7-bromo-3, 3-difluoro-1-methyl-5-phenyl-1, 3-dihydro-2h-benzo [b] azepine-2-one
The preparation of 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azazo-2-one was solved by a photochemically driven method, which solved the problem of insufficient selectivity and affinity of compounds in drug development, and reduced environmental pollution and energy consumption through green preparation methods, providing potential drugs for treating liver diseases.
Patent Information
- Application Number
- CN202510079950.6
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-01-18
- Publication Date
- 2025-04-25
AI Technical Summary
The prior art is difficult to effectively improve the selectivity and affinity of compounds for biological targets in drug development, and traditional preparation methods have problems such as heavy metal residues, environmental pollution and high energy consumption.
7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azazo-2-one was prepared by photochemically driven. The target compound was obtained by irradiating the reaction mixture with a 30W blue LED lamp at room temperature, combined with thin layer chromatography, quenching, extraction and flash column chromatography purification steps.
The high selectivity and affinity of the compounds for biological targets is achieved, the pharmacological activity is significantly improved, and environmental pollution and energy consumption are reduced through green preparation methods, providing potential drugs for the treatment of alcoholic liver disease and acute liver injury.
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Figure CN119874619A_ABST
Abstract
Description
Technical Field
[0001] The present invention belongs to the field of pharmacy, and in particular relates to a preparation method and application of 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one. Background Art
[0002] Fluorinated seven-membered nitrogen heterocyclic compounds have a wide range of applications in many fields due to their unique chemical properties. The introduction of fluorine atoms can significantly improve the lipid solubility, stability and affinity of the compounds. Especially in drug development, it can enhance the selectivity and affinity of the compounds for biological targets, thereby effectively improving their pharmacological activity. The fluorine element can also enhance the resistance of the compounds to metabolic enzymes, delay the metabolic degradation of drugs, thereby prolonging their half-life in the body and improving the stability of drugs. This property enables the drug to continue to work effectively in the body, thereby improving the therapeutic effect, reducing the frequency of administration and improving patient compliance. Fluorinated seven-membered nitrogen heterocyclic compounds also have strong target selectivity, which helps to reduce side effects in drug design and ensure that the drug can accurately act on the target cells or tissues.
[0003] Photochemically driven preparation of compounds can avoid environmental pollution and subsequent separation problems caused by heavy metal residues in the reaction, and can also effectively reduce the consumption of chemical reagents and the generation of waste, further reducing energy consumption and costs in the preparation process. As a clean and renewable energy source, light energy directly drives the reaction, greatly reducing the high energy consumption caused by harsh conditions such as high temperature and high pressure in traditional methods, and has good application prospects. Summary of the invention
[0004] The object of the present invention is to provide a preparation method and application of 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one.
[0005] The technical solution of the present invention to solve the above technical problems is as follows:
[0006] The compound of the present invention is 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one, characterized in that the structural formula of the compound is as shown in Formula 2d:
[0007]
[0008] The preparation method of 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one of the present invention comprises the following steps: at room temperature, 2-bromo-N-(4-bromo-2-(1-phenylvinyl)phenyl)-2,2-difluoro-N-methylacetamide, potassium dihydrogen phosphate, 2,4,5,6-tetrakis(9-carbazolyl)-isophthalonitrile and tetrahydrofuran are sequentially added into a reaction bottle, and a 30W blue LED lamp is used for photolysis reaction. The reaction is carried out under an air atmosphere at 25°C and monitored by thin layer chromatography until 2-bromo-N-(4-bromo-2-(1-phenylvinyl)phenyl)-2,2-difluoro-N-methylacetamide is completely consumed; after the reaction is completed, the resulting mixture is quenched and extracted; the organic solvent is concentrated in vacuo, and the residue is purified by flash silica gel column chromatography, and 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one is obtained by eluting with an eluent.
[0009] Furthermore, the molar ratio of 2-bromo-N-(4-bromo-2-(1-phenylvinyl)phenyl)-2,2-difluoro-N-methylacetamide to potassium dihydrogen phosphate is 1:1.5.
