TPA-AN and pyridine conjugate modified BODIPY fluorescent probe as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of organic synthesis, and particularly discloses a TPA-AN and pyridine conjugate modified BODIPY fluorescent probe as well as a preparation method and application thereof. According to the near-infrared NO-BDP fluorescent probe and the preparation method thereof, the mose site of BODIPY is directionally modified by TPA-AN, pyridine is unilaterally modified on the alpha site of a BODIPY parent nucleus, an expanded conjugated system is constructed through Knoevenagel condensation, and the near-infrared NO-BDP fluorescent probe is obtained. Wherein the TPA-AN provides a strong electron supply effect and an intramolecular charge transfer (ICT) channel, and the stereo steric hindrance of a pyridine ring enables a molecular structure to be twisted into a non-planar configuration, so that pi-pi accumulation is reduced, and an ACQ effect is inhibited. Wherein a TPA-AN group and a BODIPY parent nucleus have a synergistic coordination effect, so that NO-BDP can sensitively recognize a phenylalanine dipeptide structural unit in the A beta 42 protein, and then sensitive recognition of the A beta protein is realized; and a pyridine group is introduced to realize electron conduction of NO-BDP, so that a D-pi-A structure is formed, more singlet oxygen is generated, a relatively strong antibacterial effect is shown, and photodynamic antibiosis is realized. Finally, multifunctional application is realized through the NO-BDP fluorescent probe.

Description

Technical Field

[0001] This invention belongs to the field of organic synthesis technology and relates to the synthesis of BODIPY-type fluorescent probes. Specifically, it relates to a TPA-AN and pyridine conjugated modified BODIPY probe that can sensitively recognize Aβ protein and has photodynamic antibacterial properties, its preparation method, and its application in the preparation of related drugs. Background Technology

[0002] Fluoroboropyrrole (BODIPY) is considered one of the most versatile fluorophores due to its excellent photophysical properties, including high photostability, high fluorescence quantum yield, ease of modification, and narrow absorption and emission spectral peaks. It is widely used in the detection and identification of Aβ protein and in photodynamic therapy. The triphenylamine group, with its unique "equilateral triangle" non-planar molecular configuration and trilobed propeller-like structure, helps avoid aggregation-induced fluorescence quenching (ACQ) caused by π-π stacking. Typically, the electron-donating group in crops can cause a redshift in the absorption and emission spectra of organic fluorescent compounds. By binding to the phenylalanine dipeptide structural unit within the Aβ42 protein, it enhances the signal response of the BODIPY compound probe, improves the sensitivity of probe detection, and enables rapid identification of the Aβ42 protein. Furthermore, a pyridine group is introduced into one side of the NO-BDP probe, which absorbs energy to transition from a stable ground state to an unstable singlet excited state. Subsequently, the singlet excited state undergoes intersystem crossing to generate a triplet excited state. The photosensitizer in the excited state can react with surrounding macromolecules through energy transfer or electron transfer to generate more singlet oxygen, exhibiting a strong antibacterial effect and achieving photodynamic antibacterial activity. This leads to the formation of a D-π-A structure, ultimately realizing the multifunctional application of the NO-BDP probe. Summary of the Invention

[0003] To address the shortcomings of existing technologies, the present invention aims to provide a TPA-AN and pyridine conjugated BODIPY fluorescent probe. This fluorescent probe not only sensitively recognizes Aβ42 protein but also enables its own electron conduction, thereby forming a D-π-A structure, generating more singlet oxygen, exhibiting a strong antibacterial effect, and achieving photodynamic antibacterial activity. Another objective of the present invention is to provide a method for preparing this fluorescent probe.

[0004] This invention is achieved through the following technical solution: A TPA-AN conjugated with pyridine-modified BODIPY fluorescent probe has the following molecular structure: .

[0005] A further improvement to the present invention is as follows: A method for preparing a TPA-AN conjugated BODIPY fluorescent probe includes the following steps: (1) 4-(bis(4-hydroxyphenyl)amino)benzaldehyde, 2,4-dimethylpyrrole and boron fluoride diethyl ether were reacted in a one-pot reaction to give compound O-BDP; (2) Compound O-BDP is condensed with 1-formyl-methylpyrrole iodide to obtain compound NO-BDP, which is the target BODIPY fluorescent probe. The reaction route is shown below: .

