A dibenzylbutane-type lignan LCA derivative and its application

Abstract

The present invention discloses a dibenzylbutane lignan LCA derivative or a pharmaceutical salt thereof, the structure of which is shown below: #imgabs0# The dibenzylbutane lignan LCA derivative prepared by the present invention has been shown through pharmacological experiments to have significant anti-inflammatory activity, can significantly inhibit the release of inflammatory factors, and can be used to prepare anti-inflammatory drugs for diseases such as rheumatoid arthritis.

Description

Technical Field

[0001] The invention belongs to the field of medical technology, and specifically relates to a dibenzylbutane lignan LCA derivative and application thereof. Background Art

[0002] Douchi ginger is the dried root and rhizome of Litsecub cubeba (Lour.) Pers, a plant of the Lauraceae family. In clinical treatment of traditional Chinese medicine, it has the effects of treating rheumatic pain, traumatic injuries, dispelling wind, cold and dampness, warming the middle and regulating qi and relieving pain (Chinese Materia Medica Editorial Committee of the State Administration of Traditional Chinese Medicine. Chinese Materia Medica, p73.). It is a kind of ethnic medicine Dai medicine and is used as an ethnic Dai medicine in the treatment of rheumatic diseases in southwest my country. Modern medical research shows that the main components of fermented black ginger are alkaloids, volatile oils, lignans, etc. These components are closely related to the anti-tumor, anti-asthmatic, anti-allergic, anti-bacterial, anti-inflammatory and immune effects of fermented black ginger (Zhang W, et al. Antibacterial, Antifungal and Cytotoxic Isoquinoline Alkaloids from Litsea cubeba [J]. Molecules, 2012, 17 (12): 12950-12960.). In recent years, it has been reported that the active ingredient dibenzyl butane lignan LCA in the dichloromethane part of the ethanol extract of Douchi ginger has the effect of reducing proinflammatory cytokines and improving joint swelling in arthritis rats, indicating that the active ingredient LCA in Douchi ginger may become an effective drug for anti-rheumatoid arthritis (Lin B, et al. Inhibitory effects of the root extract of Litsea cubeba (lour.) pers. on adjuvant arthritis in rats [J]. Journal of Ethnopharmacology, 2013, 147 (2): 327-334). Lin Bing et al. first disclosed the study of the use of dibenzyl butane lignan LCA, an active anti-inflammatory ingredient in Douchi ginger, in anti-rheumatoid arthritis drugs. The compound structure of the dibenzyl butane lignan LCA is as follows:

[0003]

[0004] Inflammation is a defensive response of living tissues with vascular systems to damaging factors, and its typical manifestations are redness, swelling, heat, pain and dysfunction. Nowadays, inflammation is considered to be the cause or complex factor of many diseases, such as rheumatoid arthritis, diabetes, obesity, cancer, depression, Alzheimer's disease and cardiovascular disease. The research and development of new structural types of drugs with anti-inflammatory activity has important theoretical significance. Natural products are a huge treasure trove. With the development and innovation of natural product discovery, identification methods, activity screening methods and chemical informatics, natural products, especially compounds derived from plants, will continue to be an important source for drug research and development. However, the anti-inflammatory active ingredient dibenzylbutane lignan LCA in Douchi ginger has the disadvantages of easy hydrolysis and low bioavailability. Therefore, it is of great significance to design derivatives of dibenzylbutane lignan LCA and then find anti-inflammatory compounds with good anti-inflammatory effects, high bioavailability and low toxic side effects. Summary of the invention

[0005] The purpose of the present invention is to provide a dibenzylbutane lignan LCA derivative.

[0006] The second objective of the present invention is to provide a use of the dibenzylbutane lignan LCA derivative in the preparation of anti-inflammatory drugs.

