3, 3-difluoro-1, 7-dimethyl-5-phenyl-1, 3-dihydro-2h-benzo [b] azepine-2-one

Abstract

The invention belongs to the technical field of pharmacy, and particularly relates to a preparation method of 3, 3-difluoro-1, 7-dimethyl-5-phenyl-1, 3-dihydro-2H-benzo [b] azepine-2-ketone and application of the 3, 3-difluoro-1, 7-dimethyl-5-phenyl-1, 3-dihydro-2H-benzo [b] azepine-2-ketone. The compound provided by the invention is prepared by photochemical driving under the condition of no metal and no oxidant, and has antitumor activity.

Description

Technical Field

[0001] The present invention belongs to the field of pharmacy, and in particular relates to a preparation method and application of 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one. Background Art

[0002] Fluorinated and nitrogen-containing heterocyclic compounds have made significant progress in the fields of medicinal chemistry, materials science, and catalyst development in recent years due to their unique chemical properties and excellent biological activities. The introduction of fluorine atoms can significantly enhance the stability, lipid solubility, and metabolic stability of the compounds, thereby improving their pharmacokinetic properties in the body, thereby increasing bioavailability and targeting. The electronegativity of fluorine allows these compounds to form stable interactions when acting with biological molecules, reducing the side effects of drugs and enhancing therapeutic effects.

[0003] Using light energy as a driving force to replace traditional chemical catalysts or high temperature conditions can not only reduce pollution to the environment, but also conform to the concept of green chemistry. Photochemical reactions are usually carried out under mild conditions, which helps to improve the selectivity of the reaction and the purity of the product. By exciting molecules into a high-energy state through light, chemical reactions can be efficiently promoted at low or normal temperatures, thereby reducing energy consumption and improving reaction efficiency. Photochemically driven synthesis methods have high stereoselectivity and regioselectivity, making the synthesis of complex molecules more efficient and precise. Compared with traditional chemical reactions, it has fewer side reactions and by-products, and can significantly improve the yield and purity of the target product. Photochemical synthesis can also avoid the use of harmful heavy metals and oxidants, improve the safety and sustainability of the synthesis process, and conform to the core concept of green chemistry. Summary of the invention

[0004] The object of the present invention is to provide a preparation method and application of 3,3-difluoro-1,7-dimethyl-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 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepine-2-one, characterized in that the structural formula of the compound is shown in Formula 2e:

[0007]

[0008] The preparation method of 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one of the present invention comprises the following steps: at room temperature, 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-phenylvinyl)phenyl)acetamide, potassium dihydrogen phosphate, 2,4,5,6-tetra(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-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-phenylvinyl)phenyl)acetamide 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 3,3-difluoro-1,7-dimethyl-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-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-phenylvinyl)phenyl)acetamide to potassium dihydrogen phosphate is 1:1.5.

[0010] Furthermore, 0.3 mmol of 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-phenylvinyl)phenyl)acetamide, 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] Application of 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one in the preparation of anti-tumor drugs.

[0014] Application of 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one in the preparation of antibacterial drugs.

[0015] The beneficial effects of the present invention are as follows: the preparation of compounds by photochemical drive can efficiently promote chemical reactions under mild conditions, avoid the use of high temperature, high pressure and harmful catalysts, thereby reducing environmental pollution and safety requirements for experimental operations, the substrate is cheap and easy to obtain, and the reaction is green and economical. The prepared compound 2e has good anti-tumor and anti-bacterial activity and has the potential to be developed into anti-tumor and anti-bacterial drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 For compound 2e 1 H NMR spectrum;

[0017] Figure 2 For compound 2e 13 C NMR spectrum;

[0018] Figure 3 For compound 2e 19 F NMR spectrum;

[0019] Figure 4 This is the MS spectrum of compound 2e. DETAILED DESCRIPTION

[0020] 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.

