A kind of tetraphenyl ethene skeleton pyridine salt class singlet oxygen type photosensitizer and its application in preparation of bacteriostatic preparation

Abstract

The application discloses a tetraphenyl ethene skeleton pyridine salt singlet oxygen type photosensitizer and application thereof in preparation of bacteriostatic agents, wherein the tetraphenyl ethene skeleton pyridine salt singlet oxygen type photosensitizer has a general structure as shown in the following formula (I): wherein X is selected from one of I ‑ , Br ‑ , Cl ‑ , F ‑ , OH ‑ , NO3 ‑ , SHO3 ‑ and PF6 ‑ ; R1 is selected from an aromatic ring derivative electron donor group; and R2 is selected from one of an alkyl group, an alkoxy group, a 4-boronic acid phenyl group and a 4-methyl benzene boronic acid pinacol ester. The photosensitizer combines AIE fluorescence enhancement characteristics and high ROS generation capacity, significantly improves bacteriostatic selectivity and bacteriostatic efficiency, and provides a new strategy for APDT treatment of bacterial infection.

Description

TECHNICAL FIELD

[0001] The present application relates to a photosensitizer and its application, in particular to a kind of tetraphenyl ethene skeleton pyridine salt single oxygen type photosensitizer and its bacteriostatic application. BACKGROUND

[0002] Bacterial infection poses a multidimensional threat to human health, with far-reaching implications. Bacterial transmission routes are diverse and complex, including but not limited to the digestive tract, respiratory tract, and arthropod vectors. Foodborne bacterial diseases often cause gastrointestinal dysfunction in groups, with symptoms such as diarrhea and vomiting, not only affecting the quality of life of patients, but also potentially causing serious complications, including direct mortality, consumption of medical resources, exacerbation of antibiotic resistance, and long-term impact on the social economy. Antibiotics are one of the main drugs for treating bacterial infections, and they exert therapeutic effects by inhibiting bacterial growth or killing bacteria. The widespread and inappropriate use of antibiotics has led to global bacterial resistance problems, not only reducing the effectiveness of antibiotics, but also challenging the development of alternative therapies, new drug research and development, and the implementation of public health policies.

[0003] Photodynamic therapy (PDT) is an innovative treatment method, and PDT relies on the production of reactive oxygen species (ROS) by photosensitizers under light irradiation to cause damage to the structure and function of bacterial cells, achieving targeted inactivation of bacteria, and is an effective alternative or complementary treatment strategy in dealing with bacterial resistance. The performance of photosensitizers, including photochemical stability, light absorption characteristics, and ROS generation efficiency, is crucial for PDT efficacy. The aggregation of traditional photosensitizers in vivo can lead to aggregation-induced quenching (ACQ) phenomenon, reducing photochemical activity and ROS yield, thus limiting their application.

[0004] Aggregation-induced emission (AIE) is a unique photophysical phenomenon that fundamentally differs from ACQ. AIEgens exhibit weak or no fluorescence in dilute solution or single-molecule state, but their fluorescence significantly enhances in aggregated state. AIEgens have high ROS generation capacity in aggregated state, which further achieves bacterial inactivation, and exhibit significant potential for antibacterial photodynamic therapy (APDT) applications.

[0005] The inventor has made the following search on the related content of the present application:

[0006] 1, Google Scholar search results: (2024 / 7 / 8)

[0007]

[0008]

[0009] 2, CNKI search results: (2024 / 7 / 8)

[0010] Search method one:

[0011] Article title: AIE active pyridine-benzene borate single state oxygen type active oxygen photosensitizer antibacterial activity No related literature.

[0012] Article title: Pyridine-benzene borate single state oxygen type active oxygen photosensitizer antibacterial activity No related literature.

[0013] Article title: Single state oxygen type active oxygen photosensitizer antibacterial activity No related literature.

[0014] Search method two:

[0015] Full text: AIE active pyridine-benzene borate single state oxygen type active oxygen photosensitizer antibacterial activity 2, both unrelated to target compounds.