[0010] Furthermore, 0.3 mmol of 2-bromo-N-(4-bromo-2-(1-phenylvinyl)phenyl)-2,2-difluoro-N-methylacetamide, 0.45 mmol of potassium dihydrogen phosphate, 0.009 mmol of 2,4,5,6-tetrakis(9-carbazolyl)-isophthalonitrile and 3 mL of tetrahydrofuran were added to the reaction bottle.
[0011] Furthermore, the developing solvent condition of thin layer chromatography was a volume ratio of petroleum ether to diethyl ether of 3:1, and the mixture after completion of the reaction was quenched with 15 mL of a saturated aqueous solution of sodium bicarbonate, and extracted three times with 15 mL of ethyl acetate.
[0012] Furthermore, the flash column chromatography purification condition is that the volume ratio of the eluent is petroleum ether:ethyl acetate=30:1.
[0013] The invention provides an application of 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one in the preparation of a drug for treating alcoholic liver disease.
[0014] The present invention provides an application of 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one in the preparation of a drug for treating acute liver injury. 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one can be used alone or in combination with other drugs to treat acute liver injury. It is particularly effective in the ICR mouse liver injury model.
[0015] The present invention provides an application of 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one in the preparation of liver protection products, wherein the products include but are not limited to one or more of medicines, foods, and health products.
[0016] Beneficial effects of the present invention: The light-driven preparation of 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepine-2-one has a liver-protecting effect, has good application prospects in the treatment of alcoholic liver disease and acute liver injury, and can be used as a potential candidate drug for the prevention and treatment of related diseases. In addition, the light-driven preparation reaction can effectively avoid environmental pollution and separation problems caused by heavy metal residues during the reaction process, greatly reducing the energy consumption and cost of the preparation process, which is in line with the concept of green sustainable chemical development. BRIEF DESCRIPTION OF THE DRAWINGS
[0017] Figure 1 is the compound 2d of the present invention 1 HNMR spectrum;
[0018] Figure 2 is the compound 2d of the present invention 13 CNMR spectrum;
[0019] Figure 3 is the compound 2d of the present invention 19 FNMR spectrum;
[0020] Figure 4 is the MS spectrum of compound 2d of the present invention;
[0021] Figure 5 The effects of different concentrations of alcohol intervention for 24 hours on the survival rate of AML12 cells;
[0022] Figure 6 The effect of compound 2d on the survival rate of AML12 cells 24 hours after administration;
[0023] Figure 7 The effect of compound 2d on the serum ALT and AST activities in mice with liver damage induced by CCl4;
[0024] Figure 8 The effect of compound 2d on the gross and histopathological changes of mouse liver (HE staining, magnification 400×). DETAILED DESCRIPTION
[0025] The principles and features of the present invention are described below in conjunction with the accompanying drawings. The examples given are only used to explain the present invention and are not used to limit the scope of the present invention.
[0026] Potassium dihydrogen phosphate (KH2PO4) (Cat. No.: P7392), sodium bicarbonate (NaHCO3) (Cat. No.: S5240), and hematoxylin-eosin (HE) staining kit (Cat. No.: 1120) were purchased from Beijing Solebow Technology Co., Ltd.; tetrahydrofuran (THF) was purchased from Tianjin Komiou Chemical Reagent Co., Ltd.; 2,4,5,6-tetrakis(9-carbazolyl)-isophthalonitrile (4-CzIPN) was purchased from Bid Pharmaceuticals; ethyl acetate (EtOAc), petroleum ether, and anhydrous ethanol were purchased from Tianjin Tianli Chemical Reagent Factory; ether and 10% formaldehyde solution were purchased from Merck; column chromatography silica gel was purchased from Qingdao Ocean Chemical Plant Branch.
