Cationic conjugated polymer as well as preparation method and application thereof

Abstract

The invention provides a cationic conjugated polymer as well as a preparation method and application thereof. The structural formula of the cationic conjugated polymer is shown in the specification. The preparation method comprises the following steps: reacting tris (4-bromophenyl) amine and N, N-dimethyl-4-(4, 4, 5, 5-tetramethyl-1, 3, 2-dioxaborolane-2-yl) aniline in an organic solvent under the action of a catalyst and an initiator to obtain a first intermediate; the cationic conjugated polymer is prepared from the first intermediate, the second intermediate and the third intermediate under the action of a catalyst and an initiator. The cationic conjugated polymer shows good photo-thermal and photodynamic performance under the irradiation of near-infrared light with the wavelength of 980 nm, so that the effect of inhibiting the growth of bacteria is realized. # imgabs0 #

Description

Technical Field

[0001] The invention relates to a cationic conjugated polymer and a preparation method and application thereof, belonging to the technical field of cationic conjugated polymers. Background Art

[0002] In today's era, diseases and safety hazards caused by bacterial infections continue to pose a serious threat to human health. Symptoms such as pain and fever caused by infections greatly damage human well-being, making bacterial infection one of the key health problems that the world urgently needs to overcome. At present, the main means for humans to fight bacterial infections is still highly dependent on the widespread use of antibiotics. Although the large-scale popularization of antibiotics has solved many bacterial diseases to a certain extent, at the same time, it has also caused problems such as a gradual decline in treatment effects and the emergence of antibiotic resistance. Therefore, the development of new antibacterial strategies that can effectively deal with bacterial resistance and improve poor treatment effects has become increasingly urgent.

[0003] Cationic conjugated polymers are a class of polymer materials with unique structures and properties that have shown great potential for photothermal and photodynamic therapy. Their unique conjugated structure allows them to absorb near-infrared light of specific wavelengths. Upon exposure to near-infrared light, electron transitions and vibrational modes within their molecules are excited, efficiently converting the absorbed light energy into heat through non-radiative relaxation. This efficient photothermal conversion capability enables cationic conjugated polymers to generate localized high temperatures within a short period of time, thereby killing bacteria. Compared to some traditional photothermal agents, cationic conjugated polymers maintain high photothermal conversion efficiency even after repeated exposures and are less susceptible to photobleaching or photodegradation. This property enables cationic conjugated polymers to exert a sustained photothermal effect over an extended period of time, providing a stable heat source for photothermal therapy. Furthermore, upon exposure to near-infrared light, cationic conjugated polymers can transition from a ground state to an excited state. These energy is then transferred to surrounding oxygen molecules through energy transfer or electron transfer, converting them into highly oxidizing singlet oxygen. Singlet oxygen is a highly oxidizing agent that reacts with unsaturated fatty acids in cell membranes to form lipid peroxides. These peroxides alter the fluidity and permeability of cell membranes, causing damage and leakage. This allows for the leakage of intracellular substances such as electrolytes and amino acids, disrupting ion balance and normal metabolism within the cell, ultimately leading to bacterial death.

[0004] Therefore, it is necessary to further search for cationic conjugated polymers with better antibacterial effects. Summary of the Invention

[0005] Purpose of the invention: To solve the above technical problems, the present invention provides a cationic conjugated polymer and its preparation method and application. The cationic conjugated polymer obtained by the present invention has good photothermal and photodynamic properties and good antibacterial effect.

[0006] Technical solution: In order to achieve the above purpose, the present invention adopts the following technical solution:

[0007] A method for preparing a cationic conjugated polymer comprises the following steps:

[0008]

[0009] Step (1), under an inert atmosphere, tri(4-bromophenyl)amine and N,N-dimethyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline react in the presence of a catalyst and a base to produce intermediate 1;

[0010] Step (2), in an inert gas atmosphere, intermediate 1, compound 2 and compound 3 react in the presence of a catalyst to produce intermediate 4;

[0011] In step (3), intermediate 4 reacts with methyl halide to obtain a cationic conjugated polymer represented by formula (I).

