Preparation method and application of novel fluorescent macrocyclic compound

Abstract

The invention relates to the technical field of macrocyclic compound preparation, in particular to a preparation method and application of a novel fluorescent macrocyclic compound, and the preparation method comprises the following steps: synthesizing a dialdehyde substituted intermediate through Suzuki coupling reaction; reducing the obtained intermediate into benzyl alcohol, and converting the benzyl alcohol into benzyl chloride; the preparation method comprises the following steps: carrying out Friedel-Crafts reaction on benzyl chloride and 1, 4-bis (bromomethyl) benzene to obtain a monomer; under the catalysis of boron trifluoride diethyl etherate, cyclizing a monomer to obtain a bromine chain substituted macrocyclic compound; a bromine chain substituted macrocyclic compound reacts with trimethylamine in ethanol to obtain the water-soluble fluorescent macrocyclic compound, and the novel fluorescent macrocyclic compound is designed and synthesized, so that singlet oxygen can be generated in vitro, an antibacterial drug can be effectively wrapped, and bacterial imaging can be carried out. The macrocyclic compound enhances the curative effect of the antibacterial drug through the synergistic effect, overcomes the problems of drug resistance and drug effect decline of the traditional antibacterial agent, and provides a new strategy for antibacterial treatment.

Description

Technical Field

[0001] The present invention relates to the technical field of the preparation of macrocyclic compounds, and particularly to a preparation method and application of a novel fluorescent macrocyclic compound. Background Art

[0002] The effectiveness of traditional antibacterial drugs is restricted by bacterial drug resistance, and the research and development of new antibacterial strategies have become an important topic in the current medical field. In recent years, researchers have begun to explore enhancing antibacterial effects through non-traditional means. One promising strategy is to utilize reactive oxygen species (ROS) and their synergistic effects with antibacterial drugs. Singlet oxygen ( 1 O 2 ) is a highly reactive oxygen molecule, usually generated by chemical reactions or photocatalytic reactions. It has strong oxidizing ability and can destroy the cell membranes, DNA and other important cellular components of bacteria, thereby inhibiting or killing microorganisms. 1 O 2 The widespread application of 1 O 2 is also limited by its instability and low generation efficiency. Therefore, how to efficiently generate 1 O 2 in a specific environment and cooperate with antibacterial drugs to achieve stronger antibacterial effects has become a current research hotspot. Existing research mainly focuses on the combined application of photosensitive materials and antibacterial drugs, while the

[0003] synergistic mechanism and efficiency between 1 O 2 and drugs still remain an issue that needs to be explored in depth. Summary of the Invention

[0004] The purpose of the present invention is to provide a preparation method and application of a novel fluorescent macrocyclic compound to solve the problems raised in the above background art.

[0005] To achieve the above purpose, the present invention provides the following technical solutions: A novel fluorescent macrocyclic compound, and the macrocyclic compound is [2]TpB-NExP6 with the following structure: ; Among them, the macrocyclic compound can generate singlet oxygen under light illumination conditions and has a fluorescence imaging function.

[0006] As a further solution of the present invention: the fluorescence emission peak of the macrocyclic compound is located at 525 nm, presenting green fluorescence and being suitable for bacterial imaging.

[0007] A preparation method of a novel fluorescent macrocyclic compound according to the above, comprising the following steps: Step 1: Using 4,7-dibromo-2,1,3-benzothiadiazole and 4-formylphenylboronic acid as raw materials, a dialdehyde-substituted intermediate is synthesized through a Suzuki coupling reaction; Step 2: The intermediate obtained in Step 1 is reduced to benzyl alcohol, and then subjected to a chlorination reaction to obtain benzyl chloride; Step 3: Benzyl chloride and 1,4-bis(bromomethyl)benzene are subjected to a Friedel-Crafts reaction under the catalysis of aluminum trichloride to obtain a monomer; Step 4: Under the catalysis of boron trifluoride diethyl ether, the monomer is cyclized to obtain a bromine-chain-substituted macrocyclic compound; Step 5: The bromine-chain-substituted macrocyclic compound reacts with trimethylamine in ethanol to obtain a water-soluble fluorescent macrocyclic compound.

