Method for the preparation of a porphyrin-anthracene covalent organic framework for sustainable phototherapy
The porphyrin-based covalent organic framework material Por-DPA was synthesized via a Schiff base reaction solvothermal method, which solved the problems of photodamage and photosensitizer self-aggregation quenching in traditional photodynamic therapy, achieving continuous photothermal and photodynamic sterilization, and improving the therapeutic effect and safety.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- JIANGNAN UNIV
- Filing Date
- 2023-10-19
- Publication Date
- 2026-07-03
AI Technical Summary
Traditional photodynamic therapy requires continuous irradiation, which is easily affected by the duration and intensity of light exposure, and may cause damage to normal tissues. Furthermore, photosensitizers are prone to self-aggregation and quenching, resulting in poor photostability.
Por-DPA, a porphyrin-based covalent organic framework material, was synthesized using a Schiff base reaction solvothermal method. It achieves photothermal and photodynamic sterilization without continuous irradiation, and utilizes its porous structure to store oxygen and release singlet oxygen in the dark.
It achieves continuous sterilization without continuous light exposure, avoids light damage, improves photothermal and photodynamic effects, and ensures the stability and sterilization efficiency of the photosensitizer.
Smart Images
Figure CN117487158B_ABST
Abstract
Description
Technical Field
[0001] This invention relates to a method for preparing a porphyrin-anthracene covalent organic framework for sustainable phototherapy, belonging to the fields of advanced functional materials and biomedical technology. Background Technology
[0002] Bacterial infection is an acute systemic infection caused by pathogenic or opportunistic pathogens entering the bloodstream through wounds or infected lesions, multiplying, and producing toxins and other metabolic products. Bacterial infections seriously affect human health and social development. With the advent of antibiotics, this problem has been effectively alleviated. However, due to the overuse of antibiotics, bacteria have developed resistance, reducing treatment effectiveness. Therefore, developing novel therapies with low biotoxicity and without inducing drug resistance is crucial.
[0003] Photosterilization, including photothermal and photodynamic sterilization, has attracted much attention due to its advantages such as low biotoxicity, low side effects, and low likelihood of inducing drug resistance. However, traditional photosterilization relies on the duration and intensity of external light source irradiation, requiring exposure to an external light source to exert its bactericidal effect, which inevitably leads to photodamage to normal tissues. The singlet oxygen content at the infection site is limited, and traditional photodynamic sterilization methods are limited by oxygen deficiency. Furthermore, traditional photosensitizers exhibit self-aggregation and quenching, resulting in poor photostability and unsatisfactory therapeutic effects. Therefore, there is an urgent need to develop a material that does not aggregate and self-quench, enabling continuous photodynamic and photothermal sterilization for the treatment of bacterial infections. Summary of the Invention
[0004] Technical issues:
[0005] Traditional photodynamic therapy mostly requires continuous irradiation, and the therapeutic effect is affected by the duration and intensity of light exposure. It is difficult to avoid photodamage to normal tissues and is also limited by hypoxia. In addition, traditional photosensitizers are prone to self-aggregation quenching and have poor photostability.
[0006] Technical solution:
[0007] To address the above problems, the present invention aims to provide Por-DPA, synthesized by a Schiff base reaction solvothermal method, which can provide sustainable photodynamic / photothermal sterilization without continuous irradiation. It overcomes and solves the problems of unsatisfactory therapeutic effects caused by the easy aggregation and quenching of traditional photosensitizers under physiological conditions and photodamage caused by continuous irradiation.
[0008] The technical solution of this invention:
[0009] The first objective of this invention is to provide a method for preparing porphyrin-based covalent organic framework materials with sustained photothermal / photodynamic sterilization, comprising the following steps:
[0010] (1) 5,10,15,20-tetrakis(4-aminophenyl)porphyrin and 9,10-bis(4-formylphenyl)anthracene (DPA) were dispersed in a solvent, mixed at room temperature, and then the catalyst was added and mixed evenly to obtain a reaction solution;
[0011] (2) React the reaction solution at 60-150℃ for 1-4 days;
[0012] (3) After the reaction is complete, the material is centrifuged, washed, and dried to obtain the porphyrin-based covalent organic framework material, denoted as Por-DPA.