[0012] As a specific embodiment, in step (1), the catalyst is selected from tetrakis(triphenylphosphine)palladium, and the base is potassium carbonate; the solvent used in the reaction is a mixed solution of an organic solvent and water; the molar ratio of tri(4-bromophenyl)amine, N,N-dimethyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline, catalyst and base is (90-110):(15-25):(0.8-1.2):(14-18).

[0013] Furthermore, the organic solvent is selected from tetrahydrofuran, and the volume ratio of the organic solvent to water is (4-6):1.

[0014] As a specific embodiment, in step (1), the reaction temperature is 70-90°C and the reaction time is 10-14h.

[0015] As a specific embodiment, in step (2), the catalyst is selected from tris(dibenzylideneacetone)dipalladium and tri(o-tolyl)phosphine; the solvent used in the reaction is toluene; the molar ratio of the intermediate 1, compound 2, compound 3, tris(dibenzylideneacetone)dipalladium and tri(o-tolyl)phosphine is (9-11):(90-96):(100-106):(4-6):(38-44).

[0016] As a specific implementation scheme, in step (2), the reaction temperature is 80°C-90°C, and the reaction time is 20-28h.

[0017] As a specific embodiment, in step (3), the methyl halide is selected from methyl iodide; the solvent used in the reaction is dichloromethane; and the molar ratio of the intermediate 4 to the methyl halide is (1-3):(80-120).

[0018] As a specific embodiment, in step (3), the reaction temperature is 20°C-40°C, and the reaction time is 20-28h.

[0019] The present invention also provides a cationic conjugated polymer, which is prepared by the above preparation method and has the following structure:

[0020]

[0021] Finally, the present invention also provides the use of the cationic conjugated polymer in preparing an antibacterial agent.

[0022] As a specific embodiment, the cationic conjugated polymer can effectively inhibit bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA) and Escherichia coli (E. coli), under near-infrared light irradiation.

[0023] Furthermore, the cationic conjugated polymer can produce good photothermal and photodynamic properties under near-infrared light irradiation, and the synergistic effect of the two can cause a significant inhibitory effect on bacteria.

[0024] Preferably, the concentration of the cationic conjugated polymer is 50-250 μg / mL.

[0025] Preferably, the conditions for the near-infrared light irradiation are: wavelength 980nm, optical density 0.2-0.8W / cm 2 , irradiation time is 0-10min.

[0026] Beneficial effects: Compared with the prior art, the present invention has the following advantages:

[0027] 1) The cationic conjugated polymer provided by the present invention, and its preparation method and application, have simple preparation method and high repeatability. The prepared cationic conjugated polymer has good optical properties and can produce photothermal and photodynamic synergistic effects.

[0028] 2) The cationic conjugated polymer provided by the present invention is positively charged and can interact with bacteria with negative surface charges, thereby inhibiting their growth and reproduction.

[0029] 3) The cationic conjugated polymer provided by the present invention has good biocompatibility and has no obvious toxic side effects on normal cells and major organs. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 This is the H NMR spectrum of the cationic conjugated polymer prepared in Example 1.

[0031] Figure 2 This is a diagram of the hydrated particle size of the cationic conjugated polymer nanoparticles prepared in Example 3.

[0032] Figure 3 This is the surface potential diagram of the cationic conjugated polymer nanoparticles prepared in Example 4.

[0033] Figure 4 This is the temperature-time diagram of cationic conjugated polymer nanoparticles with different concentrations in Example 6.

[0034] Figure 5 This is the temperature-time diagram of the cationic conjugated polymer nanoparticles in Example 7 under laser irradiation of different powers.

[0035] Figure 6 This is the photothermal stability of the cationic conjugated polymer nanoparticles in Example 8.

[0036] Figure 7 This is the photodynamic performance of the cationic conjugated polymer nanoparticles in Example 9.

[0037] Figure 8 The ability of the cationic conjugated polymer nanoparticles in Example 10 to produce ROS in bacteria.

[0038] Figure 9 This is the antibacterial property of the cationic conjugated polymer nanoparticles in Example 11.

[0039] Figure 10 This is the cytotoxicity of the cationic conjugated polymer nanoparticles in Example 12. DETAILED DESCRIPTION

[0040] In order to make the present invention easier to understand, specific embodiments of the present invention will be further described below.

[0041] The present invention is further described below in conjunction with the embodiments and drawings, the purpose of which is to help better understand the content of the present invention, but the scope of protection of the present invention is not limited thereto.