[0027] Fetal bovine serum (catalog number: F8687) was purchased from Merck; penicillin-streptomycin-amphotericin B solution (100X), dexamethasone, insulin, CCK-8 kit (catalog number: C0038), and phosphate buffered saline (PBS) were all purchased from Beyotime Biotechnology Co., Ltd.; transferrin and DMED / F-12 culture medium were purchased from Dalian Meilun Biotechnology Co., Ltd.; carbon tetrachloride (CCl4) was purchased from Beijing Chemical Plant; silymarin was purchased from MedChemExpress; alanine aminotransferase (alanine aminotransferase / ALT / GPT) test kit (Reitman method) (catalog number: C009-1-1) and aspartate aminotransferase (aspartate aminotransferase / AST / GOT) test kit (colorimetric method) (catalog number: C010-1-1) were all purchased from Nanjing Jiancheng Bioengineering Institute.
[0028] Test cells: Mouse normal hepatocytes (AML12) (Cat. No.: CL-0602) were purchased from Wuhan Pronocell Life Science Technology Co., Ltd.
[0029] Experimental animals: ICR mice were purchased from Liaoning Changsheng Biotechnology Co., Ltd.
[0030] In the present invention, unless otherwise specified, all methods are conventional in the art. The materials, reagents, methods and instruments used are conventional in the art unless otherwise specified, and those skilled in the art can obtain them through commercial channels or prepare the solutions used in the experiment using conventional methods in the art.
[0031] Example 1
[0032] In this embodiment, the method for photochemically driving the preparation of 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one under metal-free and oxidant-free conditions is as follows:
[0033] At room temperature, 2-bromo-N-(4-bromo-2-(1-phenylvinyl)phenyl)-2,2-difluoro-N-methylacetamide 1d (0.3 mmol), KH2PO4 (1.5 equiv, 0.45 mmol), 4-CzIPN (0.009 mmol) and THF (3 mL) were added to the reaction bottle in sequence. The photolysis reaction was irradiated with a 30 W blue LED lamp and carried out in an air atmosphere maintained at 25 ° C. It was monitored by thin layer chromatography (TLC) (volume ratio of petroleum ether to diethyl ether was 3:1) until the starting material 1d was completely consumed. After the reaction was completed, the mixture was quenched with NaHCO3 (sat.aq. 15 mL) and extracted with EtOAc (3×15 mL). The organic solvent was concentrated in vacuo at 55 ° C and 80 r / min. The residue was purified by flash column chromatography using ethyl acetate and petroleum ether as eluents (the volume ratio of petroleum ether to ethyl acetate was 30:1) to give the product 2d (85.22 mg) in a yield of 78%.
[0034]
[0035] The spectrum results are as follows Figure 1-4 As shown. 1 H NMR, 13 C NMR, 19 The structure of the compound was obtained by F NMR and MS spectra. Specifically:
[0036] Compound 2d was purified by silica gel column chromatography (petroleum ether:ethyl acetate=30:1) to obtain a yellow solid (85.22 mg, 78%), mp: 160-161°C; 1 H NMR (400MHz, CDCl3) δ7.56 (dd, J = 8.7, 2.3Hz, 1H), 7.45-7.40 (m, 3H), 7.32-7.26 (m, 4H), 6.26 (t, J = 10.4Hz, 1H), 3.58 (s, 3H). 13 C NMR (100MHz, CDCl3) δ163.6 (t, J = 32Hz), 145.5-145.2 (m), 140.0 (d, J = 4.0Hz), 138.6, 133. 2,132.9,132.6,129.7,128.9,124.5,124.2,124.0,123.9,118.0,114.6-109.6(m),37.8. 19 F NMR (471MHz,CDCl3)δ-104.25(d,J=256.6Hz,1F),-108.41(d,J=256.6Hz,1F). The mass-to-nuclear ratio measured by mass spectrometry is m / z:401.9706[M+K] +, indicating that its molecular formula is C 17 H 12 BrF2NO.
[0037] Example 2
[0038] Study on the protective effect of compound 2d on alcoholic liver disease
[0039] 1 Experimental methods
[0040] 1.1 Cell culture
[0041] AML12 cells were cultured in a 37°C, 5% CO2 incubator, and DMED / F-12 complete medium containing 10% fetal bovine serum, 1% penicillin-streptomycin-amphotericin B mixture, 10 μg / mL insulin, 0.1 μmol / L dexamethasone and 5 μg / mL transferrin was added to the culture flask.