[0008] As a further solution of the present invention: in Step 1, the solvent for the Suzuki coupling reaction is a mixed solution of tetrahydrofuran and water, the reaction temperature is 85 °C, and the reaction time is 48 hours.

[0009] As a further solution of the present invention: in Step 5, the reaction temperature is 90 °C, and the reaction time is 24 hours.

[0010] An application of a novel fluorescent macrocyclic compound according to the above in an antibacterial drug carrier, wherein the macrocyclic compound can encapsulate an antibacterial drug through host-guest interaction and release singlet oxygen under light illumination conditions to achieve a synergistic antibacterial effect.

[0011] As a further solution of the present invention: the antibacterial drug is cefotaxime sodium, and the complexation ratio of the macrocyclic compound to cefotaxime sodium is 1:1.

[0012] An application of a novel fluorescent macrocyclic compound according to the above in bacterial imaging, wherein the macrocyclic compound can target Gram-positive bacteria and Gram-negative bacteria and achieve bacterial imaging through green fluorescence.

[0013] An antibacterial composition comprising the novel fluorescent macrocyclic compound described above, and the composition can significantly reduce the minimum inhibitory concentration of bacteria under light illumination conditions.

[0014] Compared with the prior art, the beneficial effects of the present invention are: In the present invention, the fluorescence emission peak of the water-soluble macrocycle (such as the macrocycle containing benzothiadiazole) is located at 525 nm, showing green fluorescence, and can be used for bacterial imaging.

[0015] Under normal temperature and pressure, the macrocycle can be excited by an artificial light source to generate 1 O 2 .

[0016] In the present invention, compared with the monomer, the macrocycle compound has a stronger ability to generate 1 O 2 .

[0017] The macrocycle compound has good biocompatibility and can undergo host-guest interactions with drug molecules, meeting the principles of green chemistry and laying a foundation for the application of a series of compounds in this field. Brief Description of the Drawings

[0018] Figure 1 It is a synthetic route diagram of the macrocycle compound in the embodiment of the present invention.

[0019] Figure 2 It is the nuclear magnetic resonance of the bromine-chain substituted macrocycle compound in the embodiment of the present invention.

[0020] Figure 3 It is the nuclear magnetic resonance of the water-soluble macrocycle compound in the embodiment of the present invention.

[0021] Figure 4 It is a schematic diagram of the study on the aggregation-induced emission properties of [2]TpB-NExP6 in the embodiment of the present invention; Among them, on the left is the fluorescence spectrum diagram at different acetone volumes in the mixed system of acetone and chloroform, and on the right is the broken line diagram of the fluorescence intensity change trend at 523 nm at different acetone volumes.

[0022] Figure 5 It is a schematic diagram of the study on the property of generating singlet oxygen by M-N and [2]TpB-NExP6 in the embodiment of the present invention; Among them, on the left is the change of the ultraviolet absorption peak of ABDA with time under illumination and in the presence of [2]TpB-NExP6, and on the right is the broken line diagram of the absorption peak change trend of ABDA alone and ABDA in the presence of the macrocycle [2]TpB-NExP6 and the monomer M-N with time.

[0023] Figure 6 It is a schematic diagram of the JobPlot experiment of [2]TpB-NExP6 and cefazolin sodium in the embodiment of the present invention; Among them, (a) is the fluorescence emission spectrum diagram at different host-guest ratios, and (b) is the JobPlot curve diagram drawn according to the fluorescence intensity.

[0024] Figure 7 Schematic diagram of the MIC value test of the macrocyclic host-guest for three bacteria in light and darkness in the embodiment of the present invention; Among them, Escherichia coli is on the left, Staphylococcus aureus is in the middle, and methicillin-resistant Staphylococcus aureus is on the right. Specific implementation manners

[0025] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

[0026] The following describes the specific implementation of the present invention in detail in conjunction with specific embodiments.

[0027] Please refer to Figures 1 - 7 , a novel fluorescent macrocyclic compound provided by an embodiment of the present invention, and the macrocyclic compound is [2]TpB-NExP6 having the following structure: ; Among them, the macrocyclic compound can generate singlet oxygen under light conditions and has a fluorescence imaging function.

[0028] The fluorescence emission peak of the macrocyclic compound is located at 525 nm, presenting green fluorescence, which is suitable for bacterial imaging.

