A fluorinated porphyrin-based covalent organic framework, its preparation method and application

Abstract

The present invention discloses a fluorinated porphyrin-based covalent organic framework, a preparation method thereof, and an application thereof. The fluorinated porphyrin-based covalent organic framework has the following structure: the fluorinated porphyrin-based covalent organic framework of the present invention is prepared by a Schiff base reaction in which an amino group and an aldehyde group are condensed to remove a molecule of water to obtain a C=N double bond (imine bond). The framework is not easily hydrolyzed and has excellent biocompatibility and stable chemical properties. The framework significantly improves photosensitivity and can efficiently load oxygen, thereby promoting photodynamic therapy under light response and improving its tumor treatment effect.

Description

Technical Field

[0001] The invention relates to a fluorine-containing porphyrin-based covalent organic framework, a preparation method and application thereof, and belongs to the technical field of polymer materials and biomaterials. Background Art

[0002] In recent years, cancer has become a major health concern with a steadily increasing incidence worldwide, posing a serious threat to human life. Hypoxia is a key characteristic of the tumor microenvironment. Hypoxia is widespread in tumor cells due to an imbalance between insufficient oxygen supply and abnormal oxygen consumption. Tumor hypoxia can lead to decreased oxygen tension in tumor tissue, abnormal tumor cell proliferation, and angiogenesis dysfunction. Severe tumor hypoxia can compromise the efficacy of cancer treatments, making addressing this issue a crucial step in combating cancer.

[0003] In this context, nanomedicine has attracted extensive attention as an emerging therapeutic method. With its unique nanoscale effect and excellent biocompatibility, nanomedicine provides new possibilities for tumor treatment.

[0004] Covalent Organic Frameworks (COFs) are a type of porous crystalline material formed by organic molecules connected by covalent bonds. Since the COFs framework structure is formed by organic building units connected by strong covalent bonds, it is not easily attacked by chemical reagents and is not easily dissociated by high temperatures, and has high thermal stability. The advantages of COFs lie in their unique structural characteristics and rich functionality. They not only have a high specific surface area and porosity, and can efficiently load drug molecules, but also can achieve directional release and targeted delivery of drug molecules through precise design and control. However, existing COFs are prone to hydrolysis, resulting in a limited duration of drug efficacy. Summary of the Invention

[0005] The present invention provides a fluorinated porphyrin-based covalent organic framework, a preparation method, and its application. The fluorinated porphyrin-based covalent organic framework of the present invention can effectively alleviate tumor hypoxia and improve its tumor treatment effect under light response. During the preparation of the present invention, a solvothermal method is used to condense an amino group with an aldehyde group to remove a molecule of water to form a Schiff base reaction of a C=N double bond (imine bond). This generates a fluorinated porphyrin-based covalent organic framework with an imine-linked Schiff base-like COF structure with excellent biocompatibility and stable chemical properties. The structure has good regularity and is not easily hydrolyzed.

[0006] In order to solve the above technical problems, the technical solutions adopted by the present invention are as follows:

[0007] A fluorinated porphyrin-based covalent organic framework (COFs) having the structure:

[0008]

[0009] The inventors characterized the structure and micromorphology of the fluorinated porphyrin-based covalent organic framework using infrared spectroscopy, ultraviolet spectroscopy, scanning electron microscopy (SEM), and dynamic light scattering, confirming the stability of the fluorinated porphyrin-based covalent organic framework. Oxygen-carrying capacity testing of the material revealed excellent oxygen-carrying properties, and photodynamic therapy testing at the cellular level demonstrated its potential for application in the treatment of tumor hypoxia.

[0010] The fluorinated porphyrin-based covalent organic framework has high photosensitivity and high oxygen-carrying capacity, and generates active oxygen after irradiation with a 660nm laser.

[0011] The preparation method of the fluorine-containing porphyrin-based covalent organic framework is prepared by reacting tetraaldehyde phenylporphyrin and 2,5-bis((3,3,4,4,5,5,6,6,6-nonafluorohexyl)oxy)terephthaloylhydrazide.

[0012] The tetraaldehyde phenylporphyrin and 2,5-bis((3,3,4,4,5,5,6,6,6-nonafluorohexyl)oxy)terephthaloylhydrazide used in the present application are both biocompatible materials.