[0007] In order to achieve the above object, the technical solution adopted by the present invention is as follows:

[0008] The first aspect of the present invention provides a dibenzylbutane lignan LCA derivative or a pharmaceutically acceptable salt thereof, the structure of which is shown below:

[0009]

[0010] X is selected from -O-, -CH2-,

[0011] R is selected from C1 to C10 chain alkyl,

[0012] R1 is selected from hydrogen, halogen (fluorine, chlorine, bromine, iodine), C1-C10 alkyl (such as methyl, ethyl, tert-butyl), C1-C10 alkoxy (such as methoxy, ethoxy, tert-butoxy), CF3;

[0013] R2 is selected from hydrogen, halogen (fluorine, chlorine, bromine, iodine), C1-C10 alkyl (such as methyl, ethyl, tert-butyl), C1-C10 alkoxy (such as methoxy, ethoxy, tert-butoxy), CF3;

[0014] R3 is selected from hydrogen, halogen (fluorine, chlorine, bromine, iodine), C1-C10 alkyl (such as methyl, ethyl, tert-butyl), C1-C10 alkoxy (such as methoxy, ethoxy, tert-butoxy), CF3;

[0015] R4 is selected from hydrogen, halogen (fluorine, chlorine, bromine, iodine), C1-C10 alkyl (such as methyl, ethyl, tert-butyl), C1-C10 alkoxy (such as methoxy, ethoxy, tert-butoxy), CF3;

[0016] R5 is selected from hydrogen, halogen (fluorine, chlorine, bromine, iodine), C1-C10 alkyl (such as methyl, ethyl, tert-butyl), C1-C10 alkoxy (such as methoxy, ethoxy, tert-butoxy), CF3;

[0017] Most preferably, the structure of the dibenzylbutane lignan LCA derivative is selected from one of the following structures:

[0018]

[0019]

[0020]

[0021] The second aspect of the present invention provides a use of the dibenzylbutane lignan LCA derivative in the preparation of anti-inflammatory drugs.

[0022] The dibenzylbutane lignan LCA derivative of the present invention can significantly inhibit the release of inflammatory factors, has good anti-inflammatory activity, and can be used as a drug with anti-inflammatory activity.

[0023] The second aspect of the present invention provides a use of the dibenzylbutane lignan LCA derivative in the preparation of a drug for treating rheumatoid arthritis.

[0024] Due to the adoption of the above technical solution, the present invention has the following advantages and beneficial effects:

[0025] The dibenzylbutane lignan LCA derivatives prepared by the present invention have been shown by pharmacological experiments that the compounds have no inhibitory effect on cell growth and no obvious cytotoxicity at a drug concentration of 20 μM, while the control drug LCA has certain cytotoxicity at a concentration of 20 μM. When stimulated by LPS, the proinflammatory factors TNFα and IL-6 of RAW 264.7 cells increased significantly, while the preferred compound 10o can significantly reduce the levels of TNFα and IL-6, and at 20 μM, the inhibitory activity of compound 10o is better than that of the control drug LCA. Under LPS stimulation, cells produce proinflammatory factors TNFα and IL-6, which in turn activate the inflammatory response pathway and lead to an increase in NO synthesis, further exacerbating the inflammatory response. The preferred compound 10o concentration-dependently inhibits LPS-induced NO generation, and the inhibitory activity is stronger than that of the control drug LCA at 20 μM, so the prepared LCA derivatives can be used to prepare anti-inflammatory drugs for diseases such as rheumatoid arthritis. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 It is a schematic diagram of the cell viability (%) results of RAW 264.7 cells when the concentration of the target compound is 20 and 50 μM.

[0027] Figure 2 It is a schematic diagram of the effect of 20 μM of each compound on LPS-induced NO production in RAW264.7 cells.

[0028] Figure 3 This is a schematic diagram of the effects of different concentrations of compound 10o on LPS-induced NO production in RAW264.7 cells.

[0029] Figure 4 This is a schematic diagram of the effects of different concentrations of compound 10o on LPS-induced TNF-α production in RAW264.7 cells.

[0030] Figure 5 This is a schematic diagram of the effects of different concentrations of compound 10o on LPS-induced IL-6 production in RAW264.7 cells. DETAILED DESCRIPTION

[0031] In order to explain the present invention more clearly, the present invention is further described below in conjunction with preferred embodiments. It should be understood by those skilled in the art that the following specific description is illustrative rather than restrictive, and should not be used to limit the scope of protection of the present invention.