[0029] In an embodiment of the present invention, a preparation method of the novel fluorescent macrocyclic compound according to the above includes the following steps: Step 1: Using 4,7-dibromo-2,1,3-benzothiadiazole and 4-formylphenylboronic acid as raw materials, synthesize a dialdehyde-substituted intermediate through a Suzuki coupling reaction; Step 2: Reduce the intermediate obtained in Step 1 to benzyl alcohol, and then carry out a chlorination reaction to obtain benzyl chloride; Step 3: Carry out a Friedel-Crafts reaction on benzyl chloride and 1,4-bis(bromomethyl)benzene under the catalysis of aluminum trichloride to obtain a monomer; Step 4: Under the catalysis of boron trifluoride diethyl ether, cyclize the monomer to obtain a bromine-chain-substituted macrocyclic compound; Step 5: React the bromine-chain-substituted macrocyclic compound with trimethylamine in ethanol to obtain a water-soluble fluorescent macrocyclic compound.

[0030] In Step 1, the solvent for the Suzuki coupling reaction is a mixed solution of tetrahydrofuran and water, the reaction temperature is 85 °C, and the reaction time is 48 hours.

[0031] In step 5, the reaction temperature is 90 °C and the reaction time is 24 hours.

[0032] In one embodiment of the present invention, there is provided an application of the novel fluorescent macrocyclic compound as described above in an antibacterial drug carrier. The macrocyclic compound can encapsulate the antibacterial drug through host-guest interaction and release singlet oxygen under light irradiation to achieve a synergistic antibacterial effect.

[0033] The antibacterial drug is cefotaxime sodium, and the complexation ratio of the macrocyclic compound to cefotaxime sodium is 1:1.

[0034] In one embodiment of the present invention, there is provided an application of the novel fluorescent macrocyclic compound as described above in bacterial imaging. The macrocyclic compound can target Gram-positive bacteria and Gram-negative bacteria and achieve bacterial imaging through green fluorescence.

[0035] In one embodiment of the present invention, there is provided an antibacterial composition comprising the novel fluorescent macrocyclic compound as described above. The composition can significantly reduce the minimum inhibitory concentration (MIC) of bacteria under light irradiation.

[0036] Through the characterization of the basic physical and chemical properties of the synthesized macrocyclic compound, the macrocycle with a fluorescent group is more suitable for subsequent antibacterial experiments. Therefore, a water-soluble macrocycle was obtained through post-modification to meet the requirement of generating 1 O 2 In this test, a light source, PBS (phosphate buffer, pH = 7.4), and 9,10-anthracenediyl-bis(methylene)dicarboxylic acid (ABDA), etc. need to be prepared.

[0037] First, dissolve the macrocyclic compound in PBS (phosphate buffer, pH = 7.4) to prepare a 0.1 mM solution. Add the ABDA (9,10-anthracenediyl-bis(methylene)dicarboxylic acid) probe (0.01 mM), and then place the solution under visible light irradiation. Monitor the change in absorbance with a UV spectrum at regular intervals. Since 1 O 2 can react with ABDA to photobleach and generate the corresponding endoperoxide. By measuring the absorbance value of the reaction solution at a specific wavelength (such as 400 nm) and according to its change trend, the ability of the macrocyclic compound to generate 1 O 2 can be tested. The light intensity and the concentration of the macrocycle solution need to be controlled well during the whole experimental process to ensure the reliability of the data.

[0038] The host-guest interaction between the macrocyclic compound and the antibacterial drug cefotaxime sodium (n / n = 1:1) was demonstrated by NMR characterization. The fluorescence spectra of the macrocyclic compound and cefotaxime sodium at different ratios were measured, and a scatter plot of their fluorescence intensity against the ratio was plotted to obtain a JobPlot curve, from which the complexation ratio was calculated to be 1:1. Then, fluorescence titration was carried out to obtain the complexation constant to prove the interaction between the macrocyclic compound and the guest molecule.

