A benzopyridazine compound and its preparation method and application

Abstract

The present invention relates to the technical field of biochemical materials, and provides a benzopyridazine compound and a preparation method and application thereof. The benzopyridazine compound provided by the present invention has a structure shown in Formula I, has an aggregation-induced emission (AIE) effect, can emit stronger fluorescence in an aggregated state, and has a strong ability to produce reactive oxygen species (ROS). Under illumination conditions, a large amount of reactive oxygen species can be generated to photodynamically kill bacteria; the benzopyridazine compound provided by the present invention has good biocompatibility and good safety, and has broad application prospects in photodynamic antibacterial chemotherapy.

Description

Technical Field

[0001] The present invention relates to the technical field of chemical materials, and in particular to a benzopyridazine compound and a preparation method and application thereof. Background Art

[0002] As antibiotic resistance is becoming more and more serious worldwide, the research on new anti-infection methods is urgent. Photodynamic antimicrobial chemotherapy (PACT) is a new anti-infection therapy based on the principle of photodynamic therapy, which uses photosensitizers and light to kill bacteria. This therapy has the advantages of safety, high efficiency, easy implementation, repeatable administration, good biocompatibility and synergy, and is very promising in antibacterial.

[0003] Photosensitizers are the core substances of photodynamic antibacterial chemotherapy, and the development of photosensitizers is the current research focus. At present, common photosensitizers include porphyrin compounds, phenothiazine compounds, riboflavin and curcumin, etc., all of which have certain photodynamic bactericidal properties. However, these traditional photosensitizers are molecules with large π conjugation, strong hydrophobicity, and easy to aggregate in aqueous solution, resulting in a decrease in the ability to produce reactive oxygen, which greatly reduces the antibacterial activity.

[0004] Benzopyridazine compounds are a class of derivatives with benzopyridazine as the core. The pyridazine structure has the characteristics of strong electron deficiency, so benzopyridazine compounds are often used to synthesize optoelectronic materials. However, there are very few reports on the application of benzopyridazine compounds in photodynamic antibacterial. Summary of the invention

[0005] In view of this, the present invention provides a benzopyridazine compound and a preparation method and application thereof. The benzopyridazine compound provided by the present invention has a high efficiency in generating reactive oxygen species (ROS), can perform efficient photodynamic killing of Gram-positive bacteria, has high antibacterial activity, and is safe.

[0006] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0007] A benzopyridazine compound having a structure shown in Formula I:

[0008]

[0009] The present invention also provides a method for preparing the benzopyridazine compound described in the above scheme, comprising the following steps:

[0010] A compound having a structure shown in Formula II, 4,4'-dimethoxydiphenylamine, a palladium catalyst, tri-tert-butylphosphine tetrafluoroborate, an organic base and a benzene solvent are mixed to carry out a Buchwald-Hartwig coupling reaction to obtain a compound having a structure shown in Formula III;

[0011]

[0012] The compound having the structure shown in formula III is mixed with diphenylacetylene, an iridium catalyst, an inorganic base, a compound containing a non-coordinating tetrafluoroborate and an organic solvent, and a cyclization reaction is carried out under a protective atmosphere to obtain the benzopyridazine compound.

[0013] Preferably, the molar ratio of the compound having the structure shown in Formula II to 4,4'-dimethoxydiphenylamine is 1:1.8-2.2; the molar ratio of the compound having the structure shown in Formula II, palladium catalyst, tri-tert-butylphosphine tetrafluoroborate and organic base is 1:0.03-0.05:0.08-0.12:2.5-3.5.

[0014] Preferably, the temperature of the Buchwald-Hartwig coupling reaction is 95-105° C. and the time is 20-25 h.

[0015] Preferably, the iridium catalyst is dichloro(pentamethylcyclopentadiene)iridium(III) dimer; the inorganic base is copper acetate; and the compound containing non-coordinated tetrafluoroborate is silver tetrafluoroborate.

[0016] Preferably, the molar ratio of the compound having the structure shown in Formula II to diphenylacetylene is 1:1.2-1.8; the molar ratio of the compound having the structure shown in Formula II, the iridium catalyst, the inorganic base and the compound containing non-coordinating tetrafluoroborate is 1:0.05-0.2:0.05-0.2:1.5-2.5.