[0021] Potassium dihydrogen phosphate (KH2PO4) (Cat. No.: P7392) and sodium bicarbonate (NaHCO3) (Cat. No.: S5240) 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 Bidex Pharmaceuticals; ethyl acetate (EtOAc) and petroleum ether were purchased from Tianjin Tianli Chemical Reagent Factory; ether was purchased from Merck; column chromatography silica gel was purchased from Qingdao Ocean Chemical Plant Branch;

[0022] DMEM medium (catalog number: MA0212) was purchased from Dalian Meilunbio Technology Co., Ltd.; RPMI-1640 medium (catalog number: R2405) and fetal bovine serum (catalog number: F8687) were purchased from Merck; MTT (catalog number: A4586) was purchased from Tianjin Alpha Biotechnology Co., Ltd.; nutrient agar medium (catalog number: 022020) was purchased from Guangdong Kaihuan Microorganisms; cisplatin and LB medium (ST163-500ml) were purchased from Beyotime Biotechnology Co., Ltd.; ciprofloxacin was purchased from MedChemExpress; dimethyl sulfoxide (DMSO) (catalog number: D6370) was purchased from Beijing Biotopped Co., Ltd.

[0023] 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.

[0024] Example 1

[0025] At room temperature, 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-phenylvinyl)phenyl)acetamide 1e (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 maintained in an air atmosphere at 25 ° C. It was monitored by thin layer chromatography (TLC) (volume ratio, petroleum ether: ether = 3: 1) until the starting material 1e was completely consumed. After the reaction was completed, the resulting 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, and the residue was purified by flash silica gel column chromatography (200-300 mesh silica gel) using ethyl acetate and petroleum ether as eluents (volume ratio, petroleum ether:ethyl acetate=30:1) to give the product 2e (64.65 mg) in a yield of 74%.

[0026]

[0027] The spectrum results are as follows Figure 1-3 As shown. 1 H NMR, 13 C NMR, 19 The structure of the compound was obtained from the F NMR spectrum. Specifically:

[0028] Compound 2e was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 30:1) to obtain a yellow solid (64.65 mg, 74%): mp: 123-124°C; 1 H NMR (400MHz, CDCl3) δ7.40-7.30 (m, 6H), 7.28 (s, 1H), 6.95 (s, 1H), 6.19 (t, J = 12Hz, 1H), 3.56 (s, 3H), 2.25 (s, 3H). 13C NMR (100MHz, CDCl3) δ163.8 (t, J = 33Hz), 146.6 (dd, J = 7, 11Hz), 139.4, 138.6 (d, J = 4Hz), 13 4.6,130.9,130.8,130.4,129.3,129.0,128.7,123.4,122.8,122.2,112.6(m),37.8,20.8. 19 F NMR (471MHz, CDCl3) δ-104.64 (dd, J=255.1, 7.6Hz, 1F), -108.01 (dd, J=255.0, 11.8Hz, 1F). like Figure 4 As shown, the mass-to-nuclear ratio measured by mass spectrometry is m / z:338.0761[M+K] + , indicating that its molecular formula is C 18 H 15 F2NO.

[0029] Example 2

[0030] Antitumor activity test

[0031] Test cells: human colon cancer cells (HCT-116) (GDC0625), human cervical cancer cells (HeLa) (GDC0009), human breast cancer cells (MCF-7) (GDC0055); human liver cancer cells (HepG-2) (Cat. No.: C6346) were purchased from Beyotime Biotech.

[0032] Human liver cancer cells (HepG-2) and human cervical cancer cells (HeLa) were cultured in DMEM medium containing 10% fetal bovine serum, and human colon cancer cells (HCT-116) and human breast cancer cells (MCF-7) were cultured in RPMI-1640 medium containing 10% fetal bovine serum. When the cells grew to 85% of the culture flask, they were digested with trypsin and subcultured, and cells in the logarithmic growth phase were selected for experiments.

[0033] The MTT colorimetric method was used to detect the inhibitory effect of compound 2e on the proliferation of various tumor cells.

[0034] Drug preparation: Compound 2e was dissolved in DMSO (final DMSO concentration <0.1%) and diluted to the required concentration in the prepared culture medium before the experiment.