[0013] In one embodiment of the present invention, the molar ratio of 5,10,15,20-tetra-(4-aminophenyl)porphyrin and 9,10-bis(4-formylphenyl)anthracene in step (1) is 1:2.
[0014] In one embodiment of the present invention, the solvent in step (1) includes one or more of mesitylene, n-butanol, dioxane, o-dichlorobenzene, and ethanol; more preferably, a mixture of mesitylene and n-butanol.
[0015] In one embodiment of the present invention, the volume ratio of mesitylene and n-butanol is (10:0):(5:5); preferably 9:1.
[0016] In one embodiment of the present invention, the ratio of the solvent and 5,10,15,20-tetra-(4-aminophenyl)porphyrin in step (1) is 2 mL: (60-80) mg; specifically, 2 mL: 67.5 mg.
[0017] In one embodiment of the present invention, the catalyst in step (1) is an acetic acid solution with a concentration of 6M.
[0018] In one embodiment of the present invention, the amount of catalyst in step (1) is 10% (V / V) of the entire reaction system.
[0019] In one embodiment of the present invention, the mixing in step (1) is ultrasonic mixing, and the ultrasonic time is 5-60 min.
[0020] In one embodiment of the present invention, the reaction solution described in step (2) needs to undergo three cycles of freezing-nitrogen purging-vacuuming-thawing before the reaction.
[0021] In one embodiment of the present invention, the centrifugation speed in step (3) is 8000-12000 rpm, and the precipitate is washed 3-6 times; the washing solvent is one or more of tetrahydrofuran and N,N-dimethylformamide.
[0022] In one embodiment of the present invention, the drying in step (3) is vacuum drying.
[0023] The second objective of this invention is to prepare porphyrin-based covalent organic framework materials using the method described herein.
[0024] In one embodiment of the present invention, the structural unit of the porphyrin-based covalent organic framework material has the structural formula as shown in Formula I:
[0025]
[0026] Formula I.
[0027] In one embodiment of the present invention, a porphyrin-based photosensitizer, as a monomer, exhibits photothermal and photodynamic activity under irradiation; anthracene derivatives, as another monomer, can capture singlet oxygen and continuously release singlet oxygen in the dark, thereby achieving a sustainable phototherapy effect; the ordered crystal structure of the covalent organic framework effectively avoids the self-aggregation quenching of the photosensitizer, and the porous structure is conducive to the storage and transport of oxygen and the generated singlet oxygen; thus, the constructed sustainable photothermal / photodynamic bactericidal material has good photothermal effect, superior ability to generate active oxygen, and continuous bactericidal performance, and has great potential in the treatment of bacterial infections.
[0028] A third objective of this invention is to provide the application of the above-mentioned porphyrin-based covalent organic framework material in the preparation of sustained photothermal / photodynamic bactericidal agents.
[0029] The fourth objective of this invention is to provide a photothermal / photodynamic sterilization method for non-disease diagnosis and treatment, wherein the method uses the porphyrin-based covalent organic framework material (Por-DPA) described in this invention as the sterilization material.
[0030] In one embodiment of the present invention, the photothermal / photodynamic sterilization method for non-disease diagnosis and treatment involves adding a porphyrin-based covalent organic framework material (Por-DPA) and then irradiating it with light.
[0031] The specific steps include: dissolving porphyrin-based covalent organic framework material (Por-DPA) in a solvent, then mixing it with bacterial suspension, irradiating it under a suitable light source for 30 minutes, then serially diluting it and coating it onto a solid culture medium, and counting the colonies after culturing for 12-24 hours to obtain the sterilization rate; wherein the solvent is a liquid culture medium, the bacterial suspension includes Staphylococcus aureus and Escherichia coli, and the light source is a white LED lamp.