[0016] Full text: Pyridine-benzene borate single state oxygen type active oxygen photosensitizer antibacterial activity No related literature.

[0017] Full text: Single state oxygen type active oxygen photosensitizer antibacterial activity 327, all unrelated to target compounds.

[0018] Search method three:

[0019] Key words: AIE active pyridine-benzene borate single state oxygen type active oxygen photosensitizer antibacterial activity No related literature.

[0020] Key words: Pyridine-benzene borate single state oxygen type active oxygen photosensitizer antibacterial activity No related literature.

[0021] Key words: Single state oxygen type active oxygen photosensitizer antibacterial activity No related literature. Summary of the invention

[0022] The present application aims at the deficiencies of the prior art, and provides a tetraphenyl ethene skeleton pyridine salt type singlet oxygen type photosensitizer and bacteriostatic application thereof.

[0023] The tetraphenyl ethene skeleton pyridine salt type singlet oxygen type photosensitizer provided by the present application has the general structure shown in the following formula (I):

[0024]

[0025] X is selected from one of iodine ion I - , bromine ion Br - , chlorine ion Cl - , fluorine ion F - , hydroxyl ion OH - , nitrate NO3 - , sulfonic acid ion SHO3 - , and hexafluorophosphate ion PF6 - . R1 is an aromatic ring derivative electron-donating group, including but not limited to diphenylamine group, triphenylamine group, ring octatetra-thiophene group, tetraphenyl ethene group, etc. R2 is selected from alkyl, alkoxy, 4-boronic acid phenyl, 4-methyl phenyl boronic acid pinacol ester, etc.

[0026] Further preferably, it is one of the structures shown in the following formula A-L:

[0027]

[0028]

[0029] Further, R1 is a tetraphenyl ethene group, R2 is a 4-boronic acid benzyl group, and X is a bromine ion.

[0030] The tetraphenyl ethene skeleton pyridine salt type singlet oxygen type photosensitizer provided by the present application is applied to the preparation of a bacteriostatic preparation.

[0031] The bacteriostatic preparation has a bacteriostatic effect under light irradiation.

[0032] The bacteria include one or more of gram-negative bacteria E. coli, gram-positive bacteria B. megaterium, B. cereus, B. subtilis WB-800 subspecies, B. subtilis 1285 subspecies, and S. aureus.

[0033] The present application simultaneously uses the unsalted pyridine derivative as the control of the singlet oxygen type photosensitizer of the tetraphenylethene skeleton pyridine salt, and the structure is shown in the following formula M:

[0034]

[0035] Compared with the prior art, the present application has the following advantages and beneficial effects:

[0036] 1. The present application synthesizes a new singlet oxygen type ROS material system with AIE performance.

[0037] 2. The present application synthesizes the singlet oxygen type photosensitizer by a simple and effective method.

[0038] 3. The singlet oxygen type photosensitizer of the present application can inhibit the growth of various bacteria at normal temperature and pressure, and has a broad-spectrum antibacterial effect.

[0039] 4. The singlet oxygen type photosensitizer of the present application has a strong antibacterial effect on the foodborne pathogenic bacteria B. cereus which can cause food poisoning. BRIEF DESCRIPTION OF DRAWINGS

[0040] Figure 1 The left graph in the middle is the fluorescence emission spectrum of TBPB (10 μM) in DMSO / Tol (v / v) mixed solvent with increasing toluene content, λ ex = 400 nm; the right graph is the change graph of the fluorescence intensity of TBPB (10 μM) in DMSO / Tol (v / v) mixed solvent with the toluene content of the solution.

[0041] Figure 2 (A) in the middle is the fluorescence intensity of DCF-H solution containing the control M (4 μM) and Ce6 (4 μM), TBPB (4 μM) and Ce6 (4 μM) under white light irradiation, and the active oxygen generation efficiency of TBPB is much higher than that of M, which is verified by the DCF-DA fluorescent probe; (B) is the degradation rate of ABDA solution containing M (10 μM) and RB (10 μM), TBPB (10 μM) and RB (10 μM) under white light irradiation, and M and TBPB both produce singlet oxygen type ROS species, which is verified by ABDA.