[0035] Take the tumor cell line in the logarithmic growth phase to prepare a cell suspension. Dilute the cell suspension to a concentration of 5×10 4 / mL, 100μL cell suspension was inoculated into each well of a 96-well culture plate. After incubation for 24h, the supernatant was removed and 100μL drug-containing culture medium was added to each well. The final concentration gradient of compound 2e was 64, 32, 16, 8, 4, and 2μg / mL. The positive control drug was cisplatin, and the blank control group was added with equal volumes of culture medium and DMSO (i.e., cells in a single well were not given drugs and were cultured as normal). Three replicate wells were set for each treatment. After culturing in a 37°C, 5% CO2 incubator for 48h, 10μL of MTT solution (5mg / mL) was added to each well, and culture was continued in a 37°C, 5% CO2 incubator for 4h. The supernatant was discarded, 150μL DMSO was added to each well, and the mixture was shaken for 10min. The absorbance value (OD) was detected at a wavelength of 490nm using an enzyme reader.

[0036] The calculation formula of the inhibition rate of compound 2e on tumor cell proliferation is:

[0037] Inhibition rate (%) = (1-OD value of experimental group / OD value of blank control group) * 100%.

[0038] Experimental Results

[0039] 1. Inhibitory effect of compound 2e on the proliferation of HepG-2 liver cancer cells

[0040] The results are shown in Table 1. The inhibitory effect of compound 2e on the proliferation of HepG-2 liver cancer cells was concentration-dependent. At a low concentration of 2 μg / mL, the inhibition rate of compound 2e was 12.4%. As the concentration increased, the inhibition rate gradually increased, reaching 76.7% at 64 μg / mL. The inhibition rate of the positive control cisplatin group was even higher, at 2 μg / mL, it was 29.5%, and at 64 μg / mL, it reached 97.7%.

[0041] Table 1 MTT test results of compound 2e on the proliferation inhibition of HepG-2 liver cancer cells

[0042]

[0043] 2. Inhibitory effect of compound 2e on the proliferation of HCT-116 colorectal cancer cells

[0044] The results are shown in Table 2. The inhibition rate of compound 2e on HCT-116 colorectal cancer cells gradually increased from 11.0% at 2 μg / mL to 99.2% at 64 μg / mL, showing significant concentration dependence. The inhibition rate of the cisplatin group was 9.48% at 2 μg / mL and 80.13% at 64 μg / mL.

[0045] Table 2 MTT test results of compound 2e on the proliferation inhibition of HCT-116 colorectal cancer cells

[0046]

[0047] 3. Inhibitory effect of compound 2e on the proliferation of HeLa cervical cancer cells

[0048] The results are shown in Table 3. In the HeLa cervical cancer cell experiment, compound 2e also exhibited a concentration-dependent inhibitory effect, with the inhibition rate increasing from 8.7% at 2 μg / mL to 68.8% at 64 μg / mL. The inhibition rate of the cisplatin group was 6.9% at 2 μg / mL and 89.2% at 64 μg / mL.

[0049] Table 3 MTT test results of compound 2e on the proliferation inhibition of HeLa cervical cancer cells

[0050]

[0051] 4. Inhibitory effect of compound 2e on proliferation of MCF-7 breast cancer cells

[0052] The results are shown in Table 4. The inhibitory effect of compound 2e on MCF-7 breast cancer cells increased with increasing concentration, from 5.7% at 2 μg / mL to 75.8% at 64 μg / mL. The inhibition rate of the cisplatin group was 25.2% at 2 μg / mL and 94.5% at 64 μg / mL.

[0053] Table 4 MTT test results of compound 2e on the proliferation inhibition of MCF-7 breast cancer cells

[0054]

[0055] 5. Inhibitory effect of compound 2e on tumor cell proliferation

[0056] IC values ​​of compound 2e and positive control drug cisplatin on four tumor cells 50 The values ​​are shown in Table 5. From the results in Table 5, it can be seen that the compound 2e of the present invention exhibited an inhibitory effect on the proliferation of four different tumor cells in the in vitro anti-tumor activity test, especially the inhibitory effect on human colon cancer cells (HCT-116) was significantly better than the positive control drug cisplatin (IC 50 =15.43 μg / mL), its IC 50 The value was 5.67 μg / mL, showing its potential for development as a new anti-tumor drug.

