Preparation method of flavonoid compound GL-V8

Abstract

The invention discloses a preparation method of a flavonoid compound GL-V8, which comprises the following steps: protecting diethanol amine by 2, 2-dimethoxypropane, carrying out nucleophilic substitution reaction with 1, 4-halogenated butane, reacting with wogonin under an alkaline condition, and removing a protecting group under an acidic condition to obtain a final product GL-V8, the purity of the final product after recrystallization is greater than 99.5%; the method has the advantages of mild reaction conditions, short line, low material cost and convenience in industrial large-scale production.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical technology, and particularly relates to a preparation method of a flavonoid compound GL-V8. Background Art

[0002] GL-V8 belongs to a derivative of the flavonoid wogonin, with the molecular formula C 24 H 29 NO7, and its chemical name is 5-hydroxy-7-(4-pyrrolidin-1-yl)butoxy-8-methoxyflavone. Its structural formula is shown in formula (I):

[0003]

[0004] GL-V8 is a small molecule compound obtained by semi-synthesis of natural products, and has the characteristics of safety, low toxicity, and low cost of natural products. Experimental studies have shown that GL-V8 has significant inhibitory effects on various tumor cells such as T-lymphocyte leukemia, colorectal cancer, and pancreatic cancer in vitro and in vivo. Its preliminary mechanism of action is to mediate lysosome-dependent cell death through lysosomal damage, thereby selectively killing malignant tumor cells, and having no obvious toxic and side effects on normal tissues and cells, which is not possessed by general chemotherapy drugs. GL-V8 can inhibit the growth of various tumor cells in vitro, induce obvious apoptosis of tumor cells, and at the same time also has an obvious apoptosis-inducing effect on clinical T-cell lymphoma samples with large individual differences, showing time- and dose-dependence, indicating that GL-V8 has a strong anti-tumor effect in vitro. In vitro studies have shown that GL-V8 has no killing effect on human normal cell lines NCM460, HLEC, HK2, and 293T at anti-tumor effective concentrations, suggesting that GL-V8 has good selectivity for tumor cells, which is of great significance for reducing toxic and side effects in clinical drug use.

[0005] HSP70 is an ATP-dependent molecular chaperone protein, which has the function of coping with various physiological and environmental stimuli (including the damage of chemotherapy drugs to cells), and helps cells survive under fatal conditions. Although HSP70 is highly expressed in tumor cells and is involved in the generation of chemotherapy resistance, its own important physiological functions limit the research and development of it as a tumor treatment drug target. Different from traditional HSP70 inhibitors, GL-V8, as a lysosome-targeted HSP70 inhibitor, only produces an inhibitory effect on HSP70 proteins at specific organelle sites, thereby avoiding the generation of toxic and side effects such as immunosuppression and myelosuppression. At the same time, due to the differences in the number and structure of lysosomes between normal cells and tumor cells, GL-V8 shows obvious cell selectivity, and thus is expected to become a highly potential anti-tumor candidate drug.

[0006] Currently, traditional preparation methods usually involve techniques such as organic synthesis and extraction and modification of natural products. The extraction methods of natural products have limited raw material sources, and existing organic syntheses face problems such as long reaction steps and low yields. For example, the prior art CN11665576A discloses a preparation method of GL-V9. On this basis, in the prior art, the end product GL-V8 can be obtained by replacing the side chain, and the synthetic route is as follows:

[0007]

[0008] This route uses chrysin as the starting material and obtains the target product GL-V8 through five steps of reactions including Elb oxidation, alkylation, hydrolysis, methylation, and nucleophilic substitution. The synthetic route is long, the yield is low, and the operation is cumbersome, which is not suitable for large-scale industrial production.

[0009] Therefore, there is an urgent need to seek a synthetic method for preparing flavonoid GL-V8 that is simple to operate, environmentally friendly, low-cost, and suitable for large-scale industrial production. Summary of the Invention

[0010] In view of the deficiencies in the prior art, the present invention provides a preparation method of flavonoid GL-V8 that is simple and stable to operate, has a high yield, is environmentally friendly, has a low production cost, and is suitable for industrial-scale production.