[0017] Preferably, the cyclization reaction is carried out at a temperature of 120 to 135° C. and for a time of 20 to 26 hours.

[0018] Preferably, the method for preparing the compound having the structure shown in Formula II comprises the following steps:

[0019] The p-bromoaniline, chloroalkane, potassium permanganate and ferrous sulfate are mixed for oxidative dimerization reaction to obtain a compound having a structure shown in formula II.

[0020] Preferably, the molar ratio of potassium permanganate to ferrous sulfate is 1:1-1.2; the molar ratio of p-bromoaniline to potassium permanganate is 1:2.8-3.5; the temperature of the oxidative dimerization reaction is 35-45°C, and the time is 5-8h.

[0021] The present invention also provides the use of the benzopyridazine compound described in the above scheme or the benzopyridazine compound prepared by the preparation method described in the above scheme in the preparation of photodynamic antibacterial drugs.

[0022] The present invention provides a benzopyridazine compound having a structure shown in Formula I. The benzopyridazine compound provided by the present invention has a benzopyridazine salt structure and a tetrafluoroborate anion in its molecular structure, which can make the compound molecule positively charged, while the bacterial surface is negatively charged. Due to electrostatic interaction, the compound can be effectively combined with the bacteria, which establishes a foundation for subsequent efficient killing; and the present invention introduces a methoxyphenyl group into the structure of the benzopyridazine compound, which is a strong electron-donating group and can rotate rapidly, and can enhance the interaction between the donor and the acceptor in the molecule, and at the same time, the molecule has an aggregation-induced emission (AIE) property, so that the molecule still has the ability to generate active oxygen in an aggregated state, which is conducive to the generation of active oxygen in an aqueous solution, and the benzopyridazine compound has better antibacterial activity.

[0023] In addition, the benzopyridazine compounds provided by the present invention also have obvious aggregation-induced emission effect (AIE). Traditional photosensitizers will undergo fluorescence quenching in the aggregated state, the ROS production will decrease, and the antibacterial effect will deteriorate. The benzopyridazine compounds provided by the present invention have aggregation-induced emission effect, show enhanced fluorescence in the aggregated state, can continuously generate ROS, and have excellent photodynamic antibacterial effect.

[0024] The results of the examples show that when the benzopyridazine compounds of the present invention are mixed with the active oxygen scavenger DCFH, under white light irradiation, the fluorescence intensity of DCFH gradually increases with the increase of illumination time and the enhancement rate is relatively fast, indicating that the benzopyridazine compounds of the present invention have a high efficiency in generating active oxygen; when the benzopyridazine compounds provided by the present invention are used for photodynamic antibacterial, the sterilization rate can reach more than 95% at a concentration of 0.6 μM; at the same time, the benzopyridazine compounds provided by the present invention also have good biocompatibility and can be used to construct safe and efficient photodynamic antibacterial drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 The fluorescence intensity test results of TPCL-MDPA in DMSO / glycerol mixed solutions with different glycerol volume fractions;

[0026] Figure 2 The fluorescence emission spectrum of the mixed solution of TPCL-MDPA and DCFH changes with the illumination time;

[0027] Figure 3 The photodynamic killing of Staphylococcus aureus by TPCL-MDPA at different concentrations;

[0028] Figure 4 The phototoxicity and dark toxicity of different concentrations of TPCL-MDPA on normal cells LO2. DETAILED DESCRIPTION

[0029] The present invention provides a benzopyridazine compound having a structure shown in Formula I:

[0030]

[0031] The present invention also provides a method for preparing the benzopyridazine compound described in the above scheme, comprising the following steps:

[0032] A compound having a structure shown in Formula II, 4,4'-dimethoxydiphenylamine, a palladium catalyst, tri-tert-butylphosphine tetrafluoroborate, an organic base and a benzene solvent are mixed to perform a Buchwald-Hartwig Cross Coupling Reaction to obtain a compound having a structure shown in Formula III;

[0033]

[0034] The compound having the structure shown in formula III is mixed with diphenylacetylene, an iridium catalyst, an inorganic base, a compound containing a non-coordinating tetrafluoroborate and an organic solvent, and a cyclization reaction is carried out under a protective atmosphere to obtain the benzopyridazine compound.