[0057] Table 5 Inhibitory effect of compound 2e on tumor cell proliferation (IC 50 μg / mL)

[0058]

[0059] Example 3

[0060] Antibacterial activity test

[0061] Test bacterial strains: Bacillus subtilis (CMCC(B)63564), Staphylococcus aureus (CMCC(B)26003), Salmonella typhi (CMCC(B)50071), Pseudomonas aeruginosa (CMCC(B)10104).

[0062] The antibacterial activity of compound 2e was determined by the microdilution method. The bacterial strains used in the activity test were placed in a 37°C constant temperature incubator for activation for 2-3 hours. After that, a small amount of the activated strains were selected and placed in sterile water, shaken and prepared at a concentration of 5×10 5 CFU / mL of bacterial suspension.

[0063] Drug preparation: Compound 2e was dissolved in DMSO to prepare a 2 mg / mL stock solution (DMSO concentration should not exceed 0.5% v / v), and the stock solution was diluted to 64 μg / mL with LB medium for later use.

[0064] 100 μL of drug-containing culture medium was added to a 96-well plate, and then 100 μL of bacterial suspension was added to each well for dilution, so that the final concentration of compound 2e was 64, 32, 16, 8, 4, 2, 1, 0.5, 0.25, and 0.125 μg / mL. Ciprofloxacin was used as a positive control, LB culture medium containing 0.5% DMSO was used as a negative control, and LB culture medium was used as a blank control. Each group of experiments was repeated 3 times. After culturing in a 37°C constant temperature incubator for 24 hours, the turbidity of the 96-well plate bacterial solution was observed. The minimum concentration at which the culture solution was clear was the minimum inhibitory concentration (MIC) of the compound.

[0065] 10 μL of the growth-inhibited bacteria was applied to nutrient agar medium from each well and cultured in a 37°C constant temperature incubator for 24 h. Each group of experiments was repeated 3 times, and the lowest concentration at which the bacteria were completely killed was the minimum bactericidal concentration (MBC).

[0066] Experimental Results

[0067] The MIC and MBC values ​​of compound 2e and the positive control drug ciprofloxacin against four standard bacterial strains are shown in Table 6. The results show that compound 2e has a certain inhibitory effect on the four strains, among which the inhibitory effect on Bacillus subtilis is more significant.

[0068] Table 6 MIC values ​​and MBC values ​​(μg / mL) of compound 2e against four tested bacterial strains

[0069]

[0070] “-” in the table means that the corresponding MIC and MBC values ​​were not detected within the experimental concentration range.

[0071] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one, characterized in that The structural formula of the compound is shown in 2e:

2. The method for preparing 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one according to claim 1, characterized in that: The method comprises the following steps: at room temperature, sequentially adding 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-phenylvinyl)phenyl)acetamide, potassium dihydrogen phosphate, 2,4,5,6-tetra(9-carbazolyl)-isophthalonitrile and tetrahydrofuran into a reaction bottle, irradiating a photolysis reaction with a 30W blue LED lamp, maintaining the reaction under an air atmosphere at 25°C, and monitoring the reaction by thin layer chromatography until the 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-phenylvinyl)phenyl)acetamide is completely consumed; after the reaction is completed, quenching and extracting the obtained mixture; vacuum concentrating the organic solvent, purifying the residue by fast silica gel column chromatography, and eluting with an eluent to obtain 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one.

3. The method for preparing 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one according to claim 2, characterized in that: The molar ratio of the 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-phenylvinyl)phenyl)acetamide to potassium dihydrogen phosphate is 1:1.

5.

4. The method for preparing 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one according to claim 3, characterized in that: The 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-phenylvinyl)phenyl)acetamide was 0.3 mmol, and potassium dihydrogen phosphate was 0.45 mmol.

5. The method for preparing 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one according to claim 2, characterized in that: The mixture of 0.009 mmol of 2,4,5,6-tetrakis(9-carbazolyl)-isophthalonitrile and 3 mL of tetrahydrofuran was quenched with 15 mL of saturated aqueous sodium bicarbonate solution and extracted three times with 15 mL of ethyl acetate; the flash column chromatography purification condition was that the volume ratio of the eluent was petroleum ether:ethyl acetate = 30:

1.

6. Application of 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepine-2-one in the preparation of anti-tumor drugs.

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