[0032] The beneficial effects of this invention are:
[0033] (1) The porphyrin-based covalent organic framework material (Por-DPA) prepared in this invention has a special porous structure that allows it to store oxygen under light and release reactive oxygen species in the dark, thereby achieving continuous phototherapy without the need for continuous irradiation. Through the dual pathways of photosensitization and cyclization reversal, it exhibits excellent photothermal conversion performance and high efficiency. 1 O2 generation capability.
[0034] (2) The pore structure of the porphyrin-based covalent organic framework material (Por-DPA) prepared by the present invention restricts the distance between photosensitizer monomer molecules, thereby reducing the self-aggregation quenching of photosensitizers, ensuring high-density aggregation of porphyrin molecules, and achieving excellent photothermal and photodynamic effects. It effectively prevents the self-aggregation quenching of photosensitizers and thermal damage caused by continuous exposure to external light sources.
[0035] (3) The material is metal-free in the synthesis process, which provides the possibility for the subsequent degradation of the material. These characteristics give Por-DPA good biosafety. Attached Figure Description
[0036] Figure 1 This is a schematic diagram of the Por-DPA prepared in Example 1, which features continuous photothermal / photodynamic sterilization.
[0037] Figure 2 Experimental and simulated X-ray powder diffraction patterns of Por-DPA prepared in Example 1.
[0038] Figure 3 The image shows a scanning electron microscope (SEM) image of the Por-DPA prepared in Example 1.
[0039] Figure 4 The image shows the X-ray powder diffraction pattern of Por-DPA prepared in Example 2.
[0040] Figure 5 The image shows the X-ray powder diffraction pattern of Por-DPA prepared in Example 3.
[0041] Figure 6 The image shows the X-ray powder diffraction pattern of Por-DPA prepared in Example 4.
[0042] Figure 7 The image shows the X-ray powder diffraction pattern of Por-DPA prepared in Example 5.
[0043] Figure 8 The photothermal image of Por-DPA prepared in Example 1.
[0044] Figure 9The graph shows the ability of Por-DPA prepared in Example 1 to generate singlet oxygen, where A0 and A are the UV absorption intensities of the ABDA solution at 377 nm before and after illumination, respectively.
[0045] Figure 10 The graph shows the ability of Por-DPA prepared in Example 1 to release singlet oxygen in the dark, where A0 and A are the UV absorption intensities of the DPBF solution at 410 nm before and after light irradiation, respectively.
[0046] Figure 11 This is a plate sterilization diagram of Escherichia coli by Por-DPA prepared in Example 1.
[0047] Figure 12 The bar charts show the bactericidal activity of Por-DPA prepared in Example 1 against Escherichia coli at concentrations of (a) 200 μg / mL and (b) 400 μg / mL. Detailed Implementation
[0048] The preferred embodiments of the present invention are described below. It should be understood that the embodiments are for better explanation of the present invention and are not intended to limit the present invention.
[0049] Example 1:
[0050] Preparation of a Por-DPA material with sustained photothermal / photodynamic sterilization: 5,10,15,20-tetrakis(4-aminophenyl)porphyrin is confined as a structural unit within a covalent organic framework. This reduces the self-aggregation quenching effect while maintaining the photosensitizer density, thus improving its photothermal and photodynamic effects and providing a framework for storing singlet oxygen. The preparation includes the following steps:
[0051] (1) 67.5 mg of 5,10,15,20-tetra(4-aminophenyl)porphyrin (TAPP, 0.10 mmol) and 77.3 mg of 9,10-bis(4-formylphenyl)anthracene (DPA, 0.20 mmol) were added to a solvent consisting of tricresyl / n-butanol (9:1, v:v, 2 mL), and sonicated for 5 minutes. Then, 1.2 mmol of acetic acid was added, and the mixture was sonicated for another 5 minutes to obtain the reaction solution.
[0052] (2) The reaction solution was subjected to a three-stage freezing-nitrogen-vacuuming-thawing cycle, and then reacted in an oil bath at 120 °C for 72 hours.
[0053] (3) After the reaction is complete, use N,NThe mixture was washed three times with dimethylformamide, then washed three times with tetrahydrofuran, and finally dried under vacuum to obtain a purplish-black powder, Por-DPA.