[0042] Figure 3Inactivation of different bacteria by TBPB under dark and light, including Gram-negative bacteria E. coli, Gram-positive bacteria B. megaterium, B. cereus and B. subtilis (three subtypes: 168, 1285, wb800), Methicillin-Resistant Staphylococcus aureus (MRSA). [TBPB] = 50 μΜ, Scale bar = 2 cm, light energy density: 26 mW cm -2 , light exposure time: 30 min.

[0043] Figure 4 Inactivation of B. cereus by different concentrations of TBPB under dark and light. Scale bar = 2 cm, light energy density: 26 mW cm -2 , light exposure time: 30 min.

[0044] Figure 5 Inactivation of B. cereus by TBPB under different light exposure times. [TBPB] = 50 μΜ, Scale bar = 2 cm, light energy density: 26 mW cm -2 .

[0045] Figure 6 Plate experiment results of M on different bacteria under dark and light, including Gram-negative bacteria E. coli, Gram-positive bacteria B. cereus and Methicillin-Resistant Staphylococcus aureus (MRSA). [M] = 50 μΜ, Scale bar = 2 cm, light energy density: 26 mW cm -2 , light exposure time: 30 min.

[0046] Figure 7 Fluorescence imaging results of M and TBPB on different bacteria, including Gram-positive bacteria E. coli, Gram-negative bacteria B. cereus and Methicillin-Resistant Staphylococcus aureus (MRSA). [M] = 10 μΜ, λ ex = 405 nm, λ em = 590-690 nm. Scale bar = 10 μm. [TBPB] = 10 μΜ, λex = 405 nm, λ em = 600-700 nm. Scale bar = 10 μm. DETAILED DESCRIPTION

[0047] The present application will be described in further detail by working examples and drawings, but embodiments of the present application are not limited thereto.

[0048] Example 1: Synthesis of pyridine salt type singlet oxygen type active oxygen photosensitizer (TBPB)

[0049]

[0050] 1. Synthesis of intermediate TPE-BTZ-Br

[0051]

[0052] In a 100 mL round bottom flask, compound [1-(4-boronic acid ester phenyl)-1,2,2-triphenyl] ethylene (1 g, 2.66 mmol) and 4,7-dibromo benzo[c]-1,2,5-thiadiazole (1.17 g, 3.99 mmol) were dissolved in 24 mL of tetrahydrofuran, and 6 mL of potassium carbonate aqueous solution (2 M) and tetrakis(triphenylphosphine)palladium were added. The reaction was performed under reflux in a nitrogen atmosphere, and TLC was used for tracking. After the reaction was completed, it was cooled to room temperature, water and dichloromethane were added for extraction, the organic layer was dried with anhydrous sodium sulfate, and after filtration, the organic layer was evaporated, and the substance was separated and purified by column chromatography (developing agent: petroleum ether:dichloromethane = 15:1, V / V), and vacuum dried to obtain a yellowish product. 1 H NMR (600 MHz, d6-DMSO), δ (ppm): 8.04 (d, J = 6.00 Hz, 1H), 7.76 (d, J = 6.00 Hz, 2H), 7.70 (d, J = 6.00 Hz, 1H), 7.16-7.07 (m, 11H), 7.04 (d, J = 12.00 Hz, 2H), 7.00 (d, J = 6.00 Hz, 2H), 6.96 (d, J = 6.00 Hz, 2H). MS (ESI): Calcd for C 32 H 21 BrN2S) 545.50, found m / z: 545.0673.

