figure 1 Schematic diagram of the synthesis and antibacterial mechanism of BPGa@COF-Cu.

[0021] Figure 2 Transmission electron micrograph of BPGa@COF-Cu.

[0022] Figure 3 Infrared spectrum of BPGa@COF-Cu.

[0023] Figure 4 XPS spectrum of BPGa@COF-Cu, where (a) is the total spectrum, (b) is the C1s spectrum, (c) is the O1s spectrum, (d) is the N1s spectrum, (e) is the P 2p spectrum, and (f) is the Cu 2p spectrum.

[0024] Figure 5 Degradation curve of DPBF by BPGa@COF-Cu within 0 - 8 minutes.

[0025] Figure 6 Degradation curve of MB by BPGa@COF-Cu within 10 minutes.

[0026] Figure 7 Heating curves of BPGa@COF-Cu at concentrations of 0, 0.3, 0.6, 0.8, and 1 mg / mL under 808 nm light irradiation.

[0027] Figure 8 Plated pictures showing the inhibition of Escherichia coli by BPGa@COF-Cu under dark conditions or red light irradiation.

[0028] Figure 9 Plated pictures showing the inhibition of Staphylococcus aureus by BPGa@COF-Cu under dark conditions or red light irradiation. Specific implementation manners

[0029] To enable those skilled in the art to better understand the technical solution of the present invention, the preferred embodiments of the present invention are described in detail below. However, the following embodiments do not limit the protection scope of the present invention.

[0030] In the embodiments of the present invention, the experimental methods without specific conditions are carried out according to the conventional methods and conditions in the art, and the materials used are commercially available as conventional without special instructions.

[0031] Reagents and instruments used in the embodiments of the present invention: The solvents used in the reaction are all of analytical grade, and the reagents used are directly applied without special treatment without any special treatment.

[0032] Black phosphorus: Nanjing Xianfeng Nano Materials Co., Ltd.;

[0033] Tetrahydrofuran, N,N-dimethylformamide (DMF): Analytical grade, Shanghai Pilot Chemical Industry Corporation;

[0034] Gallium chloride, copper chloride, terephthaloyl chloride, methylene blue (MB), 1,3-diphenylisobenzofuran (DPBF): Analytical grade, Aladdin Industrial Corporation;

[0035] 5,10,15,20-Tetra(4-aminophenyl)porphyrin: Analytical grade, Jilin Zhongke Yanshen Technology Co., Ltd.

[0036] Example 1: Preparation of BPGa@COF-Cu

[0037] In this example, the material preparation process is carried out under nitrogen protection.

[0038] (1) Take gallium black phosphorus (5 mg) and disperse it in 1 mL of tetrahydrofuran solution. Take 5,10,15,20-tetra(4-aminophenyl)porphyrin (0.15 mmol, 101 mg) and dissolve it in 5 mL of tetrahydrofuran solution. Dropwise add triethylamine (20 μL), and after fully stirring for 10 minutes, add 5 mL of a tetrahydrofuran solution of terephthaloyl chloride (0.3 mmol, 60 mg). After stirring at 25 °C for 12 hours at room temperature, let it stand to separate and remove the precipitate to obtain a suspension. The suspension is centrifuged to obtain a precipitate. After washing the precipitate with tetrahydrofuran, it is centrifuged and dried in vacuo to obtain 100 mg of red solid gallium black phosphorus porphyrin.

[0039] (2) Disperse gallium phosphoporphyrin (50 mg) in 10 mL of DMF solution, dissolve copper chloride (250 mg) in 10 mL of DMF solution. After mixing and stirring the two, heat up to 100 °C and continuously stir for 10 hours. After cooling to room temperature, let it stand and separate the lower-layer precipitate. Take the upper-layer solution and centrifuge to obtain a precipitate. Wash it three times with DMF and ethanol solutions respectively and then centrifuge. Dry it under vacuum to obtain a brick-red solid BPGa@COF-Cu (100 mg).

[0040] Figure 1 Synthesis and structural schematic diagram of BPGa@COF-Cu.

[0041] Figure 2 Transmission electron microscope image of BPGa@COF-Cu, which shows that BPGa@COF-Cu is composed of spherical porous structures with a radius of about 200 nm.

[0042] Figure 3 Fourier transform infrared spectroscopy of BPGa@COF-Cu, which shows that 1667 cm -1 is the characteristic absorption peak of the carbonyl group of the generated conjugated amide bond; 3320 cm -1 is the characteristic absorption peak of secondary amine (-NH); the results prove the formation of amide bonds.