[0011] To achieve the above object, the present invention adopts the following technical solutions:

[0012] A preparation method of flavonoid GL-V8, comprising: step (a) reacting diethanolamine with 2,2-dimethoxypropane to obtain compound 1; step (b) subjecting compound 1 to nucleophilic substitution with 1,4-halobutane to obtain compound 2; step (c) reacting compound 2 with wogonin under alkaline conditions to obtain compound 3; step (d) deprotecting compound 3 under acidic conditions to obtain the end product GL-V8, and the purity of the end product is greater than 99.5% after recrystallization and purification.

[0013] The synthetic route is as follows:

[0014]

[0015] X and Y are halogen atoms;

[0016] The preparation method comprises the following steps:

[0017] Step (a): Diethanolamine (SM1) forms a diethanolamine salt in the presence of an acid. The diethanolamine salt, 2,2-dimethoxypropane (SM2), and a catalyst react to form a compound represented by Formula 1. The reaction temperature is 25 - 100 °C, and the reaction time is 1 - 10 hours. The acid is hydrochloric acid, and the catalyst is p-toluenesulfonic acid. Among them, the molar ratio of diethanolamine, 2,2-dimethoxypropane, and the catalyst is 1:2 - 5:0.01 - 0.05;

[0018] Step (b): The compound represented by Formula 1 reacts with 1,4-halobutane in the presence of a base to form a compound represented by Formula 2. The reaction temperature is 30 - 40 °C, and the reaction time is 5 - 7 hours. Among them, the equivalent ratio of the compound represented by Formula 1 to 1,4-halobutane is 1:1 - 1:5,

[0019] Step (c): The compound represented by Formula 2 reacts with wogonin in the presence of a base to form a compound represented by Formula 3. The reaction temperature is 80 - 90 °C, and the reaction time is 2 - 6 hours. Among them, the equivalent ratio of the compound represented by Formula 2 to wogonin is 1:1 - 10:1;

[0020] Step (d): The compound represented by Formula 3 reacts in the presence of an acid to form the final product GL-V8. The reaction temperature is 30 - 100 °C, and the reaction time is 2 - 3 hours.

[0021] Furthermore,

[0022] Step (a): SM1 reacts with SM2 to form a compound represented by Formula 1

[0023]

[0024] Dissolve diethanolamine in a reaction solvent, first react with an acid to form a salt, add 2,2-dimethoxypropane and a catalyst for reflux reaction, filter, add an organic solvent to the filter cake, and adjust to alkaline to obtain 2,2-dimethyl-1,3-dioxo-6-azacyclooctane.

[0025] Among them, the acid is selected from hydrochloric acid, sulfuric acid, phosphoric acid, nitric acid, hydrofluoric acid, preferably hydrochloric acid; the equivalent of diethanolamine to the acid is 1:1 to 1:1.5, preferably 1:2; the reaction solvent is selected from dichloromethane, ethyl acetate, methyl tert-butyl ether, acetonitrile, acetone, methanol, ethanol, 1,4-dioxane, water and their mixed solvents, preferably methanol or 1,4-dioxane; the dosage of the organic solvent is 5 to 100 times the volume (V / m) of the reactants, preferably 5 to 7 times the volume (V / m); the catalyst is selected from p-toluenesulfonic acid or ammonium chloride, preferably p-toluenesulfonic acid; the molar ratio of diethanolamine, 2,2-dimethoxypropane to the catalyst is 1:2 to 5:0.01 to 0.05; for adjusting to alkaline, the base is selected from sodium bicarbonate, potassium carbonate, sodium carbonate, potassium bicarbonate, sodium hydroxide or lithium hydroxide, preferably sodium hydroxide.

[0026] Further,

[0027] Step (b): The compound shown in Formula 1 undergoes a nucleophilic substitution reaction with 1,4-halobutane to form the compound shown in Formula 2

[0028]

[0029] Dissolve the compound shown in Formula 1 and 1,4-halobutane in a reaction solvent, add an aqueous solution of a base, and heat for reaction. After the reaction is complete, add water to the reaction mixture, extract with a solvent, and concentrate to obtain the compound shown in Formula 2.