[0032] The experimental methods in the following examples without specifying specific conditions are usually carried out under conventional conditions or conditions provided by the manufacturer. All reagents used in the examples are commercially available analytically pure.

[0033] Example 1

[0034] The preparation of compounds 10a-r of the present invention is as follows:

[0035] a) Preparation of key intermediate 8, the reaction route is as follows:

[0036]

[0037]

[0038] b) Preparation of target compounds 10a-r, the reaction scheme is as follows:

[0039]

[0040] The first step is to prepare the intermediate 4-hydroxy-3,5-dimethoxycinnamic acid methyl ester (compound 2)

[0041] Sulfuric acid (98% v / v, 5 mL) was added to a solution of 4-hydroxy-3,5-dimethoxycinnamic acid (compound 1, 25.0 g, 111.6 mmol) in methanol (500 mL) and refluxed for 5 hours. After the reaction was complete, the solvent was removed by vacuum concentration, ethyl acetate and saturated sodium bicarbonate were added for extraction, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and recrystallized to obtain a white solid compound 2 (25.6 g, 96%).

[0042] Step 2: Preparation of intermediate 4-hydroxy-3,5-dimethoxyphenylpropionic acid methyl ester (Compound 3)

[0043] 10% palladium on carbon (1.5 g) was added to a methanol solution (500 mL) of methyl 4-hydroxy-3,5-dimethoxycinnamate (compound 2, 25.6 g, 107.6 mmol), and the mixture was reacted at room temperature for 10 hours in the presence of hydrogen. After the reaction, the palladium on carbon was filtered, the solvent was removed by vacuum concentration, and the mixture was purified by silica gel column chromatography to obtain a white solid compound 3 (24.0 g, 93%).

[0044] Step 3: Preparation of intermediate 4-benzyloxy-3,5-dimethoxyphenylpropionic acid methyl ester (Compound 4)

[0045] Benzyl bromide (25.6 g, 150.0 mmol) and potassium carbonate (27.6 g, 200.0 mmol) were added to a solution of methyl 4-hydroxy-3,5-dimethoxybenzenepropanoate (compound 3, 24.0 g, 100.0 mmol) in acetone (500 mL), and the mixture was refluxed for 5 hours. After the reaction, the mixture was filtered and concentrated under reduced pressure to remove the solvent, and purified by silica gel column chromatography to obtain white solid compound 4 (30.0 g, 91%).

[0046] Step 4: Preparation of intermediate 4-benzyloxy-3,5-dimethoxyphenylpropionic acid (Compound 5)

[0047] To a solution of 4-benzyloxy-3,5-dimethoxybenzenepropanoic acid methyl ester (compound 4, 30.0 g, 90.9 mmol) in methanol (500 mL) was added sodium hydroxide solution (5.5 mol / L, 50 mL), and the mixture was refluxed for 3 hours. After the reaction was complete, concentrated hydrochloric acid was added dropwise to adjust the pH value of the reaction solution to 3-4, and dichloromethane was added for extraction. The mixture was dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The mixture was purified by silica gel column chromatography to obtain white solid compound 5 (25.5 g, 89%).

[0048] Step 5: Preparation of intermediate (S)-4-benzyl-3-[3-(4-benzyloxy-3,5-dimethoxy)phenylpropionyl]oxazolidin-2-one (Compound 6)

[0049] 4-Benzyloxy-3,5-dimethoxybenzenepropionic acid (compound 5, 25.2 g, 79.7 mmol), (S)-4-benzyl-2-oxazolidinone (15.6 g, 87.8 mmol), 4-dimethylaminopyridine (1.0 g, 7.97 mmol) and 150 mL of anhydrous dichloromethane were added to the reaction flask, and an anhydrous dichloromethane solution (150 mL) of N,N'-dicyclohexylcarbodiimide (16.4 g, 79.7 mmol) was added under argon protection in an ice bath. After the addition, the reaction was slowly heated to room temperature and reacted for 24 hours. The organic phase was extracted with dichloromethane, dried over anhydrous sodium sulfate, filtered and the solvent was removed under reduced pressure, and purified by silica gel column chromatography to obtain a white solid compound 6 (24.2 g, 72%).