[0043] Figure 4 XPS spectrum of BPGa@COF-Cu, where (a) is the total spectrum, (b) is the C1s spectrum, (c) is the O1s spectrum, (d) is the N1s spectrum, (e) is the P 2p spectrum, and (f) is the Cu 2p spectrum. The total spectrum shows that BPGa@COF-Cu is mainly composed of C, N, O, P, Ga, and Cu elements; in the binding energy spectrum of C 1s, 288.1 eV corresponds to the binding energy of the C=O double bond, and in the binding energy spectrum of O 1s, 531.8 eV also corresponds to the binding energy of C=O, which can prove the existence of amide bonds; in the binding energy spectrum of N 1s, 400.2 eV corresponds to the binding energy of N-Cu; in the binding energy spectrum of P 2p, 133.7 eV corresponds to the binding energy of P-Ga. BPGa@COF-Cu degradation of DPBF experiment

[0044] Prepare an acetonitrile solution of 1,3-diphenylisobenzofuran with a concentration of 100 μM, and add BPGa@COF-Cu (10 μg / mL). After irradiating with a red LED lamp for a certain time, dilute it and use a UV spectrophotometer to record the degradation curve of DPBF, as Figure 5 shown. Figure 5Degradation curve of DPBF by BPGa@COF-Cu under red light irradiation within 0 - 8 minutes. DPBF is a sensitive singlet oxygen probe, and singlet oxygen can cause the degradation of DPBF, that is, the absorbance at 415 nm decreases. The results show that BPGa@COF-Cu can generate singlet oxygen under red light irradiation.

[0045] Experiment on the degradation of MB by BPGa@COF-Cu

[0046] Prepare an aqueous solution of methylene blue containing 1 mg / mL, add BPGa@COF-Cu (1 mg / mL), and mix and stir for 10 minutes. Add an aqueous hydrogen peroxide solution (10 μL), and after reacting for the corresponding time, dilute and use a UV spectrophotometer to record the degradation curve of MB, as Figure 6 shown. Figure 6 Degradation curve of MB by BPGa@COF-Cu within 10 minutes. MB is a sensitive hydroxyl radical probe, and hydroxyl radicals can degrade MB, resulting in a decrease in absorbance at 660 nm. The results show that BPGa@COF-Cu can catalyze hydrogen peroxide to generate hydroxyl radicals.

[0047] Figure 7 Temperature increase curves of BPGa@COF-Cu at concentrations of 0, 0.3, 0.6, 0.8, and 1 mg / mL under 808 nm light irradiation. The blank group is an aqueous solution, and it is found that the temperature does not increase significantly. However, with the increase in the concentration of BPGa@COF-Cu, the temperature increases significantly. This shows that BPGa@COF-Cu has good photothermal properties.

[0048] Antibacterial experiment of BPGa@COF-Cu

[0049] The blank group is a bacterial solution with a concentration of 10 5 CFU / mL, and the experimental group is a bacterial solution with 10 5 CFU / mL and a compound solution with a specific concentration gradient. The dark group is the compound incubated with bacteria for 2 hours, and the light group is the compound incubated with bacteria for 2 hours under red light irradiation. Finally, take 200 μL and evenly coat it on an agar plate. After incubating for 12 hours, take pictures to record the plate images, which are respectively Figure 8 and Figure 9 .

[0050] Figure 8 Are the plate images of the inhibition of Escherichia coli by BPGa@COF-Cu under dark conditions or red light irradiation. The results show that the minimum bactericidal concentration of BPGa@COF-Cu against Escherichia coli under red light irradiation is 1 μg / mL.

[0051] Figure 9The plate picture shows the inhibition of Staphylococcus aureus by BPGa@COF-Cu under dark conditions or red light irradiation. The results show that the minimum bactericidal concentration of BPGa@COF-Cu against Staphylococcus aureus under red light irradiation is 3 μg / mL.

[0052] Example 2: Preparation of BPGa@COF-Cu

[0053] In this example, the material preparation process is carried out under nitrogen protection.

[0054] (1) Take black phosphorus copper (1 mg) and disperse it in 1 mL of tetrahydrofuran solution. Take 5,10,15,20-tetrakis(4-aminophenyl)porphyrin (0.15 mmol, 101 mg) and dissolve it in 5 mL of tetrahydrofuran solution. Dropwise add triethylamine (20 μL). After stirring well at 25 °C for 10 minutes, add 5 mL of tetrahydrofuran solution containing terephthaloyl chloride (0.3 mmol, 60 mg). Stir at 25 °C for 6 hours at room temperature, then let it stand and separate to remove the precipitate to obtain a suspension. Centrifuge the suspension to obtain a precipitate. After washing the precipitate with tetrahydrofuran and centrifuging, vacuum dry to obtain 100 mg of red solid black phosphorus copper porphyrin. (2) Take black phosphorus copper porphyrin (50 mg) and disperse it in 10 mL of DMF solution. Take copper chloride (50 mg) and dissolve it in 10 mL of DMF solution. After mixing and stirring the two, raise the temperature to 10 °C and continue stirring for 1 hour. After cooling to room temperature, let it stand and separate the lower layer precipitate. Centrifuge the upper layer solution to obtain a precipitate. Wash it three times with DMF and ethanol solutions respectively and then centrifuge. Vacuum dry to obtain 80 mg of brick-red solid BPCu@COF-Cu.

[0055] Example 3: Preparation of BPGa@COF-Cu

[0056] In this example, the material preparation process is carried out under nitrogen protection.