[0035] In the present invention, the method for preparing the compound having the structure shown in Formula II comprises the following steps:

[0036] The p-bromoaniline, chloroalkane, potassium permanganate and ferrous sulfate are mixed for oxidative dimerization reaction to obtain a compound having a structure shown in formula II.

[0037] In the present invention, the ferrous sulfate is preferably ferrous sulfate heptahydrate; the molar ratio of potassium permanganate to ferrous sulfate is preferably 1:1-1.2; the molar ratio of p-bromoaniline to potassium permanganate is preferably 1:2.8-3.5, more preferably 1:3; the chloroalkane is preferably dichloromethane; the present invention preferably first mixes p-bromoaniline and chloroalkane solvent to obtain a p-bromoaniline solution, grinds and mixes potassium permanganate and ferrous sulfate, and adds the resulting mixture to the p-bromoaniline solution. In the present invention, the temperature of the oxidative dimerization reaction is preferably 35-45°C, more preferably 40°C, and the time is preferably 5-8h, more preferably 6h.

[0038] After the oxidative dimerization reaction is completed, the present invention preferably performs post-treatment on the obtained product liquid; the post-treatment preferably includes the following steps: cooling the product liquid obtained by the oxidative dimerization reaction and diluting it with chloroalkane, filtering, concentrating and column chromatography purification of the dilution in sequence to obtain a compound having a structure shown in Formula II. In the present invention, the type of chloroalkane used for dilution is consistent with the above scheme, and will not be repeated here; the filtration is preferably performed using a diatomaceous earth pad, and after the filtration is completed, the diatomaceous earth pad is preferably rinsed with a chloroalkane solvent, the rinse liquid and the filtrate are combined, and the obtained mixed liquid is subsequently concentrated; the concentration is preferably vacuum concentration; the eluent for column chromatography purification is preferably petroleum ether; the compound having a structure shown in Formula II obtained after purification is an orange-yellow solid.

[0039] In the present invention, the reaction formula of the oxidative dimerization reaction is as follows:

[0040]

[0041] After obtaining the compound having the structure shown in Formula II, the present invention mixes the compound having the structure shown in Formula II, 4,4'-dimethoxydiphenylamine, a palladium catalyst, tri-tert-butylphosphine tetrafluoroborate, an organic base and a benzene solvent to perform a Buchwald-Hartwig coupling reaction to obtain a compound having the structure shown in Formula III. In the present invention, the palladium catalyst is preferably tri(dibenzylideneacetone)dipalladium; the organic base is preferably sodium tert-butoxide, and the role of the organic base is to neutralize the HBr produced during the reaction; the role of tri-tert-butylphosphine tetrafluoroborate is a ligand of the palladium catalyst; the benzene solvent is preferably toluene; the molar ratio of the compound having the structure shown in Formula II and 4,4'-dimethoxydiphenylamine is preferably 1:1.8 to 2.2, more preferably 1:2; the compound having the structure shown in Formula II, the palladium catalyst, tri-tert-butylphosphine tetrafluoroborate, an organic base and a benzene solvent are preferably mixed to perform a Buchwald-Hartwig coupling reaction to obtain a compound having the structure shown in Formula III. The molar ratio of tert-butylphosphine tetrafluoroborate to the organic base is 1:0.03-0.05:0.08-0.12:2.5-3.5, and more preferably 1:0.05:0.1:3; the present invention has no special requirements on the amount of the benzene solvent, as long as the Buchwald-Hartwig coupling reaction can proceed smoothly; the temperature of the Buchwald-Hartwig coupling reaction is preferably 100° C., and the time is preferably 24 hours; the Buchwald-Hartwig coupling reaction is preferably carried out under nitrogen protection conditions.