[0013] In order to further improve biocompatibility and stable chemical properties, the molar ratio of tetraaldehyde phenylporphyrin to 2,5-bis((3,3,4,4,5,5,6,6,6-nonafluorohexyl)oxy)terephthaloyl dihydrazide is 1:(1.8-2.2), more preferably 1:2.

[0014] The preparation method of the above-mentioned fluorinated porphyrin-based covalent organic framework has the following synthesis path:

[0015]

[0016] As a specific implementation scheme, the preparation method of the above-mentioned fluorinated porphyrin-based covalent organic framework is as follows: tetraaldehyde phenylporphyrin and 2,5-bis((3,3,4,4,5,5,6,6,6-nonafluorohexyl)oxy)terephthaloyl dihydrazide are mixed, 1,4-dioxane, chloroform and acetic acid at a concentration of 6M are added, and ultrasonication is carried out at room temperature for 5 to 10 minutes, and vacuum nitrogen degassing is carried out. Then, the resulting mixture is sealed and reacted at 80-90°C for 2 to 4 days, naturally cooled, washed with tetrahydrofuran, and vacuum dried to obtain the final product.

[0017] The concentration of the acetic acid is 6M (mol / L). The above method significantly improves the yield (synthesis rate).

[0018] The above method uses a solvent thermal method to generate a fluorinated porphyrin-containing covalent organic framework by condensing the amino group with the aldehyde group to remove a molecule of water to form a Schiff base reaction of a C=N double bond (imine bond). The yield is greater than 90%.

[0019] The above-mentioned fluorinated porphyrin-based covalent organic framework has high photosensitivity and high oxygen-carrying capacity, and generates reactive oxygen species after laser irradiation, thereby promoting photodynamic therapy and improving its tumor treatment effect.

[0020] The 1,4-dioxane and chloroform are mainly used as reaction solvents, and acetic acid (AcOH) is mainly used as a catalyst.

[0021] The vacuum nitrogen degassing is to degas with nitrogen protection so as to carry out the reaction under a nitrogen atmosphere.

[0022] In order to improve the reaction efficiency, the volume ratio of 1,4-dioxane, chloroform and acetic acid is 7:3:1.

[0023] In order to better ensure the formation of the final product, the vacuum drying temperature is 80-90°C and the time is 4-6h.

[0024] The fluorinated porphyrin-based covalent organic framework is used to prepare targeted drugs for killing cancer cells.

[0025] The fluorinated porphyrin-based covalent organic frameworks (COFs) prepared by the above method have fluorinated porphyrin groups, which are beneficial to improving photosensitivity and efficiently loading oxygen, thereby promoting photodynamic therapy under light response and improving its tumor treatment effect.

[0026] The above-mentioned fluorinated porphyrin-based covalent organic framework is preferably used to prepare a targeted drug for killing breast cancer cells.

[0027] The technologies not mentioned in this invention are all referred to the prior art.

[0028] The fluorinated porphyrin-based covalent organic framework of the present invention is prepared by a Schiff base reaction in which an amino group and an aldehyde group are condensed to remove a molecule of water to obtain a C=N double bond (imine bond). It is not easily hydrolyzed, has excellent biocompatibility and stable chemical properties, and significantly improves drug utilization. It also significantly improves photosensitivity and generates reactive oxygen species under infrared light irradiation, thereby improving the efficacy of tumor treatment. It can efficiently load oxygen and deliver oxygen to hypoxic tumor sites, thereby increasing the content of reactive oxygen species in tumor treatment, thereby promoting photodynamic therapy under light response and improving its tumor treatment effect. It has good biocompatibility and stable chemical properties, is simple to prepare, and has a high yield. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 Infrared spectra of the fluorinated porphyrin-based covalent organic frameworks (COFs) of the present invention;

[0030] Figure 2 UV spectra of the fluorinated porphyrin-based covalent organic frameworks (COFs) of the present invention;

[0031] Figure 3 The morphology of the fluorinated porphyrin-based covalent organic framework (COFs) of the present invention is shown in a scanning electron microscope image;

[0032] Figure 4 Transmission electron microscopy images of the fluorinated porphyrin-based covalent organic frameworks (COFs) of the present invention;

[0033] Figure 5 EDS images of the fluorinated porphyrin-based covalent organic frameworks (COFs) of the present invention;