[0042] Unless otherwise defined herein, scientific and technical terms used in this application shall have the meanings commonly understood by one of ordinary skill in the art.

[0043] Example 1 Synthesis of cationic conjugated polymers PNPs.

[0044] (1) The structural formula of cationic conjugated polymer PNPs is as follows:

[0045]

[0046] (2) The synthetic route of the first intermediate is as follows:

[0047]

[0048] Tris(4-bromophenyl)amine (9.64 g, 20 mM), N,N-dimethyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline (0.99 g, 4 mM), tetrakis(triphenylphosphine)palladium (231 mg, 0.2 mM), and potassium carbonate (4.42 g, 32 mM) were added to a 100 mL round-bottom flask. The flask was evacuated and filled with dry nitrogen three times. Then, 50 mL of tetrahydrofuran and 10 mL of water were added, and the system was stirred at 80°C overnight. After cooling to room temperature, the mixture was extracted with dichloromethane. The organic phases were combined and dried over anhydrous sodium sulfate. After evaporating the solvent, the crude product was purified by silica gel column chromatography to obtain the first intermediate N,N-bis(4-bromophenyl)-N,N-dimethyl-[1,1'-biphenyl]-4,4'-diamine (yield 62%).

[0049] (3) The synthesis route of cationic conjugated polymer PNPs is as follows:

[0050]

[0051] Under nitrogen atmosphere, a mixture containing 4,8-bis(5-bromo-4-(2-octyldodecyl)thiophen-2-yl)-benzo[1,2-c;4,5-c']bis[1,2,5]thiadiazole (100.44 mg, 0.093 mM), 4,8-bis[(2-ethylhexyl)oxy]-2,6-bis(trimethyltinyl)benzo[1,2-b:4,5-b']dithiophene (8 4 mL of toluene was added to a mixture of 2-nitropropene (0.02 mg, 0.103 mM), N,N-bis(4-bromophenyl)-N,N-dimethyl-[1,1'-biphenyl]-4,4'-diamine (5.39 mg, 0.010 mM), tris(dibenzylideneacetone)dipalladium (4.74 mg, 0.005 mM) and tri(o-tolyl)phosphine (12.62 mg, 0.041 mM). The mixture was refluxed at 80°C for 24 hours. After cooling to room temperature, the mixture was poured into vigorously stirred methanol. After purification by a purification column, the obtained polymer was dissolved in 30 mL of dichloromethane, and 1 mL of iodomethane was added at 80°C (the molar ratio of polymer to iodomethane was 1:100), and the reaction was stirred for 24 hours. The mixture was then poured into vigorously stirred methanol and filtered to obtain a precipitate, which was a cationic conjugated polymer. Its nuclear magnetic hydrogen spectrum is shown as follows Figure 1 shown.

[0052] Example 2:

[0053] 1 mg of the cationic conjugated polymer synthesized in Example 1 and 2 mg of distearoylphosphatidylethanolamine-polyethylene glycol were added. 2000 The -amino group was added to 1 mL of tetrahydrofuran and sonicated to dissolve. The mixed solution was then injected into 9 mL of deionized water using a syringe and sonicated for 7 minutes. Subsequently, amphiphilic cationic conjugated polymer nanoparticles were obtained by freeze drying.

[0054] Example 3:

[0055] Take 1 mg of cationic conjugated polymer PNPs nanoparticles prepared in Example 2 and dissolve them in 10 mL of deionized water. Ultrasonicate them to make them fully dissolved. Use a pipette to draw 1 mL of the solution into a four-way cuvette. Use a laser particle size analyzer to test the hydrated particle size of the cationic conjugated polymer nanoparticles. The temperature equilibrium time is 120 s. The results are as follows Figure 2 As shown, the sizes of the obtained polymer nanoparticles are mainly concentrated in the range of 100-140 nm.

[0056] Example 4:

[0057] Take 1 mg of cationic conjugated polymer PNPs nanoparticles prepared in Example 2 and dissolve them in 10 mL of deionized water. Ultrasonicate them to make them fully dissolved. Use a pipette to draw 1 mL of the solution into the Zeta potential cell. The surface potential of the cationic conjugated polymer nanoparticles was tested using a laser particle size analyzer. The temperature equilibrium time was 120 s. The results are as follows: Figure 3 As shown, the surface potential of the prepared polymer nanoparticles is +24.4 mV.