[0042] After the Buchwald-Hartwig coupling reaction is completed, the present invention preferably performs post-treatment on the obtained product liquid; the post-treatment preferably includes the following steps: cooling the product liquid obtained by the Buchwald-Hartwig coupling reaction to room temperature and then pouring it into methanol for first precipitation, filtering to obtain a first precipitate; washing the first precipitate with methanol, disodium ethylenediaminetetraacetic acid aqueous solution, water and n-hexane in sequence, and then performing a second precipitation in a chloroform-methanol mixed solution, filtering to obtain a second precipitate, and vacuum drying the second precipitate to obtain a compound having a structure shown in formula II; the temperature of the first precipitation is preferably 25-35°C, and the time is preferably 5-8min; the temperature of the second precipitation is preferably 25-35°C, and the time is preferably 8-10min; the volume ratio of chloroform and methanol in the chloroform-methanol mixed solution is preferably 20-25:1; the compound having a structure shown in formula II is a dark red powder.

[0043] In the present invention, the reaction formula of the Buchwald-Hartwig coupling reaction is as follows:

[0044]

[0045] After obtaining the compound having the structure shown in formula III, the present invention mixes the compound having the structure shown in formula III with diphenylacetylene, an iridium catalyst, an inorganic base, a compound containing non-coordinating tetrafluoroborate and an organic solvent, and performs a cyclization reaction under a protective atmosphere to obtain a benzopyridazine compound having the structure shown in formula I. In the present invention, the iridium catalyst is preferably dichloro(pentamethylcyclopentadiene)iridium(III) dimer; the inorganic base is preferably copper acetate; the compound containing non-coordinating tetrafluoroborate is preferably silver tetrafluoroborate; the molar ratio of the compound having the structure shown in formula II to diphenylacetylene is preferably 1:1.2-1.8, more preferably 1:1.5; the molar ratio of the compound having the structure shown in formula II, the iridium catalyst, the inorganic base and the compound containing non-coordinating tetrafluoroborate is preferably 1:0.05-0.2:0.05-0.2:1.5-2.5, more preferably 1:0.1:0.1:2.

[0046] In the present invention, the temperature of the cyclization reaction is preferably 120-135°C, more preferably 125-130°C, and the time of the cyclization reaction is preferably 20-26h, more preferably 24h; the cyclization reaction is preferably carried out in a thick-walled reaction bottle.

[0047] In a specific embodiment of the present invention, it is preferred to first add a compound having a structure shown in Formula III, diphenylacetylene, an iridium catalyst and an inorganic base into a thick-walled reaction bottle, then evacuate the reaction bottle, add a compound containing a non-coordinating tetrafluoroborate and an organic solvent under nitrogen protection, then tighten the mouth of the thick-walled reaction bottle, evacuate again, and then react at the cyclization reaction temperature.

[0048] In the present invention, the reaction formula of the cyclization reaction is as follows:

[0049]

[0050] In the present invention, after the cyclization reaction is completed, it is preferred that the obtained product liquid is post-treated, and the post-treatment preferably includes the following steps: the product liquid is filtered, concentrated, and column chromatographed in sequence to obtain a benzopyridazine compound having a structure shown in Formula I; in the present invention, preferably after the reaction is completed, the product liquid is cooled to room temperature, and then filtered using diatomaceous earth, and the diatomaceous earth is rinsed with dichloromethane after the filtration is completed, and the rinse liquid and the filtrate are combined, and the combined liquid is used as the final subsequent concentration; the column chromatography eluent is preferably a mixture of dichloromethane and methanol, and the volume ratio of dichloromethane to methanol in the mixture is preferably 20: 1; after the column chromatography is completed, the present invention preferably removes the solvent in the column chromatography product. The present invention does not specifically limit the method of removing the solvent, and a conventional method of removing the solvent can be used, such as rotary evaporation.

[0051] The present invention also provides the use of the benzopyridazine compounds described in the above scheme in the preparation of photodynamic antibacterial drugs; in the present invention, the benzopyridazine compounds are preferably used as photosensitizers; the photodynamic antibacterial drugs are preferably anti-Gram-positive bacteria drugs; the Gram-positive bacteria are preferably Staphylococcus aureus; the excitation wavelength of the photodynamic antibacterial drugs is preferably 450-500nm.