[0006] Further, in step (1), 4-(bis(4-hydroxyphenyl)amino)benzaldehyde is dissolved in dichloromethane, 2,4-dimethylpyrrole is added under nitrogen protection, and then trifluoroacetic acid is added dropwise; after stirring at room temperature for a period of time, 2,3-dimethyl-5,6-dicyanobenzoquinone is added as a catalyst, and after a certain period of time, triethylamine and boron fluoride diethyl ether are slowly added, and the reaction is continued to be stirred at room temperature. After the reaction is completed, the mixed solution is washed with water, and the organic layer is concentrated and purified by column chromatography to obtain compound O-BDP.

[0007] Furthermore, the molar ratio of 4-(bis(4-hydroxyphenyl)amino)benzaldehyde, 2,4-dimethylpyrrole, trifluoroacetic acid, 2,3-dimethyl-5,6-dicyanobenzoquinone, triethylamine, and boron fluoride ether is 1:2~3:0.1~0.2:0.8~1.2:10~20:10~20.

[0008] Further, in step (2), compound O-BDP and 4-formyl-1-methylpyrrole iodide are added to toluene, along with a small amount of piperidine and acetic acid. The mixed solution is heated under nitrogen protection and the reaction is completed. After the reaction, the compound NO-BDP is obtained by separation and purification.

[0009] Furthermore, the molar ratio of the compound O-BDP, 4-formyl-1-methylpyrrole iodide, piperidine, and acetic acid is 1:2~3:2~4:0.1~0.2.

[0010] Furthermore, the heating reaction is carried out at a temperature of 120~130℃ for a time of 1~3h.

[0011] A further improvement of the present invention is as follows: The application of the above-mentioned TPA-AN conjugated with pyridine-modified BODIPY fluorescent probe in the preparation of AD diagnostic drugs.

[0012] Furthermore, the above-mentioned TPA-AN conjugated with pyridine-modified BODIPY fluorescent probe is used in the preparation of photodynamic therapy drugs.

[0013] Furthermore, the above-mentioned TPA-AN conjugated with pyridine-modified BODIPY fluorescent probe is used in the preparation of antibacterial drugs for photodynamic therapy.

[0014] Compared with the prior art, the beneficial effects of the present invention are as follows: This invention employs 4-(bis(4-hydroxyphenyl)amino)benzaldehyde (TPA-AN) to directionally modify the moose position of BODIPY, and unilaterally modifies the α-position of the BODIPY core with pyridine. An extended conjugated system is constructed via Knoevenagel condensation to obtain a near-infrared NO-BDP fluorescent probe. TPA-AN provides a strong electron-donating effect, an intramolecular charge transfer (ICT) channel, and the steric hindrance of the pyridine ring distorts the molecular structure into a non-planar configuration, reducing π-π stacking and suppressing the ACQ effect. The synergistic coordination effect between the TPA-AN group and the BODIPY core enables NO-BDP to sensitively recognize the phenylalanine dipeptide structural unit in the Aβ42 protein, thereby achieving sensitive recognition of the Aβ protein. Furthermore, the introduction of the pyridine group allows NO-BDP to achieve electron conduction, forming a D (donor)-π-A (acceptor) structure, generating more singlet oxygen and exhibiting a strong antibacterial effect, achieving photodynamic antibacterial activity. Ultimately, a multifunctional application is achieved using the NO-BDP fluorescent probe.

[0015] The NO-BDP fluorescent probe provided by this invention has a mild preparation process and simple steps, and has great potential and application value in the field of AD diagnosis and photodynamic therapy drugs. Attached Figure Description

[0016] Figure 1 The 1H NMR spectrum of the NO-BDP fluorescent probe prepared in Example 2 and conjugated with pyridine is shown. Figure 2 The mass spectrum of the NO-BDP fluorescent probe prepared in Example 2 and modified with TPA-AN and pyridine conjugation; Figure 3 This is a protein binding response diagram of the NO-BDP fluorescent probe, which is conjugated with TPA-AN and pyridine according to the present invention. Figure 4 The graph shows the generation rate of singlet oxygen by the NO-BDP fluorescent probe modified with TPA-AN and pyridine according to the present invention. Figure 5 This image shows the photodynamic sterilization effect of the NO-BDP fluorescent probe, which is conjugated with TPA-AN and pyridine according to the present invention. Detailed Implementation

[0017] The present invention will now be described in detail with reference to specific embodiments.