[0039] Therefore, the synthesis of novel macrocyclic compounds can effectively generate 1 O 2 and encapsulate antibacterial drugs, thereby achieving a synergistic antibacterial effect. Through this novel treatment strategy, the dosage of traditional antibacterial drugs can be reduced, thus reducing side effects. By designing photosensitive macrocyclic compounds, they can efficiently generate 1 O 2 under specific light conditions, thereby overcoming the problems of low generation efficiency and poor stability in the prior art. 1 O 2 The structural design of the macrocyclic compound endows it with good drug-loading capacity, enabling it to effectively encapsulate antibacterial drugs and release 1 O 2 in a local area, thereby enhancing the antibacterial effect and providing an innovative and multi-dimensional strategy for antibacterial treatment.

[0040] The present invention synthesizes the macrocyclic compound shown in Figure 1 using a five-step synthesis method. The bromine-chain substituted macrocycle can be further reacted with trimethylamine in ethanol to obtain a water-soluble fluorescent macrocycle (such as [2]TpB-NExP6 containing benzothiadiazole). We also synthesized a water-soluble monomer (such as M-N) for comparison with the macrocycle.

[0041] Example 1: Taking the macrocyclic compound [2]TpB-NExP6 constructed with benzothiadiazole (a macrocyclic compound containing benzothiadiazole, and the preparation of a water-soluble macrocycle is achieved by quaternary ammonium salt modification of it, and the specific structural formula is as shown in Figure 1 ) as an example: Weigh 5 g of 4,7-dibromo-2,1,3-benzothiadiazole, 7.6 g of 4-formylphenylboronic acid, 0.5 g of Pd(PPh 3 ) 4 0.5 g, and Na 2 CO 313 g. The dialdehyde-substituted compound was obtained by Suzuki coupling reaction in a solution of tetrahydrofuran / water (40 mL / 10 mL) at 85 °C for 48 h with a yield of 96%. It was further reduced by sodium borohydride to obtain benzyl alcohol, which was reacted with thionyl chloride in a solvent at 65 °C for 24 h to obtain benzyl chloride. Both of these two steps were equimolar reactions. Then, it reacted with 1,4-bis(bromomethyl)benzene and aluminum trichloride in dichloromethane through Friedel-Crafts reaction to obtain the monomer with a yield of 26%. Finally, the macrocyclic compound was obtained by catalysis with boron trifluoride diethyl ether with a yield of 30%, ( Figure 2 ). Subsequently, the brominated chain macrocyclic compound was dispersed in ethanol, and trimethylamine was added and reacted at 90 °C for 24 h to obtain the target compound with a yield of 96%, ( Figure 3 ). Its fluorescence was tested. By changing the content of acetone solvent (poor solvent) in the aqueous solution of the macrocyclic compound, it was proved that the target compound has green fluorescence and aggregation-induced emission properties ( Figure 4 ). And the host-guest JobPlot curve was obtained by processing the fluorescence intensity data at different host-guest ratios, proving the 1:1 complexation mode between the host and the guest ( Figure 6 ).

[0042] Example 2: Based on [2]TpB-NExP6 1 O 2 Production capacity test: First, the prepared aqueous solution of the macrocyclic compound was mixed evenly with the ABDA probe and transferred to a fluorescence cuvette. A visible light source was introduced, and the temperature of the system was controlled to remain at room temperature. UV tests were performed at regular intervals. Under the same experimental conditions, a control test was carried out with the monomer to produce 1 O 2 capacity. It was found that the macrocycle has certain advantages ( Figure 5 ).

[0043] Example 3: Antibacterial experiments were carried out on Staphylococcus aureus, Escherichia coli, and methicillin-resistant Staphylococcus aureus. First, the minimum inhibitory concentration test (MIC) of the host-guest complex was carried out under the conditions of light and without light. The bacteria were placed in a 96-well plate, and experiments were carried out on the three bacteria in two groups of light treatment and dark treatment. The experimental results showed that the minimum inhibitory concentration (MIC) values obtained under the condition of light were all smaller than those in the group without light. For example, under the condition of light, the MIC value of the macrocycle host-guest complex against Escherichia coli ( E.coli ) was 1.5 μM, and the MIC in the dark environment was 6.25 μM. The MIC values of Staphylococcus aureus ( S. aureus ) and methicillin-resistant Staphylococcus aureus (MRSA) were 31.25 μM and 250 μM respectively under light and dark conditions ( Figure 7 ). It was proved that light generates 1 O 2Its important role in the antibacterial process. In addition, the quaternary ammonium salt macrocycle can target Escherichia coli and Staphylococcus aureus, and as a fluorescent probe, it can produce bright green fluorescence after binding to bacteria, realizing the fluorescence imaging of Gram-negative and Gram-positive bacteria.