[0052] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0053] Example 1

[0054] (1) Preparation of the compound having the structure shown in Formula II

[0055] In a 250 mL eggplant-shaped bottle, p-bromoaniline (1 mmol, 172.0 mg) was dissolved in dichloromethane, and then KMnO 4Solid (3 mmol, 474.1 mg) and FeSO 4 7H 2 O solid (3mmol, 834.0mg) was mixed and ground as a catalyst and added to an eggplant-shaped bottle and stirred at 40°C for 6h. After the reaction was completed, the mixture was cooled and diluted with dichloromethane. The mixture was filtered through a diatomaceous earth pad, and the diatomaceous earth pad was washed with dichloromethane (30mL). The filtrate was concentrated under vacuum. Petroleum ether was used as an eluent, and the obtained concentrate was subjected to column chromatography to obtain an orange-yellow solid, which was a compound having the structure shown in Formula II. Yield: 70%. 1 H NMR (500 MHz, CDCl 3 )δ7.82(d,J=8.7Hz,4H),7.68(d,J=8.7Hz,4H).

[0056] (2) Preparation of the compound having the structure shown in Formula III

[0057] In a 50mL two-necked round-bottom flask, a mixture of a compound having a structure shown in Formula II (1mmol, 340.0mg), 4,4'-dimethoxydiphenylamine (2mmol, 458.5mg), tri(dibenzylideneacetone)dipalladium (0.05mmol, 45.8mg), tri-tert-butylphosphine tetrafluoroborate (0.1mmol, 29.0mg), and sodium tert-butoxide (3mmol, 288.3mg) was added and dissolved in toluene (10mL), and reacted at 100°C in a nitrogen atmosphere for 24h. After cooling to room temperature, the reaction mixture was poured into methanol (200mL). The precipitate was separated by filtration, washed with methanol, ethylenediaminetetraacetic acid disodium salt aqueous solution, water and n-hexane, reprecipitated from chloroform / methanol, and vacuum dried to obtain a dark red powder, which is a compound having a structure shown in Formula III, with a yield of 94%. 1 H NMR (500 MHz, CDCl 3 )δ7.70(d,J=8.8Hz,4H),7.11(d,J=9.0Hz,8H),6.94(d,J=9.1Hz,4H),6.86(d,J=9.0Hz,8H),3.81(s,12H).

[0058] (2) Preparation of benzopyridazine compounds having the structure shown in Formula I

[0059] The compound having the structure shown in formula III (1 mmol, 636.7 mg), diphenylacetylene (1.5 mmol, 267.3 mg), dichloro(pentamethylcyclopentadiene)iridium(III) dimer ([Cp*IrCl 2 ] 2 , 0.1mmol, 79.7mg) and copper acetate (Cu(OAc)2 , 0.1mmol, 18.2mg) was added to a thick-walled reaction bottle, and then the reaction device was ventilated. Silver tetrafluoroborate (AgBF 4 , 2mmol, 389.3mg) and 1,2-dichloroethane (10mL), tighten the thick-walled reaction bottle, perform the evacuation operation again, and stir at 130°C for 24h. After the reaction is completed, cool to room temperature, filter with diatomaceous earth, rinse the diatomaceous earth with dichloromethane (30mL), and concentrate the filtrate. Use dichloromethane / methanol (20:1, v / v) as eluent to perform column chromatography on the mixture to obtain the target product as a purple solid. Yield: 49%. The structural identification data of the product are as follows:

[0060] 1 H NMR(500MHz,MeOD)δ8.22(d,J=9.7Hz,1H),7.72(dd,J=9.7,2.6Hz,1H),7.29-7.16(m,14H),7.12(dd,J=7.4,2.1Hz,2 H),7.03-6.98(m,8H),6.91(d,J=8.9Hz,4H),6.68(d,J=9.1Hz,2H),6.53(d,J=2.5Hz,1H),3.84(s,6H),3.80(s,6H). 13 C NMR (151 MHz, CDCl 3 )δ158.91,156.73,155.62,149.79,148.55,144.84,139.57,139.40,136.32,135.94,132.85,132.01,131.34,131.1 5,129.99,129.16,128.55,128.24,128.08,128.03,127.38,127.26,118.31,115.44,114.96,105.80,55.67,55.52.

[0061] According to the above characterization data, the structural formula of the obtained target product is shown in Formula I, which is denoted as TPCL-MDPA.