[0018] Example 1: Preparation of compound O-BDP 4-(bis(4-hydroxyphenyl)amino)benzaldehyde (10 mmol) was dissolved in dichloromethane (400 mL), and 2,4-dimethylpyrrole (22 mmol) was added under nitrogen protection, followed by the dropwise addition of trifluoroacetic acid (2.2 mmol). After stirring at room temperature for 12 h, 2,3-dimethyl-5,6-dicyanobenzoquinone (10 mmol) was added as a catalyst, and stirring was continued for 2 h. Triethylamine (10 mL) and boron fluoride diethyl ether (12 mL) were slowly added, and stirring was continued for 2 h. The mixture was washed with water, and the organic layer was dried and concentrated on a rotary evaporator. Finally, it was purified by silica gel column chromatography to obtain compound O-BDP.

[0019] Example 2: Synthesis of NO-BDP fluorescent probe O-BDP (0.5 mmol) and 4-formyl-1-methylpyrrole iodide (1.5 mmol) were added to toluene (30 mL), followed by piperidine (1 mmol) and acetic acid (0.1 mmol). The mixture was heated to 125 °C for 2 h under nitrogen protection. The final product was purified by silica gel column chromatography. The compound NO-BDP was obtained, and its NMR was determined. 1 H NMR spectrum, mass spectrometry MS EI + To characterize, 1 H NMR (400 MHz, Chloroform-d) δ 8.81 – 8.78 (m, 2H), 8.10– 8.07 (m, 2H), 7.85 – 7.81 (m, 2H), 7.55 (d, J = 14.1 Hz, 1H), 7.19 – 7.15(m, 2H), 7.09 – 7.05 (m, 4H), 6.91 – 6.88 (m, 4H), 6.78 (s, 1H), 6.01 – 5.99(m, 1H), 4.38 (s, 3H), 3.80 (s, 6H), 2.57 (d, J = 1.2 Hz, 3H), 2.25 (s, 3H),1.93 (d, J = 1.5 Hz, 3H); MS EI + : 655.58.

[0020] Example 3: NO-BDP fluorescent probe for Aβ42 protein recognition analysis: Diphenylalanine dipeptide (3 mg) was dissolved in 45 μl of hexafluoroisopropanol, and then 105 μl of water was injected. The mixture was sonicated at room temperature for 5 minutes, and the diphenylalanine dipeptide self-assembled into nanofibers. Hexafluoroisopropanol was removed by water dialysis, and the resulting diphenylalanine dipeptide nanofibers were stored in water for later use. A NO-BDP probe (1 mg) was dissolved in 45 μl of hexafluoroisopropanol and injected into the diphenylalanine dipeptide nanofiber aqueous solution. After 2 hours, fluorescently labeled diphenylalanine dipeptide nanofibers were obtained, achieving action on Aβ protein and producing a fluorescent response. Figure 3 As shown.

[0021] Example 4: Determination of singlet oxygen by NO-BDP fluorescent probe under in vitro illumination To evaluate whether the prepared NO-BDP probe can generate [something] under laser irradiation. 1 O2 was determined by the characteristic absorption peak changes of 1,3-bis(4-phenyl)vinylbenzofuran (DPBF) to measure the in vitro production of BODIPY. 1 The ability to absorb O2 was assessed. DPBF (2 mg / mL, 30 μL) was added to the NO-BDP probe solution, with only DPBF and EtOH as controls. The UV-Vis absorption spectrum was measured every 30 s under laser irradiation (λ=545 nm, 100 mW), and the change in absorbance at 415 nm was recorded. Figure 4 As shown, the singlet oxygen decrease rate reaches 80%.