[0034] Figure 6 Particle size distribution of the fluorinated porphyrin-based covalent organic frameworks (COFs) of the present invention;

[0035] Figure 7 Oxygen loading capacity diagram of the fluorinated porphyrin-based covalent organic frameworks (COFs) of the present invention;

[0036] Figure 8 Cell viability graph of the fluorinated porphyrin-based covalent organic frameworks (COFs) of the present invention;

[0037] Figure 9 The active oxygen generation diagram of the fluorinated porphyrin-based covalent organic frameworks (COFs) of the present invention;

[0038] Figure 10 The effect diagram of the fluorinated porphyrin-based covalent organic framework loaded with oxygen for tumor treatment under 660nm laser irradiation;

[0039] Figure 11 The fluorine-containing porphyrin-based covalent organic framework of the present invention generates reactive oxygen species (ROS) after laser irradiation. DETAILED DESCRIPTION

[0040] In order to better understand the present invention, the content of the present invention is further illustrated below in conjunction with the examples, but the content of the present invention is not limited to the following examples.

[0041] In each case, the room temperature was 15-25° C. The stirring speed, unless otherwise specified, was 60 rpm.

[0042] Example 1

[0043] Preparation of Fluorinated Porphyrin-Based Covalent Organic Frameworks:

[0044] 14.53 mg (0.02 mmol) of tetraaldehyde phenylporphyrin (TFPP) and 28.7 mg (0.04 mmol) of 2,5-bis((3,3,4,4,5,5,6,6,6-nonafluorohexyl)oxy)terephthaloyl dihydrazide (TH-F9) were weighed and added to a Pyrex tube. 0.7 ml of 1,4-dioxane and 0.3 ml of chloroform were added as reaction solvents. 0.1 ml of 6 M acetic acid (AcOH) was then added to the solution as a catalyst. The tube was sealed with adhesive tape, and the solution was placed in an ultrasonic cleaner for ultrasonic treatment (200 W) for 30 minutes to uniformly distribute the monomers in the solvent. After ultrasonication, a purple suspension was obtained. After freezing with liquid nitrogen for 3 minutes, the sealing film was removed and the Pyrex tube was connected to the vacuum tube. After three cycles of liquid nitrogen freezing-vacuuming-nitrogen filling to remove air from the reaction system, the system was vacuumed again for 3-5 minutes. Finally, the tube mouth was sealed under vacuum. The Pyrex tube was sealed with an 80°C flame. After the temperature rose to room temperature, the tube was placed in a constant temperature oven at 80-90°C for reaction for 96 hours.

[0045] After the reaction is completed, cool to room temperature, break the Pyrex tube with tweezers in a fume hood, transfer the sample inside to a 10 ml container bottle with a spoon, add 10 ml of tetrahydrofuran to soak, and replace the tetrahydrofuran every 24 hours until the supernatant liquid is clear to obtain a soaking solution mixed with the product.

[0046] Install the sand core funnel, connect the suction pump, place a piece of filter paper on the sand core funnel, press it with the funnel bottle, turn on the suction pump, and use a pipette to draw 1ml of tetrahydrofuran to soak the filter paper. Shake the soaking solution mixed with the product to distribute the product in the solution, then pour it into the sand core funnel for suction filtration. Use tetrahydrofuran to promptly rinse off the product stuck to the funnel wall. Filter for 6 minutes. After completion, use a spoon to scrape the product off the filter paper and transfer it to a 25ml solvent bottle. Close the bottle stopper tightly, plug the side branch with cotton, place the 25ml solvent bottle together with the heating module on the heater, and vacuum dry at 100℃ for 4 hours. After the solvent bottle has cooled, pour it into a mortar and grind it, pick out the glass slag, and grind the sample again until the particle size is less than 100 mesh. Then transfer it to a weighing paper and weigh it, and record the weight to obtain a fluorinated porphyrin-based covalent organic framework (synthesis rate 91%). Finally, transfer it to a 5 ml EP tube (plastic centrifuge tube) and store it in a glass bottle with silica gel desiccant in the dark.