[0062] Performance Test:

[0063] (1) AIE property test of TPCL-MDPA: Glycerol is a high viscosity solvent. DMSO solution (1 mM) of TPCL-MDPA was added to DMSO / glycerol mixed solutions with different glycerol volume fractions to obtain TPCL-MDPA solutions with a concentration of 10 μM. The fluorescence intensity of TPCL-MDPA in the mixed solutions with different glycerol volume fractions was measured.

[0064] Figure 1 The fluorescence intensity test results of TPCL-MDPA in DMSO / glycerol mixed solutions with different glycerol volume fractions (the corresponding glycerol volume fractions from low to high are 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99%, and the excitation light wavelength is 390nm). Figure 1 It can be seen that with the increase of glycerol volume fraction, the fluorescence intensity of TPCL-MDPA gradually increases, indicating that TPCL-MDPA has AIE characteristics.

[0065] (2) Test of TPCL-MDPA's ability to produce active oxygen: prepare the DMSO stock solution of the sample to be tested, take 20 μL of the sample to be tested and 50 μL of DCFH, add them to a cuvette containing 2 mL of PBS buffer solution, so that the final test concentration of TPCL-MDPA is 10 μM and the final concentration of DCFH is 40 μM. Use FL-4700 fluorescence spectrometer for testing, set the excitation wavelength to 489 nm, first measure the fluorescence spectrum in the absence of light (0 min), and then measure the sample spectrum at the same interval (interval time is 5 s) to obtain the ratio of real-time luminescence intensity to initial luminescence intensity; at the same time, use a single DCFH solution (concentration of 40 μM) and a single TPCL-DMA solution (concentration of 10 μM) as controls. The results are as follows: Figure 2 shown.

[0066] Figure 2 The fluorescence emission spectrum of the mixed solution of TPCL-MDPA and DCFH changes with the illumination time. Figure 2 It can be seen that with the increase of illumination time, the fluorescence intensity of the mixed solution gradually increases, indicating that TPCL-MDPA has a strong ability to produce active oxygen.

[0067] (3) Test of the photodynamic killing ability of TPCL-MDPA on Staphylococcus aureus. The specific steps are as follows:

[0068] The Staphylococcus aureus strain was revived and inoculated into LB liquid medium and cultured in a shaker at 37°C and 220 r / min for 16 h. At this time, the concentration of Staphylococcus aureus in the bacterial suspension was about 2×10 9CFU / mL; 1 mL of cultured Staphylococcus aureus was taken, the culture medium was removed, and it was washed with PBS; different volumes of DMSO solution of TPCL-MDPA were added to make the final concentration of TPCL-MDPA 0 μM, 0.2 μM, 0.4 μM, and 0.6 μM, respectively. The bacterial suspension was incubated in a shaker at 37°C and 220 r / min for 10 min, and then the bacterial suspension was illuminated for 20 min using a solar simulator equipped with a 400 nm filter; after the illumination, the bacterial suspension was gradiently diluted to 1×10 7 The cells were plated on LB agar solid medium and cultured in a 37°C incubator for 16 h before colony counting and survival rate calculation, which was used as the phototoxicity group. The cells were cultured and counted without light treatment, which was used as the dark toxicity group.

[0069] Figure 3 The photodynamic killing of Staphylococcus aureus by TPCL-MDPA at different concentrations. Figure 3 It can be seen that with the increase of TPCL-MDPA concentration, the survival rate of Staphylococcus aureus in the phototoxic group decreased significantly. When the concentration of TPCL-MDPA was 0.6 μM, the survival rate of Staphylococcus aureus in the phototoxic group was only about 1%, while the survival rate of Staphylococcus aureus in the dark toxicity group decreased slightly, indicating that TPCL-MDPA has a highly efficient photodynamic killing effect on Staphylococcus aureus.