[0050] Step 6: Preparation of the intermediate (2R,3S)-1,4-bis[(S)-4-benzyl-2-keto-3-oxazolidinyl]-2,3-bis[(4-benzyloxy-3,5-dimethoxy)benzyl]butane-1,4-dione (Compound 7)

[0051] Compound 6 (24.2 g, 56.7 mmol) and anhydrous tetrahydrofuran (500 mL) were added to a reaction flask, cooled to -78 ° C, and lithium diisopropylamide (2.0 M, 56.7 mL, 113.3 mmol) was added. After reacting for 1 hour, iodophenyl diacetic acid (36.7 g, 113.3 mmol) was added dropwise to the reaction solution, and the reaction was continued at -78 ° C for 1 hour. Then hydrochloric acid (1 M) was added to quench the reaction, and ethyl acetate was added for extraction. The organic layer was washed with dilute hydrochloric acid, 10% sodium thiosulfate solution and saturated sodium bicarbonate aqueous solution in turn, and the organic phase was dried over anhydrous sodium sulfate. After filtering and removing the solvent under reduced pressure, it was purified by silica gel column chromatography to obtain white solid compound 7 (15.2 g, 63%).

[0052] Step 7: Preparation of intermediate (2R,3S)-2,3-bis[(4-benzyloxy-3,5-dimethoxy)benzyl)]succinic acid (Compound 8)

[0053] At 0°C, 30% hydrogen peroxide solution (43.6 mL, 35 mmol) and lithium hydroxide monohydrate (588 mL, 14 mmol) were added to a mixed solution of compound 7 (3.00 g, 3.5 mmol) in tetrahydrofuran (33 mL) and water (20 mL). After 15 minutes, the reaction solution was slowly warmed to room temperature and stirred overnight. After the reaction was complete, 10% hydrochloric acid was used to adjust the pH value to 3, dichloromethane was added for extraction, the organic phase was dried over anhydrous sodium sulfate, filtered and the solvent was removed under reduced pressure, and then purified by silica gel column chromatography to obtain white solid compound 8 (1.87 g, 85%).

[0054] Step 8: Preparation of the target compound (2R, 3S)-2,3-bis[(4-hydroxy-3,5-dimethoxy)benzyl)]-1,4-bis(4-substituted phenylpiperazine-1-yl)butane-1,4-dione

[0055] 1) Preparation of the intermediate (2R,3S)-2,3-bis[(4-benzyloxy-3,5-dimethoxy)benzyl)]-1,4-bis(4-phenylpiperazine-1-yl)butane-1,4-dione (Compound 9a)

[0056] Benzotriazole-N,N,N',N'-tetramethyluronium hexafluorophosphate (216.2 mg, 0.57 mmol), N,N-diisopropylethylamine (248.1 mg, 1.92 mmol) and 1-phenylpiperazine (116.8 mg, 0.72 mmol) were added to a solution of compound 8 (150 mg, 0.24 mmol) in N,N-dimethylformamide (1.5 mL), and the mixture was reacted for 4 hours under argon protection. After the reaction was complete, the mixture was extracted with ethyl acetate, the organic phase was dried over anhydrous sodium sulfate, filtered, the solvent was removed under reduced pressure, and purified by silica gel column chromatography to obtain compound 9a (170 mg, 77%) as a white solid.

[0057] 2) Preparation of the target compound (2R,3S)-2,3-bis[(4-hydroxy-3,5-dimethoxy)benzyl)]-1,4-bis(4-phenylpiperazine-1-yl)butane-1,4-dione (Compound 10a)

[0058] 10% palladium on carbon (20 mg) was added to a solution of compound 9a (170 mg, 0.19 mmol) in methanol (20 mL), and the mixture was reacted at room temperature for 3 hours in the presence of hydrogen. After the reaction was complete, the palladium on carbon was filtered and the solvent was removed under reduced pressure, and the mixture was purified by medium pressure preparative chromatography to obtain compound 10a (120 mg, 86%) as a white solid.