[0030] Among them, X and Y are halogen atoms, selected from fluorine atom, chlorine atom, bromine atom or iodine atom; the reaction solvent is selected from tetrahydrofuran, 2-methyltetrahydrofuran, N,N-dimethylformamide, acetonitrile, acetone, dichloromethane, dimethyl sulfoxide, methanol, water and their mixed solvents, preferably N,N-dimethylformamide; the heating reaction temperature is 30 to 100 °C, preferably 30 to 40 °C; the equivalent of the compound shown in Formula 1 to 1,4-halobutane is 1:1 to 1:5, preferably 1:1.5; the base is selected from potassium carbonate, sodium carbonate, sodium bicarbonate, potassium bicarbonate, sodium hydroxide, potassium hydroxide, tetramethylammonium hydroxide, preferably potassium carbonate; the extraction solvent is selected from 2-methyltetrahydrofuran, dichloromethane, ethyl acetate, isopropyl acetate, isopropyl ether, diethyl ether, preferably ethyl acetate.

[0031] Further,

[0032] Step (c): The compound shown in Formula 2 reacts with wogonin in the presence of a base to form the compound shown in Formula 3

[0033]

[0034] Dissolve the compound shown in Formula 2 and wogonin in an organic solvent, add a base, and heat for reaction. After the reaction is complete, filter to obtain 2-phenyl-5-hydroxy-7-[4-(2,2-dimethyl-1,3,6-dioxazocan-6-yl)butoxy]-8-methoxy-4H-chromen-4-one.

[0035] Among them, the organic solvent is selected from tetrahydrofuran, 2-methyltetrahydrofuran, N,N-dimethylformamide, acetonitrile, acetone, dichloromethane, dimethyl sulfoxide, methanol, water and their mixed solvents, preferably dimethyl sulfoxide; the equivalent ratio of the compound shown in Formula 2 to wogonin is 1:1 to 10:1, preferably 1:1 to 3:1; the base is selected from potassium carbonate, sodium carbonate, sodium bicarbonate, potassium bicarbonate, sodium hydroxide, potassium hydroxide, tetramethylammonium hydroxide, preferably potassium carbonate; the heating reaction temperature is 30 to 100 °C, preferably 80 to 90 °C.

[0036] Furthermore,

[0037] Step (d): The compound shown in Formula 3 is deprotected in the presence of an acid to generate the final product GL-V8

[0038]

[0039] Add the compound shown in Formula 3 to an acid and a solvent, heat to a high temperature for reaction, and process to obtain the final product GL-V8.

[0040] Among them, the acid is formic acid, hydrochloric acid, acetic acid, trifluoroacetic acid, sulfuric acid, preferably acetic acid; the solvent is selected from acetonitrile, methanol, ethanol, tetrahydrofuran, 2-methyltetrahydrofuran, water and their mixed solvents, preferably tetrahydrofuran.

[0041] Advantages of the present invention:

[0042] 1) The process route of the present invention is short, the operation steps are simple, the overall yield is significantly improved compared with the existing process, the production cost is greatly reduced, and the market competitiveness of the product is improved.

[0043] 2) The present invention avoids the use of toxic reagents such as dimethyl sulfate and hydrobromic acid in the existing synthesis process, and has the advantages of environmental friendliness and green environmental protection.

[0044] 3) The step involving the use of flavonoid raw materials in the present invention is at the end of the process route, significantly reducing the material cost and improving the utilization rate.

[0045] 4) The reaction process and post-treatment involved in the present invention are simple, the process reproducibility is good, and it can be mass-produced industrially. Description of the Drawings

[0046] Figure 1This is the 1H-NMR spectrum of GL-V8 in Example 1.

[0047] Figure 2 This is the HRMS spectrum of GL-V8 in Example 1.