[0058] Example 5:

[0059] 0.5, 1, 1.5, 2, and 2.5 mg of the cationic conjugated polymer PNPs nanoparticles prepared in Example 2 were added to 10 mL of deionized water, and ultrasonicated to fully dissolve them to obtain cationic conjugated polymer aqueous solutions with concentrations of 50, 100, 150, 200, and 250 μg / mL.

[0060] Example 6:

[0061] 0.5 mL of the cationic conjugated polymer PNPs aqueous solution of different concentrations (50, 100, 150, 200, 250 μg / mL) prepared in Example 5 was added to 1.5 mL centrifuge tubes and irradiated with a 980 nm near-infrared laser for 10 min. The laser power density was set to 0.4 W / cm 2 , use an infrared camera to record the temperature changes during the irradiation process, and the results are as follows Figure 4 As shown in the figure, the temperature of PNPs solutions with different concentrations can reach above 50°C after irradiation with 980nm near-infrared laser for 10 minutes, showing a good heating trend.

[0062] Example 7:

[0063] The cationic conjugated polymer PNPs aqueous solution (0.5 mL, 100 μg / mL) prepared in Example 5 was placed in a 1.5 mL centrifuge tube. 980 nm near-infrared laser was used for 10 min, and the laser power density was set to 0.2, 0.4, 0.6, and 0.8 W / cm 2 , use an infrared camera to record the temperature changes during the irradiation process, and the results are as follows Figure 5 As shown in Figure 2, the 100 μg / mL PNPs solution showed a large laser power dependence. When the power was greater than 0.4 W / cm 2 When the temperature of 100 μg / mL PNPs solution can reach above 50°C, it has certain antibacterial ability.

[0064] Example 8:

[0065] The cationic conjugated polymer PNPs aqueous solution (0.5 mL, 100 μg / mL) prepared in Example 5 was placed in a 1.5 mL centrifuge tube. 980 nm near-infrared laser irradiation was used for 10 min, followed by laser irradiation. Five cycles were performed, with the laser power density set to 0.4 W / cm 2 , use an infrared camera to record the temperature changes during the irradiation process, and the results are as follows Figure 6 As shown, PNPs have good photothermal stability.

[0066] Example 9:

[0067] The cationic conjugated polymer PNPs aqueous solution (3 mL, 100 μg / mL) prepared in Example 5 was mixed with a 1,3-diphenylisobenzofuran solution (10 mM) and placed in a quartz cuvette. The solution was irradiated with a 980 nm laser for 10 min, and the laser power density was set to 0.4 W / cm 2 Then, the UV absorption intensity was measured every 2 minutes using a UV spectrophotometer. The results were as follows: Figure 7 As shown in the figure, with the increase of irradiation time, the absorption value of DPBF at around 410 nm continues to decrease, indicating that PNPs have the ability to generate singlet oxygen under 980 nm laser irradiation.

[0068] Example 10:

[0069] 2',7'-dichlorodihydrofluorescein (10 μM) was incubated with methicillin-resistant Staphylococcus aureus (MRSA) and Escherichia coli (E. coli) suspensions at 37°C for 20 min. The mixture was centrifuged (3000 rpm, 5 min), washed three times with PBS, and then resuspended in a cationic conjugated polymer PNPs solution (100 μg / mL). The bacterial suspension was incubated at 37°C for 6 h and then irradiated with a 980 nm laser (0.4 W / cm 2 , 10min) and immediately observed under an inverted fluorescence microscope. The corresponding control group was set up under no light conditions. The results are as follows Figure 8 As shown, in the absence of laser irradiation, no fluorescence was produced in the bacteria co-cultured with PNPs, but after laser irradiation, bright green fluorescence was produced, indicating that PNPs have the ability to produce ROS in bacteria under laser irradiation.