[0054] The prepared Por-DPA was subjected to performance testing. Figure 2 The results show that the characteristic diffraction peaks of the porphyrin-based covalent organic framework material (Por-DPA) are similar to those of the simulated AA stacking diffraction peaks, but differ significantly from those of the simulated AB stacking diffraction peaks, indicating that the obtained porphyrin-based covalent organic framework is constructed using the AA stacking mode.
[0055] Example 2:
[0056] The mixed solvents of mesitylene / n-butanol in Example 1 were adjusted to mesitylene / n-butanol (5:5, v:v), mesitylene / dioxane (5:5, v:v), o-dichlorobenzene / ethanol (5:5, v:v), and o-dichlorobenzene / n-butanol (5:5, v:v), while other aspects remained the same as in Example 1, to obtain the porphyrin-based covalent organic framework material (Por-DPA).
[0057] The obtained porphyrin-based covalent organic framework material (Por-DPA) was subjected to performance testing, and the test results are as follows:
[0058] The X-ray powder diffraction pattern of the obtained material is as follows: Figure 4 As shown, Por-DPA obtained with tricresyl / dioxane, o-dichlorobenzene / ethanol, or o-dichlorobenzene / n-butanol as the reaction system has a lower characteristic diffraction peak at 3.5°, indicating that its crystallinity is lower than that of Por-DPA with tricresyl / n-butanol as the reaction system.
[0059] High crystallinity facilitates the transfer and transport of singlet oxygen, effectively preventing the self-aggregation and quenching of photosensitizers. Low crystallinity results in a lack of porous structure in COFs, leading to poor stability and an inability to effectively limit monomer self-aggregation. It also hinders the storage and transport of generated singlet oxygen.
[0060] Example 3:
[0061] The ratio of the trimethylbenzene / n-butanol mixed solvent in Example 1 was adjusted to (10:0), (9:1), (7:3), (5:5), (3:7), (1:9) (v:v), while other aspects remained the same as in Example 1, to obtain the porphyrin-based covalent organic framework material (Por-DPA).
[0062] The obtained porphyrin-based covalent organic framework material (Por-DPA) was subjected to performance testing, and the test results are as follows:
[0063] X-ray powder diffraction of the obtained material is as follows Figure 5As shown in the figure, the characteristic diffraction peaks of Por-DPA obtained with reaction systems of (10:0), (7:3), (5:5), (3:7), and (1:9) are lower at 3.5°, indicating that their crystallinity is lower than that of Por-DPA with reaction system of mesitylene / n-butanol (9:1, v:v).
[0064] Example 4:
[0065] By adjusting the reaction temperature in Example 1 to 60℃, 90℃, or 150℃, while keeping other parameters consistent with Example 1, a porphyrin-based covalent organic framework material (Por-DPA) was obtained.
[0066] The obtained porphyrin-based covalent organic framework material (Por-DPA) was subjected to performance testing, and the test results are as follows:
[0067] The X-ray powder diffraction pattern of the obtained material is as follows: Figure 6 As shown, Por-DPA obtained by adjusting the reaction temperature to 60℃, 90℃ or 150℃ has a lower characteristic diffraction peak at 3.5°, indicating that its crystallinity is lower than that of Por-DPA obtained at a reaction temperature of 120℃.
[0068] Example 5:
[0069] The reaction time in Example 1 was adjusted to 1, 2, and 4 days, while other aspects remained the same as in Example 1, resulting in a porphyrin-based covalent organic framework material (Por-DPA).
[0070] The obtained porphyrin-based covalent organic framework material (Por-DPA) was subjected to performance testing, and the test results are as follows:
[0071] The X-ray powder diffraction pattern of the obtained material is as follows: Figure 7 As shown, the characteristic diffraction peak at 3.5° of Por-DPA obtained by adjusting the reaction time to 1, 2, and 4 days is lower, indicating that its crystallinity is lower than that of Por-DPA obtained by adjusting the reaction time to 3 days.