[0059] The preparation method of compound 10b-r is the same as above, except that the raw material 1-phenylpiperazine in the eighth step is replaced by 1-(2-fluorophenyl)piperazine, 1-(2-chlorophenyl)piperazine, 1-(3-chlorophenyl)piperazine, 1-(4-chlorophenyl)piperazine, 1-(2-bromophenyl)piperazine, 1-(3-bromophenyl)piperazine, 1-(4-bromophenyl)piperazine, 1-(2-methylphenyl)piperazine, 1-(4-methylphenyl)piperazine, 1-(3-methoxyphenyl)piperazine, 1-(2-trifluoromethylphenyl)piperazine, 1-(4-trifluoromethylphenyl)piperazine, 1-(2,3-dichlorophenyl)piperazine, 1-(2,4-difluorophenyl)piperazine, 1-methylpiperazine, morpholine, piperidine, and the rest is the same as in Example 1, to obtain compounds 10b-r respectively.

[0060] The structure, yield, H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of the compound of the present invention are shown in Table 1:

[0061]

[0062] Table 1 Yield, H NMR spectrum, C NMR spectrum and high resolution mass spectrum data of the compounds of the present invention

[0063]

[0064]

[0065]

[0066]

[0067]

[0068] Example 2

[0069] Pharmacological experiments of the compounds prepared by the present invention

[0070] I. Cytotoxicity experiments of compounds

[0071] 1. Experimental method: CCK-8 method was used to determine the cytotoxicity of the target compound.

[0072] 1) Cell selection: mouse RAW 264.7 cells.

[0073] 2) Preparation of drugs and reagents: Accurately weigh the sample to be tested, add it to a 0.5 mL centrifuge tube, and add the corresponding amount of DMSO to make 2×10 4 μM drug concentration, stored at 4°C. When used, take an appropriate amount of high-glucose complete medium with 10% fetal bovine serum and dilute to 20 and 50 μM respectively.

[0074] 3) Cell culture and passaging: All cells were cultured in high-glucose complete medium containing 10% fetal bovine serum in a 37°C, 5% CO2 incubator. After the cells covered the bottom of the dish, the medium was aspirated, washed twice with sterile PBS, and 3 mL of complete medium was added. The cells were pipetted with a pipette, centrifuged, and divided into 3 new culture dishes. 8 mL of complete medium was added and cultured continuously.

[0075] 4) Drug treatment: RAW 264.7 cells cultured to the logarithmic growth phase were collected and the cell concentration was adjusted to a certain concentration (the concentration of mouse mononuclear macrophage RAW 264.7 was 5×10 5 / mL), add 100μL of cell suspension to each well of a 96-well cell culture plate. Place the culture plate in a carbon dioxide incubator for 24 hours, remove the original culture medium, add 100μL of fresh culture medium containing 20 and 50μM drugs, set 6 replicates for each concentration, and set up 6 replicates for the control group (with cells), blank group (without cells), and positive control drug LCA group, and continue to culture in a carbon dioxide incubator for 24 hours. Take out the culture plate and add 10μL of CCK-8 reagent to each well. Place the 96-well plate in a carbon dioxide incubator and continue to culture for 30 minutes. The absorbance value is measured by an enzyme reader at 450nm to calculate the inhibition rate of the drug on cell proliferation.

[0076] Inhibition rate = [(OD 对照孔 -OD 实验孔 ) / (OD 对照孔 -OD 空白孔 )]×100%.

[0077] 2. Test results

[0078] The inhibition rate of the target compound on the proliferation of RAW 264.7 cells is shown in Figure 1 As shown, Figure 1 The figure is a schematic diagram of the cell viability (%) results of RAW 264.7 cells when the target compound concentration is 20 and 50 μM. It can be seen from the figure that 20 μM of the compound of the present invention has no obvious cytotoxicity to RAW 264.7 cells, which provides a basis for the subsequent dose design of the anti-inflammatory activity screening experiment.