[0048] Figure 3 This is the HPLC spectrum of GL-V8 in Example 1. DETAILED DESCRIPTION

[0049] In order to further illustrate the present invention, a series of examples are given below. These examples are purely illustrative and are only used to specifically describe the present invention, and should not be understood as limiting the present invention.

[0050] The experimental methods in the following examples of the present invention are generally carried out under conventional conditions, with specific conditions being noted.

[0051] Unless otherwise specified, the raw materials or reagents used in the following examples of the present invention are commercially available.

[0052] The room temperature described in the following examples of the present invention is 20-35° C. Unless otherwise specified, the reagents are used directly without purification. The reaction is analyzed by TLC or HPLC, and the termination of the reaction is determined by the consumption of the starting materials.

[0053] Example 1

[0054] Step a: Synthesis of Intermediate 1.

[0055]

[0056] First, diethanolamine (5 kg, 47.58 mol, 1.0 eq) was added to 25 L of methanol, and hydrogen chloride (1.2 eq) was introduced at 0-10°C. The resulting solution was stirred at room temperature for 1 hour, and concentrated under reduced pressure to obtain diethanolamine hydrochloride, which can be used for the reaction.

[0057] Dissolve diethanolamine hydrochloride (3kg, 21.19mol, 1.0eq) in 15L of 1,4-dioxane, add 2,2-dimethoxypropane (11kg, 105.62mol, 5.0eq) and anhydrous p-toluenesulfonic acid (110g, 638.79mmol, 0.03eq) in turn, and heat the resulting solution to 80-85°C and stir for 3-4 hours. Filter, wash the filter cake with an appropriate amount of acetone, dissolve it in dichloromethane, adjust the pH value to 9-10 with 2N sodium hydroxide aqueous solution, separate the organic layer, extract the aqueous phase with dichloromethane, combine the organic layers, and concentrate under reduced pressure to remove the solvent to obtain 3.84kg of light yellow oil, with a yield of 92%. HRMS (ESI) m / z calculated for C7H 15NO2[M+H] + 146.1176, found 146.1175.

[0058] Step b: Synthesis of Intermediate 2

[0059]

[0060] Dissolve Intermediate 1 (2 kg, 13.68 mol, 1.0 eq) and 1-bromo-4-chlorobutane (3.5 kg, 20.41 mol, 1.5 eq) in 10 L of DMF, add an aqueous potassium carbonate solution (3.8 kg, 27.50 mol, 2.00 eq, dissolved in 2 L of water), heat the resulting solution to 30 - 40 °C and stir for reaction for 5 - 7 hours. After the reaction is complete, cool to room temperature, add 12 L of water to the reaction mixture, extract with ethyl acetate, separate the organic layer, and concentrate under reduced pressure to obtain 2.42 kg of a yellow oil, with a yield of 75%. HRMS (ESI) m / z calculated for C 11 H 22 ClNO2[M+H] + 236.1412.1255, found 236.1410.

[0061] Step c: Synthesis of Intermediate 3

[0062]

[0063] Under nitrogen protection, dissolve wogonin (1 kg, 3.52 mol, 1.0 eq), Intermediate 2 (3.4 kg, 14.42 mol, 3 eq), and potassium carbonate (2.0 kg, 14.47 mol, 4 eq) in 5 L of dimethyl sulfoxide, stir well, heat to 80 °C, and hold for reaction for 3 hours. After the reaction is complete, cool to room temperature, filter, add 7.5 L of water to the filtrate, stir at room temperature for 30 minutes, then filter by suction to obtain a filter cake, and dry to obtain 1.32 kg of a yellow solid, with a yield of 80%. HRMS (ESI) m / z calculated for C 27 H 33 NO7[M+H] + 484.2330, found 484.2331.