[0047] (1) Infrared and UV spectra of fluorinated porphyrin-based covalent organic frameworks:

[0048] The successful synthesis of the fluorinated porphyrin-based covalent organic framework (TT-COF) was analyzed and verified by Fourier transform infrared absorption spectroscopy. Figure 1The analysis showed that the infrared spectra of tetraaldehyde phenylporphyrin (TFPP), 2,5-bis((3,3,4,4,5,5,6,6,6-nonafluorohexyl)oxy) terephthaloylhydrazide (TH-F9) and TT-COF were compared. The infrared spectrum of TT-COF showed a peak at 3400 cm -1 The NH stretching vibration absorption peak of the amino group disappears, and the C=O stretching vibration signal of the aldehyde group is at 1700cm -1 The peak at 1676cm is also significantly weakened. -1 The characteristic stretching vibration peak of C=N bond appeared at . The results showed that tetraaldehyde phenylporphyrin (TFPP) reacted with the aldehyde group and amino group in 2,5-bis((3,3,4,4,5,5,6,6,6-nonafluorohexyl)oxy) terephthaloyl dihydrazide (TH-F9) to generate a product connected by imine bond, namely porphyrin-based covalent organic framework (TT-COF) was successfully synthesized. The UV spectrum was measured as follows Figure 2 , which are the UV spectra of TH-F9, TFPP and TT-COF respectively.

[0049] (II) Morphological characterization of fluorinated porphyrin-based covalent organic frameworks:

[0050] 1 mg of the product was dissolved in 1 ml of deionized water, sonicated to dissolve it, and a drop was placed on an organic copper mesh. After the water evaporated, the morphology was observed using a scanning electron microscope. Figure 3 The morphology of the fluorinated porphyrin-based covalent organic framework is shown in the scanning electron microscope image. Figure 3 It can be seen that the size of the fluorinated porphyrin-based covalent organic framework is about 100 nm and is spherical under the microscopic view. Figure 4 Transmission electron microscopy (TEM) image of the nanoparticles shows clear lattice fringes, further demonstrating the synthesis of TT-COF materials. Figure 5 This is the EDS image of nanoparticles, from which we can see the distribution of C, O, N, and F in the material.

[0051] (III) Particle size distribution of fluorinated porphyrin-based covalent organic frameworks:

[0052] 1 mg of the product was dissolved in 1 ml of deionized water and sonicated to dissolve it. 1 ml of the supernatant was placed in a four-sided light-transmitting cuvette and its particle size distribution was measured using a Malvern laser particle size analyzer. Figure 6 is the particle size distribution of the fluorinated porphyrin-based covalent organic framework, Figure 6 It can be seen that the particle size distribution of the fluorinated porphyrin-based covalent organic framework is consistent with the scanning electron microscopy observation.

[0053] (IV) Stability evaluation test of fluorinated porphyrin-based covalent organic frameworks:

[0054] 1 mg of the product was dissolved in 1 ml of deionized water and sonicated to dissolve it. 1 ml of the supernatant was placed in a four-sided light-transmitting cuvette and its particle size distribution was measured using a Malvern laser particle size analyzer for six consecutive days. The particle size of the fluorinated porphyrin-based covalent organic framework remained relatively stable, proving that it is not easily hydrolyzed and has stable properties. Figure 7 Stability evaluation of covalent organic frameworks containing fluorinated porphyrin groups.

[0055] (V) Oxygen-carrying test of fluorinated porphyrin-based covalent organic frameworks:

[0056] 10 mg of the fluorinated porphyrin-based covalent organic framework prepared in Example 1 was dissolved in 10 ml of water. Nitrogen was then introduced for 15 minutes to remove the pre-dissolved gas in the water. Oxygen was then introduced for 15 minutes to allow the material to be fully loaded with oxygen. The oxygen concentration was measured using a portable dissolved oxygen meter. The amount of dissolved oxygen far exceeded that in the control group. Figure 8 is the oxygen loading capacity of the fluorinated porphyrin-based covalent organic framework.

[0057] Example 2

[0058] Cell viability:

[0059] The fluorinated porphyrin-based covalent organic framework obtained in Example 1 was prepared into a 10 mg / mL stock solution with PBS (phosphate buffered saline), and diluted to 1000 μg / mL and 100 μg / mL before use.