[0022] Example 5: In vitro colony counting experiment using NO-BDP fluorescent probe under light irradiation conditions The in vitro antibacterial activity of the NO-BDP probe (3.20% (v / v)) was evaluated using a live colony counting method. First, Staphylococcus aureus (10... 4 CFU / mL) was mixed with PBS, DMSO, EtOH, and NO-BDP, respectively. The mixtures were irradiated with a 545 nm laser at 100 mW for 30 min, while the control group was not irradiated. The mixtures were then spread on LB agar plates and incubated at 37°C for 24 h. Colony growth was recorded by photographing, and bacterial survival rates in different treatment groups were calculated. This experiment consisted of four control groups: samples with BODIPY and light (+BDP, +light), samples with BODIPY and light-protected (+BDP, -light), samples without BODIPY and light-protected (-BDP, +light), and samples without BODIPY and light-protected (-BDP, -light). Figure 5 As shown.

[0023] The above description of the embodiments is only for illustrating the technical concept and features of the present invention. Its purpose is to enable those skilled in the art to understand the content of the present invention and implement it accordingly. Those skilled in the art can obviously easily make various modifications to these embodiments and apply the general principles described herein to other embodiments without creative effort. Therefore, the above embodiments should not be used to limit the scope of protection of the present invention. All improvements and modifications made by those skilled in the art based on the disclosure of the present invention without departing from the scope of the present invention should be covered within the scope of protection of the present invention.

Claims

1. A TPA-AN conjugated with pyridine-modified BODIPY fluorescent probe, characterized in that, The molecular structure of the fluorescent probe is shown in the following formula: 。 2. The method for preparing a TPA-AN conjugated and pyridine-modified BODIPY fluorescent probe as described in claim 1, characterized in that, Includes the following steps: (1) 4-(bis(4-hydroxyphenyl)amino)benzaldehyde, 2,4-dimethylpyrrole and boron fluoride diethyl ether were reacted in a one-pot reaction to give compound O-BDP; (2) Compound O-BDP is condensed with 1-formyl-methylpyrrole iodide to obtain compound NO-BDP, which is the target BODIPY fluorescent probe. The reaction route is shown below: 。 3. The preparation method according to claim 2, characterized in that: In step (1), 4-(bis(4-hydroxyphenyl)amino)benzaldehyde was dissolved in dichloromethane, and 2,4-dimethylpyrrole was added under nitrogen protection, followed by the dropwise addition of trifluoroacetic acid. After stirring at room temperature for a period of time, 2,3-dimethyl-5,6-dicyanobenzoquinone was added as a catalyst, and the reaction was continued for a certain period of time. Triethylamine and boron fluoride ether were then slowly added, and the reaction was continued to be stirred at room temperature. After the reaction was completed, the mixed solution was washed with water, and the organic layer was concentrated and purified by column chromatography to obtain compound O-BDP.

4. The preparation method according to claim 3, characterized in that: The ratio of 4-(bis(4-hydroxyphenyl)amino)benzaldehyde, 2,4-dimethylpyrrole, trifluoroacetic acid, 2,3-dimethyl-5,6-dicyanobenzoquinone, triethylamine, and boron fluoride ether is 1:2~3:0.1~0.2:0.8~1.2:10~20:10~20.

5. The preparation method according to claim 2, characterized in that: In step (2), compound O-BDP and 4-formyl-1-methylpyrrole iodide are added to toluene, along with a small amount of piperidine and acetic acid. The mixed solution is heated under nitrogen protection and the reaction is completed. After the reaction, the compound NO-BDP is obtained by separation and purification.

6. The preparation method according to claim 5, characterized in that: The ratio of the compound O-BDP, 4-formyl-1-methylpyrrole iodide, piperidine, and acetic acid is 1:2~3:2~4:0.1~0.

2.

7. The preparation method according to claim 5, characterized in that: The heating reaction is carried out at a temperature of 120~130℃ for 1~3 hours.

8. The application of the TPA-AN and pyridine conjugated modified BODIPY fluorescent probe as described in claim 1 in the preparation of AD diagnostic drugs.

9. The application of the TPA-AN and pyridine conjugated modified BODIPY fluorescent probe as described in claim 1 in the preparation of photodynamic therapy drugs.

10. The use of the TPA-AN and pyridine conjugated modified BODIPY fluorescent probe as described in claim 1 in the preparation of antibacterial drugs for photodynamic therapy.

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