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

Abstract

The application belongs to the technical field of pharmacy, and particularly 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. The compound is prepared under the conditions of no metal and no oxidant by photochemical driving, and the compound has antitumor activity.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of pharmacy, and particularly 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

[0002] Fluorine-containing nitrogen-containing heterocyclic compounds have made remarkable progress in the fields of pharmaceutical chemistry, material 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 compounds, improving their pharmacokinetic properties in the body, thereby increasing their bioavailability and targeting. The electronegativity of fluorine can enable these compounds to form stable interactions when interacting with biological molecules, reducing drug side effects and enhancing therapeutic effects.

[0003] Using light energy as a driving force instead of traditional chemical catalysts or high-temperature conditions not only reduces environmental pollution but also meets 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 driven at low or room temperature, thereby reducing energy consumption and improving reaction efficiency. Photochemical synthesis driven by light has 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, which can significantly improve the yield and purity of the target product. Photochemical synthesis can also avoid the use of harmful heavy metals and oxidizing agents, improving the safety and sustainability of the synthesis process and meeting the core concept of green chemistry. SUMMARY

[0004] The present application aims 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 application to solve the above technical problems is as follows:

[0006] The compound of the present application is 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one, and the structural formula of the compound is as 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 comprises the following steps: at room temperature, 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-styryl)phenyl)acetamide, potassium dihydrogen phosphate, 2,4,5,6-tetrakis(9-carbazolyl)-m-benzene dicarbonitrile and tetrahydrofuran are sequentially added into a reaction bottle, a photolysis reaction is irradiated using a 30W blue LED lamp, and the reaction is carried out in an air atmosphere kept at 25 DEG C, and is monitored by thin layer chromatography until 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-styryl)phenyl)acetamide is completely consumed; after the reaction is completed, the obtained mixture is quenched and extracted; the organic solvent is concentrated under vacuum, and the residue is purified by a fast silica gel column chromatography to obtain 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one by elution with an eluent.

[0009] Further, the molar ratio of 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-styryl)phenyl)acetamide to potassium dihydrogen phosphate is 1:1.5.

[0010] Further, 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-styryl)phenyl)acetamide 0.3 mmol, potassium dihydrogen phosphate 0.45 mmol, 2,4,5,6-tetrakis(9-carbazolyl)-m-benzene dicarbonitrile 0.009 mmol and tetrahydrofuran 3 mL are added into the reaction bottle.

[0011] Further, the thin layer chromatography developing agent is petroleum ether and ethyl ether with a volume ratio of 3:1, the mixture after the reaction is completed is quenched with 15 mL of a saturated aqueous sodium bicarbonate solution and extracted with 15 mL of ethyl acetate for 3 times.

[0012] Further, the fast column chromatography purification condition is that the volume ratio of the eluent is petroleum ether: ethyl acetate = 30:1.

[0013] 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one is used for preparing an antitumor drug.

[0014] 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2H-benzo[b]azepin-2-one is used for preparing an antibacterial drug.

[0015] The present application has the following beneficial effects: the chemical reaction can be efficiently promoted under mild conditions by photochemical driving preparation of the compound, avoiding the use of high temperature, high pressure and harmful catalysts, thereby reducing the pollution to the environment and the safety requirements for experimental operation, the substrate is cheap and easy to obtain, and the reaction is green and economical. The prepared compound 2e has good antitumor and antibacterial activity, and has the potential to develop into an antitumor and antibacterial drug. BRIEF DESCRIPTION OF DRAWINGS

[0016] Figure 1 The H NMR spectrum of compound 2e is as follows: 1 The H NMR spectrum of compound 2e is as follows:

[0017] Figure 2 The C NMR spectrum of compound 2e is as follows: 13 The C NMR spectrum of compound 2e is as follows:

[0018] Figure 3 The F NMR spectrum of compound 2e is as follows: 19 The F NMR spectrum of compound 2e is as follows:

[0019] Figure 4 The MS spectrum of compound 2e is as follows. DETAILED DESCRIPTION

[0020] The principles and characteristics of the present application are described below in conjunction with the accompanying drawings, and the examples are only used to explain the present application and are not used to limit the scope of the present application.

[0021] Potassium dihydrogen phosphate (KH2PO4) (item number: P7392), sodium bicarbonate (NaHCO3) (item number: S5240) were purchased from Beijing Solabio Science and Technology Co., Ltd.; tetrahydrofuran (THF) was purchased from Tianjin Kemio Chemical Reagent Co., Ltd.; 2,4,5,6-tetra(9-carbazolyl)-isophthalonitrile (4-CzIPN) was purchased from Bide Pharmaceutical; ethyl acetate (EtOAc) and petroleum ether were purchased from Tianjin Tianli Chemical Reagent Factory; diethyl ether was purchased from Merck; column chromatography silica gel was purchased from Qingdao Haizhuan Chemical Factory Branch;

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

[0023] In the present application, the methods are conventional methods in the art unless otherwise specified. The materials, reagents, methods and instruments used are conventional materials, reagents, methods and instruments in the art unless otherwise specified, which can be obtained through commercial channels or prepared by conventional methods in the art to prepare solutions for experiments.