[0053] 2. Synthesis of control M

[0054]

[0055] In a 100 mL round flask, intermediate TPE-BTZ-Br (500 mg, 916.59 pmol) and 4-pyridineboronic acid (169 mg, 1.37 mmol) and 24 mL THF were added, after it was dissolved, 63 mg tetrakis(triphenylphosphine)palladium and 2 M K2C03solution (6 mL) were added, and the reaction was refluxed under nitrogen atmosphere. TLC was used to track the reaction, after it was completed, it was extracted with CH2CI2, the organic layer was dried over anhydrous Na2S04, filtered and rotary evaporated, and purified by silica gel column chromatography (eluent: DCM:EA = 9: 1, V / V) to obtain orange reference M. 1 HNMR (700 MHz, d6-DMSO): δ 8.75 (d, J = 7.00 Hz, 1H), 8.12 (d, J = 7.00 Hz, 1H), 8.06 (d, J = 7.00 Hz, 1H), 7.97 (m, J = 7.00 Hz, 2H), 7.86 (t, J = 10.50 Hz, 3H), 7.22-7.12 (m, 14H), 7.09 (d, J = 7.00 Hz, 3H), 7.06 (d, J = 7.00 Hz, 2H), 7.02 (d, J = 7.00 Hz, 2H), 1.23 (s, 2H). MS (ESI): Calcd for (C 37 H 25 N3S) 543.69, found m / z: [TBP + H] + 544.18.

[0056] 3. Synthesis of target product TBPB

[0057]

[0058] In a 25 mL round flask, reference M (100 mg, 0.22 mmol), 4-bromomethylphenylboronic acid pinacol ester (65 mg, 0.22 mmol) and 5 mL DMF were added, and the reaction was refluxed under nitrogen atmosphere. After it was completed, excess toluene was added, and black precipitate was separated, and the black solid product TBPB was obtained by filtration. 1 HNMR (700 MHz, d6-DMSO): δ 8.75 (d, J = 7.00 Hz, 1H), 8.12 (d, J = 7.00 Hz, 1H), 8.06 (d, J = 7.00 Hz, 1H), 7.97 (m, J = 7.00 Hz, 2H), 7.86 (t, J = 10.50 Hz, 3H), 7.22-7.12 (m, 14H), 7.09 (d, J = 7.00 Hz, 3H), 7.06 (d, J = 7.00 Hz, 2H), 7.02 (d, J = 7.00 Hz, 2H), 1.23 (s, 2H). MS (ESI): Calcd for (C 29 H 20 N4S) 758.54, found m / z: [TBPB-Br + 678.24.

[0059] Example 2: AIE property characterization of pyridine salt singlet oxygen type active oxygen photosensitizer (TBPB)

[0060] Figure 1 is the fluorescence spectrum of the material obtained based on Example 1 under different toluene content conditions. As can be seen from the figure, the example material emits little light in pure dimethyl sulfoxide solvent, and the fluorescence gradually increases with the addition of toluene, which is a poor solvent. When the toluene content is increased to 99%, the fluorescence intensity of TBPB reaches the strongest state, indicating that TBPB photosensitizer has AIE property.

[0061] Example 3: Active oxygen species identification of singlet oxygen type active oxygen photosensitizer (TBPB)

[0062] Figure 2 is the active oxygen species identification based on the material obtained in Example 1. As can be seen from Figure 2 A, when DCFDA fluorescent probe is used to detect active oxygen, after adding photosensitizer TBPB and control M and irradiating with white light, the DCF fluorescence intensity is obviously enhanced, indicating that M and TBPB can both produce active oxygen. By comparing the fluorescence intensity changes of the two groups, it can be observed that the active oxygen production efficiency of TBPB is obviously higher than that of M. Figure 2 B, 9, 10-Anthracenediyl-bis(methylene) dimalonic acid (ABDA) is used to directly prove that M and TBPB produce singlet oxygen type active oxygen. ABDA is used as a singlet oxygen trapping agent, and after photoactivation, ABDA+M group and ABDA+TBPB group show obvious ABDA ultraviolet absorption reduction. This result shows that M and TBPB both produce singlet oxygen type ROS species under light conditions.