[0044] In summary, by designing a new type of photosensitive fluorescent macrocyclic compound, the present invention can generate 1 O 2 under certain conditions and effectively encapsulate antibacterial drugs therein. This macrocyclic compound can not only bind to antibacterial drugs through host-guest interaction, but also further enhance the antibacterial effect of the drugs by generating 1 O 2 The high oxidizing property of 1 O 2 can directly damage biological macromolecules such as the cell membrane and DNA of bacteria, thereby achieving the effect of rapid sterilization. Compared with traditional single antibacterial drug treatment, this synergistic effect can effectively improve the antibacterial effect and reduce the generation of drug-resistant bacteria, especially in the treatment of complex infections and drug-resistant bacteria infections, which has important application prospects. It provides a theoretical basis for the development of new antibacterial strategies and a new direction for the innovative design of future antibacterial systems.

[0045] It should be noted that in the present invention, it should be understood that although this specification is described according to the embodiments, not each embodiment only contains an independent technical solution. This narrative way of the specification is only for clarity. Those skilled in the art should regard the specification as a whole, and the technical solutions in each embodiment can also be appropriately combined to form other embodiments that can be understood by those skilled in the art.

Claims

1. A novel fluorescent macrocyclic compound, characterized in that: The macrocyclic compound is [2]TpB-NExP6 having the following structure: ； Among them, macrocyclic compounds can produce singlet oxygen under light conditions and have fluorescence imaging capabilities.

2. The novel fluorescent macrocyclic compound according to claim 1, characterized in that: The fluorescence emission peak of the macrocyclic compound is located at 525 nm, exhibiting green fluorescence, and is suitable for bacterial imaging.

3. A method for preparing the novel fluorescent macrocyclic compound according to claim 1, characterized in that: The following steps are involved: Step 1, using 4,7-dibromo-2,1,3-benzothiadiazole and 4-formylphenylboronic acid as raw materials, synthesizing a dialdehyde-substituted intermediate through a Suzuki coupling reaction; Step 2, reducing the intermediate obtained in step 1 to benzyl alcohol, and then subjecting the intermediate to chlorination to obtain benzyl chloride; Step 3, performing Friedel-Crafts reaction on benzyl chloride and 1,4-di(bromomethyl)benzene under the catalysis of aluminum chloride to obtain a monomer; Step 4, cyclizing the monomer under the catalysis of boron trifluoride ether to obtain a bromine chain substituted macrocyclic compound; Step 5: reacting the bromine chain-substituted macrocyclic compound with trimethylamine in ethanol to obtain a water-soluble fluorescent macrocyclic compound.

4. The method for preparing the novel fluorescent macrocyclic compound according to claim 3, characterized in that: In step 1, the solvent for the Suzuki coupling reaction is a mixed solution of tetrahydrofuran and water, the reaction temperature is 85° C., and the reaction time is 48 hours.

5. The method for preparing the novel fluorescent macrocyclic compound according to claim 3, characterized in that: In step 5, the reaction temperature is 90° C. and the reaction time is 24 hours.

6. Use of the novel fluorescent macrocyclic compound according to claim 1 in an antibacterial drug carrier, characterized in that: The macrocyclic compound can encapsulate the antibacterial drug through host-guest interaction and release singlet oxygen under light conditions to achieve a synergistic antibacterial effect.

7. Use of the novel fluorescent macrocyclic compound in an antibacterial drug carrier according to claim 6, characterized in that: The antibacterial drug is cefuroxime sodium, and the complexation ratio of the macrocyclic compound to cefuroxime sodium is 1:

1.

8. Use of the novel fluorescent macrocyclic compound according to claim 1 in bacterial imaging, characterized in that: The macrocyclic compound can target Gram-positive and Gram-negative bacteria and achieve bacterial imaging through green fluorescence.

9. An antibacterial composition, characterized in that The composition comprises the novel fluorescent macrocyclic compound as claimed in claim 1, and the composition can significantly reduce the minimum inhibitory concentration of bacteria under illumination conditions.

Images