[0060] Experimental steps: Primary cultured 4T1 cells (provided by Hebei University of Technology) were cultured at 5×10 3 / mL was inoculated into a 96-well plate with a volume of 90μL per well and cultured in a CO2 incubator for 24h. After the cells adhered to the wall, 10μL of PBS solution of fluoroporphyrin-containing covalent organic framework was added to form different concentrations of fluoroporphyrin-containing covalent organic framework TT-COF. After 48h of culture, 10μL of MTT (3-(4,5-dimethylthiazol-2)-2,5-diphenyltetrazolium bromide) solution (5mg / mL) was added to each well and continued to incubate in the incubator for 4h. After pouring off the supernatant, 100μL of DMSO (dimethyl sulfoxide) was added to each well. After shaking, the absorbance of each well was measured at 570nm using a microplate reader. At the same time, a blank group (same volume of PBS) was set up to calculate the cell viability (Cell viability) = (OD value of the extract group / OD value of the blank group) × 100%. The results are shown as follows. Figure 9 Without 660nm laser irradiation, the cell survival rate exceeded 98% after incubation with different concentrations of TT-COF for 24h, confirming that TT-COF has good biocompatibility. In contrast, 4T1 cells treated with TT-COF irradiated with 660nm laser showed obvious cytotoxicity. Figure 9It can be seen that TT-COF itself is non-toxic to cells, but becomes cytotoxic after laser irradiation and can be used for targeted drug therapy; Figure 9 In the figure, TT-COF indicates no laser irradiation, and TT-COF+L indicates laser irradiation. At the same concentration in the figure, the left side is TT-COF and the right side is TT-COF+L. At the same time, the fluoroporphyrin-based covalent organic framework TT-COF was loaded with oxygen (nitrogen was introduced and stirred for 15 minutes to remove the gas pre-dissolved in water, and then oxygen was introduced and stirred for 15 minutes to fully load the material with oxygen). The above operation was carried out in two groups: one with 660nm laser irradiation and the other without 660nm laser irradiation. Figure 10 It can be seen that the fluorinated porphyrin-based covalent organic framework TT-COF loaded with oxygen has a better effect in tumor treatment under 660nm laser irradiation. Figure 11 The fluorinated porphyrin-based covalent organic framework generates reactive oxygen species (ROS) after laser irradiation, which can effectively kill cancer cells.

Claims

1. A photosensitive, highly oxygen-loaded, fluorinated porphyrin-based covalent organic framework, characterized by: Its structure is: 。 2. A method for preparing the fluorinated porphyrin-based covalent organic framework according to claim 1, characterized in that: It is prepared by the reaction of tetraaldehyde phenylporphyrin and 2,5-bis((3,3,4,4,5,5,6,6,6-nonafluorohexyl)oxy)terephthaloylhydrazide.

3. The method for preparing a fluorinated porphyrin-based covalent organic framework according to claim 2, wherein: The molar ratio of tetraaldehyde phenylporphyrin to 2,5-bis((3,3,4,4,5,5,6,6,6-nonafluorohexyl)oxy)terephthaloyl dihydrazide is 1:(1.8~2.2).

4. The method for preparing a fluorinated porphyrin-based covalent organic framework according to claim 2 or 3, wherein: The synthesis path is: 。 5. The method for preparing a fluorinated porphyrin-based covalent organic framework according to claim 2 or 3, wherein: Tetraaldehyde phenylporphyrin and 2,5-bis((3,3,4,4,5,5,6,6,6-nonafluorohexyl)oxy)terephthaloylhydrazide were mixed, 1,4-dioxane, chloroform and acetic acid were added, and ultrasonic treatment was performed at room temperature for 5-10 minutes. The mixture was degassed with nitrogen in a vacuum. Then, the resulting mixture was sealed and reacted at 80-90°C for 2-4 days. The mixture was naturally cooled, washed with tetrahydrofuran, and vacuum dried to obtain a fluorinated porphyrin-based covalent organic framework.

6. The method for preparing a fluorinated porphyrin-based covalent organic framework according to claim 2 or 3, wherein: The volume ratio of 1,4-dioxane, chloroform and acetic acid is 7:3:

1.

7. The method for preparing a fluorinated porphyrin-based covalent organic framework according to claim 2 or 3, wherein: The vacuum drying temperature is 80~90℃ and the time is 4-6h.

8. A use of the fluorinated porphyrin-based covalent organic framework according to claim 1, characterized in that: Used to prepare targeted drugs to kill cancer cells.

9. The use of the fluorinated porphyrin-based covalent organic framework according to claim 8, characterized in that: Used to carry oxygen.

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