[0063] Example 4: Inactivation performance of pyridine salt singlet oxygen type active oxygen photosensitizer (TBPB) on bacteria

[0064] Figure 3 is the inactivation performance experiment results of TBPB under dark and light conditions on different bacteria. The experimental bacteria include gram-negative bacteria E. coli, gram-positive bacteria B. megaterium, B. cereus, B. subtilis WB-800 subspecies and B. subtilis 1285 subspecies, and Methicillin-Resistant Staphylococcus aureus (MRSA). After the bacteria were cultured for 18 hours, 50 μM TBPB was added to the bacterial solution, and white light was irradiated for 30 min. 1 x 105 CFU / mL of bacteria solution 200 μL was uniformly spread on solid medium (LB agar) and incubated at 37 °C for 18 hours. From the results, it can be seen that TBPB can inhibit the growth of bacteria in the range of investigation, has broad-spectrum bacteriostatic property, and has strong bacteriostatic effect on drug-resistant bacteria MRSA. TBPB can completely kill B. s and B. m, and has bactericidal property.

[0065] Figure 4 The experimental results of the inactivation performance of TBPB on B. cereus under dark and light conditions at different concentrations. After the bacteria were cultured for 18 hours, 10, 30, 50, 70, and 90 μM TBPB were added to the bacterial solution, and white light was irradiated for 30 min. 1 x 10 5 CFU / mL of bacteria solution 200 μL was uniformly spread on solid medium (LB agar) and incubated at 37 °C for 18 hours. From the results, it can be seen that 10 μM TBPB can completely inactivate B. cereus.

[0066] Figure 5 The experimental results of the inactivation performance of TBPB on B. cereus under different light irradiation times. After the bacteria were cultured for 18 hours, 50 μM TBPB was added to the bacterial solution, and white light was irradiated for 5, 10, 15, 20, 25, and 30 min, respectively. 1 x 10 5 CFU / mL of bacteria solution 200 μL was uniformly spread on solid medium (LB agar) and incubated at 37 °C for 18 hours. From the results, it can be seen that 10 μM TBPB can completely inactivate B. cereus.

[0067] Example 5: Inactivation performance of control M on bacteria

[0068] Figure 6 The experimental results of the inactivation performance of control M on different bacteria under dark and light conditions. The experimental bacteria include gram-negative bacteria E. coli, gram-positive bacteria B. cereus, and Methicillin-Resistant Staphylococcus aureus (MRSA). After the bacteria were cultured for 18 hours, 50 μM control M was added to the bacterial solution, and white light was irradiated for 30 min. 1 x 10 5 CFU / mL of bacteria solution 200 μL was uniformly spread on solid medium (LB agar) and incubated at 37 °C for 18 hours. From the results, it can be seen that M has no bacteriostatic property on gram-positive bacteria and gram-negative bacteria.

[0069] Example 6: Fluorescent imaging of bacteria by singlet oxygen-type reactive oxygen light sensitizer TBPB and control M

[0070] Figure 7 For the fluorescent imaging of TBPB and control M on various bacteria, including gram-negative bacteria E. coli, gram-positive bacteria B. cereus and Methicillin-Resistant Staphylococcus aureus (MRSA). After the bacteria were cultured for 18 hours, 10 μM TBPB and 10 μM control M were added to the bacterial solution, respectively, and acted for 10 min at 37°C, then washed with PBS for 3 times, and then imaged and characterized by laser confocal microscope. From the results, it can be seen that TBPB can be uniformly distributed on the surface of the bacteria, realizing the imaging of the morphology of the bacteria, while the control M cannot act on the bacteria, thus it is difficult to perform fluorescent imaging, which is also the reason why M has no antibacterial effect.

Claims

1. A tetraphenylethylene skeleton pyridine salt singlet oxygen-type photosensitizer, characterized in that... Its structural formula is shown below: 。 2. The application of the tetraphenylethylene skeleton pyridine salt singlet oxygen photosensitizer according to claim 1 in the preparation of antibacterial agents, characterized in that: The bacteria in question is Bacillus cereus.

3. The application of the tetraphenylethylene skeleton pyridine salt singlet oxygen photosensitizer according to claim 1 in the preparation of antibacterial agents, characterized in that: The antibacterial agent has an inhibitory effect on bacteria under light conditions; The bacteria are one or more of Bacillus megaterium, Bacillus cereus, and methicillin-resistant Staphylococcus aureus.

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