[0079] 2. Effects of compounds on LPS-induced nitric oxide (NO) production

[0080] 1. Experimental method: Griess method was used to determine the effect of target compounds on LPS-induced NO production

[0081] 1) Preparation of drugs and reagents: Accurately weigh the sample to be tested, add it to a 0.5 mL centrifuge tube, and add the corresponding amount of DMSO to make 2×10 4 μM drug concentration, stored at 4°C. When used, take an appropriate amount of high-glucose complete medium with 10% fetal bovine serum and dilute to 20 μM.

[0082] 2) Cell culture and passaging: All cells were cultured in high-glucose complete medium containing 10% fetal bovine serum in a 37°C, 5% CO2 incubator. After the cells covered the bottom of the dish, the medium was aspirated, washed twice with sterile PBS, and 3 mL of complete medium was added. The cells were pipetted with a pipette, centrifuged, and divided into 3 new culture dishes. 8 mL of complete medium was added and cultured continuously.

[0083] 3) Drug treatment: RAW 264.7 cells cultured to the logarithmic growth phase were collected and the cell concentration was adjusted to a certain concentration (the concentration of mouse mononuclear macrophage RAW 264.7 was 5×10 5 / mL), add 100μL of cell suspension to each well of a 96-well cell culture plate. Place the culture plate in a carbon dioxide incubator for 24 hours, remove the original culture medium, add 100μL of fresh culture medium containing 20μM of the drug, set up 6 replicates for each concentration, and set up 6 replicates for the control group (with cells, no LPS stimulation), model group (LPS stimulation), and positive control drug LCA group, and continue to culture in a CO2 incubator for 4 hours. Take out the culture plate, remove the original culture solution, prepare 20μM of the drug with a culture medium containing LPS (100ng / mL), and add 200μL to each well of the 96-well plate. Place the 96-well plate in an incubator and continue to culture for 24 hours. Add 100μL / well of the sample to each new 96-well plate, and add Griess Reagent I and Griess Reagent II at 50μL / well. The absorbance value was measured by an enzyme-labeled instrument at a wavelength of 540 nm, and the NO concentration in the sample was calculated based on the OD value of each well and the standard curve.

[0084] 2. Test results

[0085] Effects of target compounds on LPS-induced NO production in RAW264.7 cells are shown in Figure 2 As shown, Figure 2 The figure is a schematic diagram of the effect of 20 μM compounds on the NO production of RAW264.7 cells induced by LPS. It can be seen from the figure that most of the target compounds have an inhibitory effect on the NO production of RAW 264.7 cells induced by LPS, among which compounds 10c, 10l, 10o, 10p and 10q have stronger inhibitory effects, and 10o has the strongest inhibitory effect.

[0086] 3. Effects of different concentrations of compound 10o on LPS-induced NO production in RAW 264.7 cells Figure 3 As shown, Figure 3 Schematic diagram of the effect of different concentrations of compound 10o on NO production in RAW264.7 cells induced by LPS. For the preferred compound 10o, the effect of different concentrations on NO production in RAW 264.7 cells induced by LPS was further investigated using the same method as above.

[0087] NO participates in platelet regulation and immune system response in the body, and plays a key regulatory role in inflammation. When it is expressed in large quantities, it will cause vasodilation, increase in exudates such as plasma and leukocytes, and cause vascular and tissue damage. Therefore, the content of NO can be used as a standard for assessing the severity of inflammation. As can be seen from the figure, different concentrations of 10o (1.25, 2.5, 5, 10, 20 μM) have an inhibitory effect on LPS-induced NO production in RAW 264.7 cells, and are dose-dependent. As the concentration increases, the inhibitory effect increases. Therefore, the target compound can inhibit the production of NO and has anti-inflammatory activity.