[0072] Example 6:
[0073] The performance of the Por-DPA prepared in Example 1 was tested, mainly focusing on its photothermal and photodynamic properties. The specific steps are as follows:
[0074] (1) First, the photothermal properties of Por-DPA were studied: Por-DPA was dispersed in ultrapure water to obtain Por-DPA solutions of different concentrations (0 µg mL). 1 100 µg mL 1 200 µg mL 1 and 400 µg mL 1 Then, 1 mL of the solution was added to a 48-well cell culture plate, and the plate was then illuminated with a white LED light for 30 min (200 µg / mL). 1 TAPP was used as a control, with a power density of 100 mW / cm². -2 Temperature changes during the irradiation process were recorded using a FLIR thermal imager.
[0075] like Figure 8 As shown, at 100 mW·cm -2 After 30 min of white light irradiation, Por-DPA exhibited good photothermal properties, showing a concentration- and time-dependent effect, containing 0 µg mL 1 100 µg mL -1 200 µg mL -1 and 400 µg mL 1 The temperature of Por-DPA solution systems of different concentrations increased by 4℃, 13℃, 16℃ and 17℃, respectively, while the temperature of TAPP, as a control group, increased by only 6℃.
[0076] (2) The photodynamic properties of Por-DPA were then studied: using 1 The photodynamic properties of Por-DPA were investigated using 9,10-anthratrium-bis(methylene)dimalonic acid (ABDA) as an O2 indicator. Specifically, 100 µL of ABDA solution (5 × 10⁻⁶) was added... 3 mol L 1 The solution was mixed with different concentrations of Por-DPA solution (9.9 mL) in 50 mL centrifuge tubes, resulting in solutions with concentrations of 0 µg / mL. 1 100 µg mL 1 200 µg mL 1 and 400 µg mL 1 TAPP (100 µg mL) -1 () was used as a control group. Then, a power density of 100 mW / cm² was used. -2 After irradiating the above solution with a white LED lamp for different times, 1 mL of the irradiated solution was taken out, centrifuged, and the ultraviolet absorption spectrum of the supernatant after centrifugation was measured using an ultraviolet-visible-near-infrared spectrophotometer.
[0077] like Figure 9 As shown, Por-DPA exhibits good photodynamic performance, which is time- and concentration-dependent. After 30 minutes of illumination, 200 µg / mL 1 The absorbance of ABDA treated with Por-DPA decreased to approximately 14% of its initial value, 400 µg mL 1 The absorbance of ABDA treated with Por-DPA was close to zero, indicating the generation of singlet oxygen. In contrast, the absorbance of ABDA alone and ABDA treated with TAPP showed almost no significant change throughout the process. These results demonstrate that the ordered structure of Por-DPA effectively enhances the singlet oxygen production capability of TAPP.
[0078] (3) Then, using DPBF as a probe, the continuous release of Por-DPA after the illumination period was further investigated. 1 The performance of O2. Specifically, the procedure involves adding 600 µL of acetonitrile solution of DPBF (1×10⁻⁶). 4 mol L 1 The solution was mixed with 400 µL of an aqueous solution of Por-DPA, resulting in a final concentration of 200 µg / mL. 1 Then, a power density of 600 mW / cm² was used. -2 After irradiating the above solution with an 808 nm laser for 3 min, it was stored in the dark for different periods of time, centrifuged, and the ultraviolet absorption spectrum of the supernatant after centrifugation was measured using an ultraviolet-visible-near-infrared spectrophotometer.
[0079] like Figure 10 As shown, DPBF remained stable under these conditions for 24 minutes after irradiation ceased, with almost no change in absorbance at 410 nm. However, the absorbance of DPBF mixed with pre-excited Por-DPA decreased to approximately 75.0% of its initial value, confirming that Por-DPA continues to be released in the dark. 1 O2.
[0080] However, the porphyrin covalent organic framework materials reported in CN115232271A and CN113087863A do not exhibit sustained release. 1 O2 is no longer released after irradiation stops. 1 O2 makes continuous phototherapy impossible.