[0064] Step d: Synthesis of GL-V8

[0065]

[0066] Dissolve 1.2 kg of Intermediate 3 in a mixed solution of formic acid (3.6 kg, 78.21 mol) and methanol (1 L), heat up to 80 °C, and react for 2 - 3 hours. After the reaction is complete, concentrate under reduced pressure to obtain a yellow solid, recrystallize with methanol / water, and dry to obtain 950 g of GL-V8 solid with a yield of 86%. HRMS (ESI) m / z calculated for C 24 H 29 NO7[M+H] + 444.2017, found 444.2004. 1 H NMR (300 MHz, Chloroform-d) δ 12.59 (s, 1H), 8.00 - 7.98 (m, 2H), 7.61 - 7.58 (m, 3H), 6.73 (s, 1H), 6.46 (s, 1H), 4.18 (t, J = 12.0 Hz, 2H), 3.98 (s, 3H), 3.78 (t, J = 9.0 Hz, 1H), 2.88 - 2.82 (m, 6H), 2.09 - 1.95 (m, 2H), 1.86 - 1.83 (m, 2H).

[0067] Total yield of the four-step reaction: 47.5%

[0068] Example 2

[0069] Compared with Example 1, the difference lies in step a:

[0070] Dissolve diethanolamine hydrochloride (800 g, 5.65 mol, 1.0 eq) in 4 L of acetone, successively add 2,2-dimethoxypropane (2.6 kg, 24.96 mol, 4.5 eq), and anhydrous p-toluenesulfonic acid (30 g, 174.21 mmol, 0.03 eq). Heat the resulting solution to 80 - 90 °C and stir for 2 - 5 hours. Filter, wash the filter cake with an appropriate amount of acetone, dissolve it in dichloromethane, adjust the pH value to 9 - 10 with 2N sodium hydroxide aqueous solution, separate the organic layer, extract the aqueous phase with dichloromethane, combine the organic layers, and concentrate under reduced pressure to remove the solvent to obtain Intermediate 1 (738 g, yield 90%).

[0071] Example 3

[0072] Compared with Example 1, the difference lies in step a:

[0073] Dissolve diethanolamine hydrochloride (500 g, 3.53 mol, 1.0 eq) in 1.2 L of N,N-dimethylformamide. Then, add 2,2-dimethoxypropane (1.47 kg, 14.11 mol, 4.0 eq) and anhydrous p-toluenesulfonic acid (18 g, 104.52 mmol, 0.03 eq) successively. Heat the resulting solution to 90 - 100 °C and stir for 1 - 2 hours. Filter, wash the filter cake with an appropriate amount of acetone, dissolve it in dichloromethane, adjust the pH value to 9 - 10 with 2N aqueous sodium hydroxide solution, separate the organic layer, extract the aqueous phase with dichloromethane, combine the organic layers, and concentrate under reduced pressure to remove the solvent to obtain Intermediate 1 (451 g, yield 88%).

[0074] Example 4

[0075] Compared with Example 1, the difference lies in step c:

[0076] Under nitrogen protection, dissolve wogonin (200 g, 703.57 mmol, 1.0 eq), Intermediate 2 (330 g, 1.40 mol, 2.0 eq), and potassium carbonate (290 g, 2.10 mol, 3 eq) in 1.4 L of acetone, stir well, heat to 90 °C, and keep the reaction for 2 hours. After the reaction is complete, cool to room temperature, filter, add 2 L of water to the filtrate, stir at room temperature for 30 minutes, then filter by suction to obtain the filter cake, and dry to obtain Intermediate 3 (255 g, yield 75%).

[0077] Example 5

[0078] Compared with Example 1, the difference lies in step c:

[0079] Under nitrogen protection, dissolve wogonin (150 g, 527.68 mmol, 1.0 eq), Intermediate 2 (311 g, 1.32 mol, 2.5 eq), and potassium carbonate (290 g, 2.10 mol, 3 eq) in 1 L of N,N-dimethylformamide, stir well, heat to 80 °C, and keep the reaction for 4 hours. After the reaction is complete, cool to room temperature, filter, add 1.5 L of water to the filtrate, stir at room temperature for 30 minutes, then filter by suction to obtain the filter cake, and dry to obtain Intermediate 3 (194 g, yield 76%).