[0042] 1.2 Cellular intervention
[0043] AML12 cells were cultured at 15×10 4 Each well was inoculated in a six-well plate, and the experimental intervention was performed when the uniform growth reached a density of about 80% under an inverted microscope. In the preliminary experiment, each well was added with culture medium containing different concentrations of alcohol (0, 100, 200, 300, 400, 500, 600, 700 mmol / L) for 24 hours to determine the modeling concentration.
[0044] The formal experiment was divided into a blank control group, an alcohol modeling group, a positive control group (treated with a medium containing 10 μg / mL silymarin for 24 hours, and then treated with a medium containing 300mmol / L alcohol for 24 hours) and a compound 2d intervention group (treated with a medium containing 0.625, 1.25, 2.5, 5, 10, 20 and 40 μg / mL compound 2d for 24 hours, and then treated with a medium containing 300mmol / L alcohol for 24 hours), with 6 replicates in each group. The normal control group and the alcohol modeling group were cultured with complete medium for 24 hours, and the compound 2d intervention group was intervened for 24 hours according to the experimental group; the original medium was discarded, and PBS was washed twice. The alcohol modeling group and the compound 2d intervention group were both treated with a medium containing 300mmol / L alcohol for 24 hours, and the normal control group was replaced with fresh medium for 24 hours.
[0045] 1.3 CCK-8 assay to detect cell proliferation
[0046] AML12 cells were cultured at 1×10 4100 μL of culture medium containing CCK-8 reagent was added to each well (the ratio of CCK-8 reagent to complete culture medium was 1:10), and then placed in a 37°C constant temperature incubator for 2 hours, and then the absorbance value was measured at 450 nm. The cell survival rate was calculated according to the following formula:
[0047] Cell survival rate (%) = (OD a -OD b ) / (OD c -OD b )×100, where: OD a : Absorbance values of wells with cells, culture medium, drugs and CCK-8 solution; OD b : Absorbance value of wells with culture medium and CCK-8 solution; OD c : Absorbance values of wells with cells, culture medium and CCK-8 solution.
[0048] 2 Experimental Results
[0049] 2.1 Alcohol modeling conditions
[0050] AML12 cells were used to construct an in vitro model of alcoholic liver injury. In order to clarify the modeling conditions, AML12 cells were treated with different concentrations of alcohol for 24 hours, and the changes in cell survival rate were detected. Figure 5 As shown, alcohol treatment can lead to a decrease in the survival rate of AML12 cells, and the effect is dose-dependent. The condition of cell survival rate between 50% and 60% was selected as the basis for constructing a cell oxidative damage model. When the alcohol concentration was 300mmol / L, the cell survival rate was 68.45%. At this time, the degree of damage to AML12 cells was close to half, and it was still in a reversible stage, which was more in line with the early state of alcoholic liver disease. Therefore, in subsequent experiments, the alcoholic liver injury model used a complete culture medium containing 300mmol / L alcohol to treat cells for 24 hours. ( Figure 5 Compared with 0mmol / L, *P<0.05, **P<0.01).
[0051] 2.2 Effect of compound 2d on AML12 cell activity
[0052] The experimental results are as follows Figure 6 As shown, in the alcohol injury model constructed with AML12 cells, compound 2d showed a significant protective effect on cell proliferation in the concentration range of 0.625μg / mL to 10μg / mL. At concentrations of 5μg / mL and 10μg / mL, cell survival rate was significantly improved, indicating that compound 2d can effectively resist alcohol-induced cytotoxicity. ( Figure 6 Compared with the control group, *P<0.05, **P<0.01; compared with the model group, #P<0.05, ##P<0.01).