[0081] Example 7:
[0082] The bactericidal ability of Por-DPA prepared in Example 1 was investigated. The specific steps are as follows:
[0083] Plate coating experiment: Using typical Gram-negative Escherichia coli as model bacteria, the bactericidal performance of Por-DPA was evaluated. Bacterial suspensions were mixed with different concentrations of Por-DPA solution (200 µg / mL). 1 400 µg mL 1 Mix and then incubate on a shaking table at 37°C and 200 rpm for 30 minutes. Subsequently, the bacteria in the two models were divided into the following groups: (1) bacteria in unirradiated PBS (No treatment + No irradiation), (2) bacteria in PBS irradiated for 30 minutes (No treatment + irradiation), (3) bacteria treated with Por-DPA in PBS (Por-DPA + No irradiation), (4) bacteria treated with pre-irradiated Por-DPA in PBS (Por-DPA + pre-irradiation), (5) bacteria incubated with Por-DPA in PBS irradiated for 10 minutes, 20 minutes, and 30 minutes respectively (irradiation + Por-DPA-I / Por-DPA-II / Por-DPA-III). White light (100 mW cm⁻¹) was used. 2 Irradiation. Extract 100 μL of the bacterial suspension from each group and serially dilute it 10⁻⁶ times. 4 After dilution, spread the mixture evenly onto solid culture medium. Incubate at 37 ℃ for 12 to 24 hours, then count the number of colonies.
[0084] The results are as follows Figure 11-12 As shown in Table 1, Por-DPA exhibits a significant bactericidal effect under white light irradiation, and the bactericidal effect is positively correlated with concentration and irradiation time. At 100 mW / cm², -2 After 30 minutes of white light irradiation, the bactericidal rate of different concentrations of Por-DPA against Escherichia coli was 100%.
[0085] Table 1
[0086]
Claims
1. A method for preparing porphyrin-based covalent organic framework materials with sustained photothermal / photodynamic sterilization, characterized in that, Includes the following steps: (1) 5,10,15,20-tetra(4-aminophenyl)porphyrin and 9,10-bis(4-formylphenyl)anthracene were dispersed in a solvent, mixed at room temperature, and then a catalyst was added and mixed until homogeneous to obtain a reaction solution; (2) React the reaction solution at 60-150℃ for 1-4 days; (3) After the reaction is complete, the material is centrifuged, washed, and dried to obtain the porphyrin-based covalent organic framework material, denoted as Por-DPA.
2. The method according to claim 1, characterized in that, The molar ratio of 5,10,15,20-tetra-(4-aminophenyl)porphyrin and 9,10-bis(4-formylphenyl)anthracene in step (1) is 1:2; the ratio of solvent to 5,10,15,20-tetra-(4-aminophenyl)porphyrin is 2 mL: (60-80) mg.
3. The method according to claim 1, characterized in that, The solvent mentioned in step (1) includes one or more of the following: mesitylene, n-butanol, dioxane, o-dichlorobenzene, and ethanol.
4. The method according to claim 1, characterized in that, The solvent mentioned in step (1) is a mixture of mesitylene and n-butanol.
5. The method according to claim 4, characterized in that, The volume ratio of mesitylene and n-butanol is (10:0):(5:5).
6. The method according to any one of claims 1-5, characterized in that, The catalyst in step (1) is an acetic acid solution with a concentration of 6M; the amount of catalyst is 10% of the total volume of the reaction system.
7. The porphyrin-based covalent organic framework material with sustained photothermal / photodynamic sterilization prepared by the method according to any one of claims 1-6.
8. The porphyrin-based covalent organic framework material for sustained photothermal / photodynamic sterilization according to claim 7, characterized in that, Its structural unit structure is shown in Equation I: Formula I.
9. The application of the porphyrin-based covalent organic framework material according to claim 7 or 8 in the preparation of sustained photothermal / photodynamic bactericidal agents.
10. A photothermal / photodynamic sterilization method for non-disease diagnosis and treatment, characterized in that, The method uses the porphyrin-based covalent organic framework material as described in claim 7 or 8 as the bactericidal material.