[0080] Example 6

[0081] Compared with Example 1, the difference lies in step d:

[0082] Dissolve 120 g of Intermediate 3 in a mixed solution of trifluoroacetic acid (600 ml) and tetrahydrofuran (600 ml), heat to 50 °C, and react for 5 - 6 hours. After the reaction is complete, concentrate under reduced pressure to obtain a yellow solid, recrystallize with methanol / water, and dry to obtain GL-V8 (88 g, yield 80%).

[0083] Example 7

[0084] Compared with Example 1, the difference lies in step d:

[0085] Take 110 g of Intermediate 3, mix it with 110 mL of tetrahydrofuran and 550 mL of water, slowly add dropwise 6.7 g of 98% sulfuric acid (diluted with 60 mL of water), heat up to 50 °C, and react for 3 hours. After the reaction is complete, concentrate under reduced pressure to obtain a yellow solid, recrystallize with methanol / water, and dry to obtain GL-V8 (76 g, yield 75%).

Claims

1. A preparation method of flavonoid compound GL-V8, characterized in that: The synthetic route is as follows: X and Y are halogen atoms; The preparation method includes the following steps: Step (a): Diethanolamine forms a diethanolamine salt in the presence of an acid. The diethanolamine salt, 2,2-dimethoxypropane and a catalyst react to form the compound shown in Formula 1. The reaction temperature is 25 - 100 °C, and the reaction time is 1 - 10 hours. The acid is hydrochloric acid, and the catalyst is p-toluenesulfonic acid; wherein, the molar ratio of diethanolamine, 2,2-dimethoxypropane and the catalyst is 1:2 - 5:0.01 - 0.05; Step (b): The compound shown in Formula 1 reacts with 1,4-halobutane in the presence of a base to form the compound shown in Formula 2. The reaction temperature is 30 - 40 °C, and the reaction time is 5 - 7 hours; wherein, the equivalent ratio of the compound shown in Formula 1 to 1,4-halobutane is 1:1 - 1:5; Step (c): The compound shown in Formula 2 reacts with wogonin in the presence of a base to form the compound shown in Formula 3. The reaction temperature is 80 - 90 °C, and the reaction time is 2 - 6 hours; wherein, the equivalent ratio of the compound shown in Formula 2 to wogonin is 1:1 - 10:1; Step (d): The compound shown in Formula 3 reacts in the presence of an acid to form the final product GL-V8. The reaction temperature is 30 - 100 °C, and the reaction time is 2 - 3 hours.

2. The preparation method according to claim 1, characterized in that: In the process of diethanolamine forming a diethanolamine salt in the presence of an acid, the reaction solvent is selected from one or more of dichloromethane, ethyl acetate, methyl tert-butyl ether, acetonitrile, acetone, methanol, ethanol, 1,4-dioxane, N,N-dimethylformamide, and water.

3. The preparation method according to claim 1, characterized in that: In step (b), the 1,4-halobutane is 1-bromo-4-chlorobutane. The reaction solvent is selected from one or more of tetrahydrofuran, 2-methyltetrahydrofuran, N,N-dimethylformamide, acetonitrile, acetone, dichloromethane, dimethyl sulfoxide, methanol, and water. The base is selected from potassium carbonate, sodium carbonate, sodium bicarbonate, potassium bicarbonate, sodium hydroxide, potassium hydroxide, or tetramethylammonium hydroxide.

4. The preparation method according to claim 1, wherein: In step (c), the base is selected from potassium carbonate, sodium carbonate, sodium bicarbonate, potassium bicarbonate, sodium hydroxide, potassium hydroxide, or tetramethylammonium hydroxide. The reaction solvent is selected from one or more of tetrahydrofuran, 2-methyltetrahydrofuran, N,N-dimethylformamide, acetonitrile, acetone, dichloromethane, dimethyl sulfoxide, methanol, and water.

5. The preparation method according to claim 1, characterized in that: In step (d), the acid is selected from formic acid, hydrochloric acid, acetic acid, trifluoroacetic acid, or sulfuric acid. The reaction solvent is selected from one or more of acetonitrile, methanol, ethanol, tetrahydrofuran, 2-methyltetrahydrofuran, and water.

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