[0057] (1) Disperse black phosphorus zinc (10 mg) in 1 mL of tetrahydrofuran solution. Dissolve 5,10,15,20 - tetra(4 - aminophenyl)porphyrin (0.15 mmol, 101 mg) in 5 mL of tetrahydrofuran solution. Dropwise add triethylamine (20 μL). After stirring well at 25 °C for 10 minutes, add 5 mL of a tetrahydrofuran solution of terephthaloyl chloride (0.3 mmol, 60 mg). Stir at 25 °C for 24 hours at room temperature, then let it stand and separate to remove the precipitate to obtain a suspension. Centrifuge the suspension to obtain a precipitate. Wash the precipitate with tetrahydrofuran and then centrifuge again. Dry it under vacuum to obtain 100 mg of red solid black phosphorus zinc porphyrin. (2) Disperse black phosphorus zinc porphyrin (50 mg) in 10 mL of DMF solution. Dissolve copper chloride (500 mg) in 10 mL of DMF solution. Mix the two and stir. Then heat to 150 °C and continue stirring for 20 hours. After cooling to room temperature, let it stand and separate the lower - layer precipitate. Centrifuge the upper - layer solution to obtain a precipitate. Wash the precipitate three times with DMF and ethanol solutions respectively and then centrifuge. Dry it under vacuum to obtain 86 mg of brick - red solid BPGa@COF - Cu.

Claims

1. A nanospherical porous copper porphyrin, characterized in that, The mass percentage contents of gallium and copper in the nano-spherical porous copper porphyrin are 0.7% and 4.2% respectively; the nano-spherical porous copper porphyrin material contains gallium black phosphorus and copper porphyrin, denoted as BPGa@COF-Cu; The constituent elements of the nano-spherical porous copper porphyrin include C, N, Ga, O, P, and Cu; infrared spectroscopy and XPS spectra show the presence of P-Ga bonds, amide bonds, and N-Cu bonds; BPGa@COF-Cu is a spherical nanoparticle with a specific surface area of 17.40 m 2 / g, an average pore volume of 0.05 m 3 / g, an average pore diameter of 18.36 nm, and an average particle size of the nanoparticles of 344.8 nm; Preparation method of BPGa@COF-Cu: First, black phosphorus crystals are obtained by grinding bulk black phosphorus; gallium black phosphorus is prepared in one step by liquid-phase exfoliation ultrasound, where gallium and black phosphorus are connected by P-Ga bonds; subsequently, using gallium black phosphorus as a template, 5,10,15,20-tetra(4-aminophenyl)porphyrin and terephthaloyl chloride are induced to form amide bonds and wrap gallium black phosphorus, and finally, porous spherical BPGa@COF-Cu is obtained by coordinating and assembling the N atoms of porphyrin with copper ions, where copper ions and porphyrin are connected by N-Cu bonds.

2. The preparation method of the nano-spherical porous copper porphyrin according to claim 1, characterized in that It includes the following steps: (1) Disperse gallium black phosphorus compound, 5,10,15,20-tetra(4-aminophenyl)porphyrin, and terephthaloyl chloride in tetrahydrofuran solution respectively. Slowly drip the gallium black phosphorus tetrahydrofuran solution into the 5,10,15,20-tetra(4-aminophenyl)porphyrin tetrahydrofuran solution, then add the terephthaloyl chloride tetrahydrofuran solution, and finally add triethylamine. Mix evenly and stir to react. After the reaction is completed, centrifuge, wash with tetrahydrofuran, and dry in vacuum to obtain gallium black phosphorus porphyrin compound; (2) Disperse gallium black phosphorus porphyrin compound and copper chloride in N,N-dimethylformamide (DMF) solution respectively. Drip the copper chloride DMF solution into the gallium black phosphorus porphyrin DMF solution, stir to react. After the reaction is completed, centrifuge, wash with DMF, and dry in vacuum to obtain gallium black phosphorus porphyrin copper compound.

3. The preparation method according to claim 2, characterized in that, In step (1), the dosage ratio of gallium black phosphorus, 5,10,15,20-tetra(4-aminophenyl)porphyrin, terephthaloyl chloride, and triethylamine is 1 - 10 mg: 10 - 200 mg: 10 - 100 mg: 10 - 50 μL, the reaction temperature is 10 - 50 °C, and the reaction time is 1 - 24 hours.

4. The preparation method according to claim 3, wherein The dosage ratio of gallium black phosphorus, 5,10,15,20-tetra(4-aminophenyl)porphyrin, terephthaloyl chloride, and triethylamine is 5 mg: 101 mg: 60 mg: 20 μL, the reaction temperature is 25 °C, and the reaction time is 12 hours.

5. The preparation method according to claim 2, wherein, In step (2), the mass ratio of gallium black phosphorus porphyrin and copper chloride is 5:1 - 1:10, the reaction temperature is 10 - 150 °C, and the reaction time is 1 - 20 hours.

6. The preparation method according to claim 5, characterized in that, The mass ratio of gallium black phosphorus porphyrin and copper chloride is 1:5, the reaction temperature is 100 °C, and the reaction time is 10 hours.

7. Application of the nano-spherical porous copper porphyrin prepared by the method according to claim 1 or any one of claims 2 - 6 in inhibiting Escherichia coli or Staphylococcus aureus in the field for non-disease diagnosis and treatment purposes.

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