[0024] Example 1

[0025] To the reaction flask was added 2-bromo-2,2-difluoro-N-methyl-N-(4-methyl-2-(1-styryl)phenyl)acetamide 1e (0.3 mmol), KH2PO4 (1.5 equiv, 0.45 mmol), 4-CzIPN (0.009 mmol) and THF (3 mL) successively at room temperature. The photolysis reaction was carried out using a 30 W blue LED lamp irradiation, maintained at 25 °C in an air atmosphere, monitored by thin layer chromatography (TLC) (volume ratio, petroleum ether: diethyl 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 x 15 mL). The organic solvent was concentrated under vacuum at 55 °C with a rotation speed of 80 r / min, and the residue was purified by flash silica gel column chromatography (200-300 mesh silica gel) eluted with ethyl acetate and petroleum ether as eluent (volume ratio, petroleum ether: ethyl acetate = 30: 1) to obtain the product 2e (64.65 mg) with a yield of 74%.

[0026]

[0027] The spectral results are shown in Figures 1-3 The structure of compound 2e was determined by 1 H NMR, 13 C NMR, 19 F NMR spectra. 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 (400 MHz, CDCl3) δ 7.40-7.30 (m, 6H), 7.28 (s, 1H), 6.95 (s, 1H), 6.19 (t, J = 12 Hz, 1H), 3.56 (s, 3H), 2.25 (s, 3H). 13C NMR (100 MHz, CDC13) δ 163.8 (t, J = 33 Hz), 146.6 (dd, J = 7, 11 Hz), 139.4, 138.6 (d, J = 4 Hz), 134.6, 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 (471 MHz, CDC13) δ -104.64 (dd, J = 255.1, 7.6 Hz, IF), -108.01 (dd, J = 255.0, 11.8 Hz, IF). The mass spectrum gave the mass-to-charge ratio as m / z: 338.0761 [M+K] Figure 4 as shown in the mass spectrum, the mass-to-charge ratio was m / z: 338.0761 [M+K] + , indicating that the molecular formula was C 18 H 15 F2NO.

[0029] Example 2

[0030] Anti-tumor 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 hepatoma cells (HepG-2) (Cat. No. C6346) were purchased from Biyun Tian Biotechnology.

[0032] Human hepatoma cells (HepG-2) and human cervical cancer cells (HeLa) were cultured in DMEM medium containing 10% fetal bovine serum, 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 bottle, they were trypsinized and subcultured, and the cells in the logarithmic growth phase were selected for the experiment.

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

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

[0035] Logarithmic growth phase tumor cell lines were taken to prepare cell suspensions. The cell suspension concentration was diluted to 5 x 10 4Cells were seeded at 100 μL / mL in each well of a 96-well plate and incubated for 24 h. After incubation, the supernatant was removed, and 100 μL of drug-containing medium was added to each well. The final concentration gradient of compound 2e was 64, 32, 16, 8, 4, and 2 μg / mL. Cisplatin was used as the positive control, and an equal volume of medium and DMSO was added to the blank control group (i.e., single-well cells were cultured without drug administration, using standard methods). Each treatment was performed in triplicate. After culturing at 37°C and 5% CO2 for 48 h, 10 μL of MTT solution (5 mg / mL) was added to each well, and the cells were incubated at 37°C and 5% CO2 for another 4 h. The supernatant was discarded, and 150 μL of DMSO was added to each well. The plates were shaken for 10 min, and the absorbance (OD) was measured at 490 nm using a microplate reader.

[0036] The formula for calculating the inhibition rate of tumor cell proliferation by compound 2e is as follows:

[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%. With increasing concentration, the inhibition rate gradually increased, reaching 76.7% at 64 μg / mL. The positive control cisplatin group showed a higher inhibition rate, at 29.5% at 2 μg / mL and reaching 97.7% at 64 μg / mL.

[0041] Table 1. MTT assay results showing the inhibitory effect of compound 2e on the proliferation of HepG-2 liver cancer cells.

[0042]

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

[0044] The results are shown in Table 2. The inhibition rate of compound 2e against HCT-116 colon cancer cells gradually increased from 11.0% at 2 μg / mL to 99.2% at 64 μg / mL, showing a significant concentration-dependent effect. 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 assay results showing the inhibitory effect of compound 2e on the proliferation of HCT-116 colon 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 assay, 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 in the cisplatin group was 6.9% at 2 μg / mL and 89.2% at 64 μg / mL.

[0049] Table 3. MTT assay results showing the inhibitory effect of compound 2e on the proliferation of HeLa cervical cancer cells.