[0088] Effect of compound 10o on LPS-induced production of pro-inflammatory factors TNF-α and IL-6

[0089] 1. Experimental method: ELISA method was used to determine the effect of the target compound on LPS-induced production of pro-inflammatory factors TNF-α and IL-6

[0090] 1) Preparation of drugs and reagents: Accurately weigh the sample to be tested, add it to a 0.5 mL centrifuge tube, and add the corresponding amount of DMSO to make 2×10 4 μM drug concentration, stored at 4°C. When used, take an appropriate amount of high-glucose complete medium with 10% fetal bovine serum and dilute to 1.25, 2.5, 5, 10, 20 μM respectively.

[0091] 2) Cell culture and passaging: All cells were cultured in high-glucose complete medium containing 10% fetal bovine serum in a 37°C, 5% CO2 incubator. After the cells covered the bottom of the dish, the medium was aspirated, washed twice with sterile PBS, and 3 mL of complete medium was added. The cells were pipetted with a pipette, centrifuged, and divided into 3 new culture dishes. 8 mL of complete medium was added and cultured continuously.

[0092] 3) Drug treatment: RAW 264.7 cells cultured to the logarithmic growth phase were collected and the cell concentration was adjusted to a certain concentration (the concentration of mouse mononuclear macrophage RAW 264.7 was 1.6×10 5 / mL), add 5mL of cell suspension to each well of a 24-well cell culture plate. Place the culture plate in a carbon dioxide incubator for 24 hours, remove the original culture medium, add 5mL of fresh culture medium containing 1.25, 2.5, 5, 10, and 20μM of each drug, set up 6 replicates for each concentration, and set up 6 replicates for the positive control drug LCA group (20μM) and the model group (LPS stimulation), and continue to culture in a CO2 incubator for 2 hours. Take out the culture plate, remove the original culture solution, prepare 1.25, 2.5, 5, 10, and 20μM drugs with a culture medium containing LPS (100ng / mL), and add 5mL to each well of the 24-well plate. Place the 24-well plate in an incubator and continue to culture for 4 hours. Determine the content of IL-6 and TNF-α according to the method provided by the ELISA kit (Xinbosheng).

[0093] 2. Test results

[0094] The effects of compound 10o on LPS-induced TNF-α and IL-6 production in RAW264.7 cells are shown in Figure 4 and Figure 5 , Figure 4 This is a schematic diagram of the effects of different concentrations of compound 10o on LPS-induced TNF-α production in RAW264.7 cells. Figure 5 This is a schematic diagram of the effect of different concentrations of compound 10o on the production of IL-6 in RAW264.7 cells induced by LPS. TNF-α and IL-6 are both important pro-inflammatory factors in inflammatory responses. Inhibiting the production of pro-inflammatory factors can inhibit the progress of inflammatory responses. Figure 4 and Figure 5 It can be seen that compared with the inflammation model group, the levels of TNF-α and IL-6 in compound 10o were significantly reduced, indicating that the compound can inhibit the production of proinflammatory factors TNF-α and IL-6, thereby inhibiting the progress of the inflammatory response. Therefore, the target compound has anti-inflammatory activity and can be used to prepare anti-inflammatory drugs.

[0095] The above is only a preferred embodiment of the present invention, and does not limit the present invention in any form. Although the present invention has been disclosed as a preferred embodiment, it is not used to limit the present invention. Any technician familiar with this patent can make some changes or modify the technical contents suggested above into equivalent embodiments without departing from the scope of the technical solution of the present invention. However, any simple modification, equivalent change and modification made to the above embodiments according to the technical essence of the present invention without departing from the content of the technical solution of the present invention still fall within the scope of the solution of the present invention.

Claims

1. A dibenzylbutane lignan LCA derivative or a pharmaceutically acceptable salt thereof, characterized in that: The structure of the dibenzylbutane lignan LCA derivative is selected from one of the following structures:

2. Use of the dibenzylbutane lignan LCA derivative according to claim 1 in the preparation of anti-inflammatory drugs.

3. Use of the dibenzylbutane lignan LCA derivative according to claim 1 in the preparation of a medicament for treating rheumatoid arthritis.

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