[0070] Example 11:

[0071] Methicillin-resistant Staphylococcus aureus (MRSA) and Escherichia coli (E. coli) were inoculated into liquid culture medium and cultured in an incubator shaker at 37°C and 120 rpm for 12 hours. The bacterial solution was then diluted with liquid culture medium until the OD value at 600 nm reached 0.1. The bacteria were divided into four groups for treatment: PBS (control group), PNPs, PNPs+0.2W / cm 2 、PNPs+0.4W / cm 2 After treatment, continue to culture for 4 hours and dilute the bacterial solution to 10 -5 The plate was coated at multiple times, and the coated plate was incubated at 37℃ for 12 hours, and photographed and recorded. The results are as follows Figure 9 As shown, PNPs themselves have almost no antibacterial ability, but using 0.2W / cm 2 Under laser irradiation, the photodynamic properties of PNPs were activated, giving them a certain antibacterial ability. When the power reached 0.4W / cm 2 When the photothermal and photodynamic properties of PNPs are activated simultaneously, the two work synergistically to have strong antibacterial properties.

[0072] Example 12:

[0073] Mouse fibroblasts (L929) were cultured and seeded in 96-well plates. After the cells attached to the plates, they were divided into two groups. Equal amounts of culture medium and cationic conjugated polymer PNPs solution (100 μg / mL) were added to each group. After incubation at 37°C and 5% CO2 for 24 hours, the cells were washed twice with PBS. After that, calcein solution (5 μM) was added to each group. After staining in the dark for 30 minutes, cell activity was observed using an inverted fluorescence microscope. The results are shown in Figure 2. Figure 10 As shown in the figure, 100 μg / mL of PNPs had no obvious toxicity to cells.

[0074] Finally, it should be noted that the above specific implementation methods are only used to illustrate the technical solutions of the present invention and are not limiting. Although the present invention has been described in detail with reference to examples, those skilled in the art should understand that the technical solutions of the present invention can be modified or replaced by equivalents without departing from the spirit and scope of the technical solutions of the present invention, which should all be included in the scope of the claims of the present invention.

Claims

1. A method for preparing a cationic conjugated polymer, characterized in that: The following steps are involved: Step (1), under an inert atmosphere, tri(4-bromophenyl)amine and N,N-dimethyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline react in the presence of a catalyst and a base to produce intermediate 1; Step (2), in an inert gas atmosphere, intermediate 1, compound 2 and compound 3 react in the presence of a catalyst to produce intermediate 4; In step (3), intermediate 4 reacts with methyl halide to obtain a cationic conjugated polymer represented by formula (I).

2. The method for preparing a cationic conjugated polymer according to claim 1, wherein In step (1), the catalyst is selected from tetrakis(triphenylphosphine)palladium, and the base is potassium carbonate; the solvent used in the reaction is a mixed solution of an organic solvent and water; the molar ratio of tri(4-bromophenyl)amine, N,N-dimethyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline, catalyst and base is (90-110):(15-25):(0.8-1.2):(14-18).

3. The method for preparing a cationic conjugated polymer according to claim 1, wherein In step (1), the reaction temperature is 70-90°C and the reaction time is 10-14h.

4. The method for preparing a cationic conjugated polymer according to claim 1, wherein In step (2), the catalyst is selected from tris(dibenzylideneacetone)dipalladium and tri(o-tolyl)phosphine; the solvent used in the reaction is toluene; the molar ratio of the intermediate 1, compound 2, compound 3, tris(dibenzylideneacetone)dipalladium and tri(o-tolyl)phosphine is (9-11):(90-96):(100-106):(4-6):(38-44).

5. The method for preparing a cationic conjugated polymer according to claim 1, wherein In step (2), the reaction temperature is 80°C-90°C and the reaction time is 20-28h.

6. The method for preparing a cationic conjugated polymer according to claim 1, wherein In step (3), the methyl halide is selected from methyl iodide; the solvent used in the reaction is dichloromethane; and the molar ratio of the intermediate 4 to the methyl halide is (1-3):(80-120).

7. The method for preparing a cationic conjugated polymer according to claim 1, wherein In step (3), the reaction temperature is 20°C-40°C, and the reaction time is 20-28h.

8. A cationic conjugated polymer, characterized in that The cationic conjugated polymer is prepared by the preparation method according to any one of claims 1 to 7.

9. Use of the cationic conjugated polymer according to claim 8 in the preparation of an antibacterial agent.

10. The use according to claim 9, characterized in that The cationic conjugated polymer can effectively inhibit bacteria under near-infrared light irradiation.

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