[0050]

[0051] 4. Inhibitory effect of compound 2e on the 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 in the cisplatin group was 25.2% at 2 μg / mL and 94.5% at 64 μg / mL.

[0053] Table 4. MTT assay results showing the inhibitory effect of compound 2e on the proliferation of MCF-7 breast cancer cells.

[0054]

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

[0056] IC50 of compound 2e and the positive control drug cisplatin on four types of tumor cells 50 The values ​​are shown in Table 5. As can be seen from the results in Table 5, compound 2e of the present invention exhibited inhibitory effects on the proliferation of four different tumor cell lines in the in vitro antitumor activity assay, especially showing significantly better inhibitory effects on human colon cancer cells (HCT-116) than the positive control drug cisplatin (IC50). 50 =15.43 μg / mL), its IC50 is 15.43 μg / mL. 50 The value was 5.67 μg / mL, demonstrating its potential as a novel antitumor drug.

[0057] Table 5. Inhibitory effect of compound 2e on tumor cell proliferation (IC50) 50 μg / mL)

[0058]

[0059] Example 3

[0060] Antibacterial activity test

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

[0062] The antibacterial activity of compound 2e was determined using the microdilution method. The bacterial strains used in the activity test were activated in a 37°C incubator for 2-3 hours. Afterward, a small amount of the activated strain was picked, dissolved in sterile water, and shaken well to prepare a concentration of 5 × 10⁻⁶. 5 CFU / mL bacterial suspension.

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

[0064] 100 μL of drug-containing culture medium was added to each well of a 96-well plate, followed by dilution with 100 μL of bacterial suspension to achieve final concentrations of compound 2e of 64, 32, 16, 8, 4, 2, 1, 0.5, 0.25, and 0.125 μg / mL. Ciprofloxacin was used as a positive control, LB medium containing 0.5% DMSO as a negative control, and LB medium as a blank control. Each experiment was repeated three times. After incubation at 37°C for 24 hours, the turbidity of the bacterial suspension in the 96-well plate was observed. The minimum concentration at which the culture medium became clear was defined as the minimum inhibitory concentration (MIC) of the compound.

[0065] The bacteria whose growth was inhibited were aspirated into 10 μL from each well and spread onto nutrient agar medium. The culture was carried out in a constant temperature incubator at 37°C for 24 h. Each experiment was repeated 3 times. 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 all four strains, with the inhibitory effect on Bacillus subtilis being more significant.

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

[0069]

[0070] In the table, "-" indicates 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 principles of the present invention should be included within the protection scope of the present invention.

Claims

1,3,3-Difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2 H -benzo[ b A method for preparing azepam-2-one, characterized in that, The steps include: at room temperature, adding 2-bromo-2,2-difluoro- N -methyl- N The photolysis reaction was carried out using 4-methyl-2-(1-styryl)phenyl)acetamide, potassium dihydrogen phosphate, 2,4,5,6-tetra(9-carbazolyl)-isophthalonitrile, and tetrahydrofuran, under irradiation with a 30 W blue LED lamp and maintained at 25°C in air. Monitoring was performed by thin-layer chromatography until the reaction proceeded to 2-bromo-2,2-difluoro- N -methyl- N -(4-methyl-2-(1-styryl)phenyl)acetamide was completely consumed; after the reaction was complete, the resulting mixture was quenched and extracted; the organic solvent was concentrated under vacuum, and the residue was purified by rapid silica gel column chromatography, eluting with eluent to obtain the 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2-ethylhexylene ... H -benzo[ b Azazo-2-one, the 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2-one H -benzo[ b The structural formula of azepine-2-one is shown in 2e: 。 2. The 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2-di ... H -benzo[ b A method for preparing azepam-2-one, characterized in that, The 2-bromo-2,2-difluoro N -methyl- N The molar ratio of 4-methyl-2-(1-styryl)phenyl)acetamide to potassium dihydrogen phosphate is 1:1.

5.

3. The 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2-diphenyl-1,3-dihydro-2-diphenyl-1,3-difluorodiphenyl-1,3-dihydrodiphenyl-1,3- ... H -benzo[ b A method for preparing azepam-2-one, characterized in that, The 2-bromo-2,2-difluoro N -methyl- N -(4-methyl-2-(1-styryl)phenyl)acetamide 0.3 mmol, potassium dihydrogen phosphate 0.45 mmol.

4. The 3,3-difluoro-1,7-dimethyl-5-phenyl-1,3-dihydro-2-di ... H -benzo[ b A method for preparing azepam-2-one, characterized in that, The mixture of 0.009 mmol of 2,4,5,6-tetra(9-carbazolyl)-isophthalonitrile and 3 mL of tetrahydrofuran was quenched with 15 mL of saturated sodium bicarbonate aqueous solution and extracted three times with 15 mL of ethyl acetate. The rapid column chromatography purification conditions were as follows: the volume ratio of eluent was petroleum ether:ethyl acetate = 30:1.

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