[0070] (4) Phototoxicity and dark toxicity test of TPCL-MDPA on normal cells LO2. The specific steps are as follows:

[0071] LO2 cells were seeded in 96-well plates and incubated at 37°C and 5% CO 2 , 20% O 2 The cells were cultured in a humidified incubator in DMEM medium (containing 10% FBS and 1% penicillin / streptomycin). Then, the cells in the logarithmic growth phase were harvested and plated at 5×10 3 The density of cells was seeded in a 96-well plate and incubated for 24 hours. Subsequently, the culture medium was replaced with fresh culture medium containing different concentrations of TPCL-MDPA (final concentrations were 0 μM, 0.2 μM, 0.4 μM, 0.6 μM, respectively). After further incubation for 20 hours, the cells were exposed to white light irradiation for 30 minutes. At the same time, cells incubated with TCPL-MDPA without white light irradiation were used as dark control groups. After further incubation for 4 hours, the culture medium was removed and washed 3 times with PBS. The cells were then incubated in the dark for 1 hour with fresh serum-free culture medium containing 10% CCK-8. Finally, the absorbance of the product was measured at a wavelength of 450 nm using an ELISA reader. The results are expressed as the survival percentage of cells after different treatments relative to control cells without any treatment.

[0072] Figure 4 The phototoxicity and dark toxicity of TPCL-MDPA at different concentrations to normal cells LO2. It can be seen from the figure that TPCL-MDPA has no toxicity to normal cells LO2 under both light and dark conditions, and the cell survival rate is above 90%, indicating that TPCL-MDPA has good biocompatibility.

[0073] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A benzopyridazine compound, characterized in that: It has the structure shown in formula I:

2. The method for preparing the benzopyridazine compound according to claim 1, characterized in that: The following steps are involved: A compound having a structure shown in Formula II, 4,4'-dimethoxydiphenylamine, a palladium catalyst, tri-tert-butylphosphine tetrafluoroborate, an organic base and a benzene solvent are mixed to carry out a Buchwald-Hartwig coupling reaction to obtain a compound having a structure shown in Formula III; the benzene solvent is toluene; The compound having the structure shown in formula III is mixed with diphenylacetylene, an iridium catalyst, an inorganic base, a compound containing a non-coordinating tetrafluoroborate and an organic solvent, and a cyclization reaction is carried out under a protective atmosphere to obtain the benzopyridazine compound.

3. The preparation method according to claim 2, characterized in that: The molar ratio of the compound having the structure shown in formula II to 4,4'-dimethoxydiphenylamine is 1:1.8-2.2; the molar ratio of the compound having the structure shown in formula II, palladium catalyst, tri-tert-butylphosphine tetrafluoroborate and organic base is 1:0.03-0.05:0.08-0.12:2.5-3.

5.

4. The preparation method according to claim 2, characterized in that: The temperature of the Buchwald-Hartwig coupling reaction is 95-105° C. and the time is 20-25 hours.

5. The preparation method according to claim 2, characterized in that: The iridium catalyst is dichloro(pentamethylcyclopentadiene)iridium(III) dimer; the inorganic base is copper acetate; and the compound containing non-coordinated tetrafluoroborate is silver tetrafluoroborate.

6. The preparation method according to claim 2, characterized in that: The molar ratio of the compound having the structure shown in formula II to diphenylacetylene is 1:1.2-1.8; the molar ratio of the compound having the structure shown in formula II, iridium catalyst, inorganic base and compound containing non-coordinating tetrafluoroborate is 1:0.05-0.2:0.05-0.2:1.5-2.

5.

7. The preparation method according to claim 2, characterized in that: The temperature of the cyclization reaction is 120-135° C. and the time is 20-26 hours.

8. The preparation method according to claim 2, characterized in that: The method for preparing the compound having the structure shown in Formula II comprises the following steps: The p-bromoaniline, chloroalkane, potassium permanganate and ferrous sulfate are mixed for oxidative dimerization reaction to obtain a compound having a structure shown in formula II.

9. The preparation method according to claim 8, characterized in that: The molar ratio of potassium permanganate to ferrous sulfate is 1:1-1.2; the molar ratio of p-bromoaniline to potassium permanganate is 1:2.8-3.5; the temperature of the oxidative dimerization reaction is 35-45°C, and the time is 5-8h.

10. Use of the benzopyridazine compound according to claim 1 or the benzopyridazine compound prepared by the preparation method according to any one of claims 2 to 9 in the preparation of photodynamic antibacterial drugs; the bacteria are bacteria.

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