[0053] The experimental results showed that compound 2d had a significant protective effect in the in vitro alcoholic liver injury model. It not only increased the survival rate of AML12 cells after alcohol treatment, but also showed an effect comparable to or even better than the positive control silymarin. This discovery provides an important basis for the potential application of compound 2d in the prevention and treatment of alcoholic liver disease, and also lays the foundation for subsequent in vivo research and mechanism exploration.
[0054] Example 3
[0055] 1 Experimental methods
[0056] 1.1 Animal grouping, drug administration and modeling
[0057] Fifty male ICR mice (SPF grade, 6-8 weeks old, weighing 24-26 g) were randomly divided into a blank control group, a model group, and a low-dose group (20 mg kg -1 ·d -1 ), high-dose group (60 mg·kg -1 ·d -1 ), positive drug group (silymarin 60 mg kg -1 ·d -1 ) 10 mice in each group. Adaptive feeding was performed for 3 days, and then each dose group was gavaged with low and high doses of compound 2d, while the blank group and model group were gavaged with equal amount of distilled water for 7 consecutive days. One hour after the last gavage, each dose group of compound 2d, positive drug group and model group were intraperitoneally injected with 5mL kg of 0.3% CCl4 vegetable oil solution -1 The blank control group was gavaged with the same amount of vegetable oil. After fasting for 16 hours, the blood was collected from the mouse orbits and centrifuged to obtain serum for testing.
[0058] 1.2 Detection of ALT and AST in serum
[0059] After blood was collected from mice, the samples were centrifuged at 3500 r / min for 10 min. Serum was collected and the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were detected using an ultraviolet spectrophotometer.
[0060] 1.3 Pathological staining of liver tissue
[0061] The mouse liver was fixed with 10% formaldehyde solution for 24 hours, dehydrated, transparently immersed in wax, embedded, sectioned, and stained with HE. The changes in mouse liver tissue and fibrosis were observed under a microscope.
[0062] 2 Experimental results
[0063] 2.1 Effect of compound 2d on serum ALT and AST activities in mice with CCl4-induced liver injury
[0064] The experimental results are as follows Figure 7 As shown in the figure, compared with the control group, the ALT and AST activities in the serum of the mice in the model group were significantly increased (P < 0.01), indicating that the CCl4-induced liver injury model was successfully established. Compared with the model group, the ALT and AST activities in the serum of the silymarin group, the high-dose and low-dose groups of compound 2d were significantly reduced (P < 0.01), showing a significant liver protective effect. ( Figure 7 Compared with the control group, *P<0.05, **P<0.01; compared with the model group, #P<0.05, ##P<0.01).
[0065] 2.2 Effects of compound 2d on the gross and histopathological characteristics of mouse liver
[0066] The experimental results are as follows Figure 8 As shown in the figure, the liver lobules and hepatocytes of the mice in the control group were normal and clearly identifiable; the liver lobules of the mice in the model group were damaged, the hepatocytes around the central vein were necrotic, the hepatocytes were significantly enlarged, the nuclei were of different sizes, or even disappeared, indicating that the modeling was successful. As the dose increased, the liver structure of the mice in each dose group gradually became clearer, the lesion range gradually decreased, and the size of the hepatocytes and nuclei gradually returned to normal, indicating that the compound 2d can alleviate the degree of hepatocyte lesions in mice, and the high-dose group has a better effect. It can be seen that the compound 2d has a certain effect on alleviating the pathological damage of liver tissue caused by CCl4.
[0067] Compound 2d showed significant hepatoprotective effects in the CCl4-induced mouse liver injury model. Compared with the model group, both the high- and low-dose groups of compound 2d could effectively reduce the activity of ALT and AST in serum and alleviate the pathological damage of liver tissue caused by CCl4 in a dose-dependent manner. The high-dose group had a more significant effect, which could significantly improve the liver lobule structure and hepatocyte morphology, indicating that compound 2d had a good protective and repairing effect on CCl4-induced liver injury.
Claims
1. 7-Bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one, characterized in that The structural formula of the compound is shown in 2d:
2. Use of 7-bromo-3,3-difluoro-1-methyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one according to claim 1 in the preparation of a drug for treating alcoholic liver disease.