Synthetic method of natural product flavonoid compound

Abstract

The invention discloses a synthetic method of a natural product flavonoid compound, and belongs to the technical field of synthesis of flavone derivatives, the synthetic method comprises eight specific steps, the synthetic method adopts an artificial synthesis method to synthesize the flavonoid compound 6-methoxyl-4 ', 5, 6'-tetramethyl-4 ', 5'-tetramethyl-4 ', 5'-tetramethyl-4 ', 5'-tetramethyl-4 ', 5'-tetramethyl-4 '-tetramethyl-4', 5 '-tetramethyl-4' Compared with the prior art, the method provided by the invention has the advantages of high yield and low cost, and can replace the method for separating and preparing the compound from the plant, and the method provided by the invention can be used for preparing the compound from the plant in the presence of the 2, 3, 7-trihydroxy-8-(1 ''-(3'' '-methoxyl-4 '''-hydroxyphenyl) ethyl) isoflavone.

Description

Technical Field

[0001] The present invention belongs to the technical field of the synthesis of flavone derivatives, and particularly relates to a method for synthesizing a natural product flavonoid compound. Background Art

[0002] The Chinese medicinal material Belamcanda chinensis is the dried rhizome of the plant Iris tectorum Maxim. in the family Iridaceae. Its functions and indications are clearing heat and detoxifying, resolving phlegm, and relieving sore throat. It is used for the treatment of heat-toxin phlegm-fire stasis, sore throat, excessive phlegm, cough, and asthma. Clinically, it has a good effect on the treatment of inflammatory diseases, cough, and liver-related diseases. The flavonoid compound 6-methoxy-4',5,7-trihydroxy-8-(1''-(3'''-methoxy-4'''-hydroxyphenyl)ethyl)isoflavone is a new compound extracted and separated from Belamcanda chinensis by methods such as silica gel column chromatography, Sephadex LH-20, ODS, and high-performance liquid chromatography. The structural formula of the compound is shown in the following figure. This flavonoid compound has good anti-inflammatory effects and safety, and has the characteristic of selectively inhibiting mPGES-1. It can be used for the treatment of inflammatory diseases, AD, OA, Parkinson's disease, colorectal cancer, liver-related diseases, and myocardial ischemia-reperfusion, and has the prospect of further research and drug development.

[0003]

[0004] The prior art CN115093388A discloses a preparation method of the flavonoid compound 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl)isoflavone. That is, after using 21 kg of crushed Chinese medicinal material Belamcanda chinensis as a raw material to obtain an extract by ethanol extraction, after obtaining the 70% ethanol elution part by macroporous resin column chromatography, using dichloromethane-methanol as the mobile phase to obtain fractions by silica gel column chromatography, and then successively passing through silica gel column chromatography, ODS column chromatography, Sephadex LH-20 column chromatography, and high-performance liquid chromatography to obtain 10 mg of this flavonoid compound.

[0005] This flavonoid compound is derived from plants and is prepared by using Belamcanda chinensis as a raw material through an extraction and separation method. However, this method has a low yield, high preparation cost, and is difficult to achieve large-scale production, which restricts the application of this type of compound.

[0006] Therefore, there is an urgent need to find a method for preparing the flavonoid compound 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl)isoflavone with a higher yield and suitable for scale-up production. Summary of the Invention

[0007] To solve the above problems, the present invention provides a novel chemical synthesis method for flavonoid compound 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl) isoflavone with high yield and suitable for large-scale production.

[0008] To achieve the above object, the technical solution adopted by the present invention is:

[0009] A synthesis method for a natural product flavonoid compound, and the synthesis method is carried out according to the following route:

[0010]

[0011] The synthesis method includes the following specific steps:

[0012] (1) Dissolve 2,4,6-trihydroxybenzaldehyde compound in dichloromethane, then add N,N-diisopropylethylamine compound, degas and purge with nitrogen 3-4 times, and charge with inert gas protection. Stir at -2 - 0 °C for 10 - 15 min, and add bromo(methoxy)methane. Stir at room temperature for 15 - 18 h to obtain a red solution. After monitoring the completion of the reaction, pour the red solution into 200 - 250 mL of water, extract with ethyl acetate 3 - 4 times, 200 - 250 mL each time. Combine the extracted organic phases, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel column chromatography using petroleum ether and ethyl acetate as eluents to obtain product 2;

[0013] (2) Dissolve product 2 in dichloromethane, add m-chloroperoxybenzoic acid, stir at room temperature for 5 - 20 h, then add 10 - 12% sodium hydroxide solution and methanol, stir at room temperature to obtain a reaction solution. After monitoring the completion of the reaction, acidify the reaction solution with 2 - 3 times the amount of hydrochloric acid solution, and then extract with ethyl acetate 3 - 4 times, 200 - 280 mL each time. Combine the extracted organic phases, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel column chromatography using petroleum ether and ethyl acetate as eluents to obtain product 3;

[0014] (3) Dissolve product 3 in N,N-dimethylformamide, add cesium carbonate, degas and purge with nitrogen 3 - 4 times, then fill with nitrogen for protection. Add methyl iodide, fill with inert gas for protection, and stir at room temperature for 15 - 18 h to form a yellow solution. After monitoring the completion of the reaction, pour the yellow solution into 180 - 200 mL of water, extract with ethyl acetate 3 - 4 times, 200 - 250 mL each time. Combine the extracted organic phases, wash with saturated brine, dry over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain the crude product. Purify the crude product by silica gel column chromatography using petroleum ether and ethyl acetate to obtain product 4;

[0015] (4) Dissolve product 4 in methanol, add 36 - 38% concentrated hydrochloric acid for mixing, degas and purge with nitrogen 3 - 4 times, then fill with inert gas for protection. Stir at room temperature for 3 - 4 h to form a yellow solution. After monitoring the completion of the reaction, concentrate the yellow solution under reduced pressure and dry to obtain the intermediate product. Dissolve the intermediate product in 150 - 200 mL of water, then extract with ethyl acetate 3 - 4 times, 150 - 200 mL each time. Combine the extracted organic phases, dry over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain the crude product. Purify the crude product by silica gel column chromatography using dichloromethane and methanol to obtain product 5;

[0016] (5) Mix product 5 with 4-hydroxyphenylacetonitrile and ether, add zinc chloride to obtain a reaction solution. Degas the reaction solution and purge with nitrogen 3 - 4 times. Slowly introduce hydrogen chloride gas into the reaction solution at a temperature below 0 °C for 1 - 5 h, and fill with inert gas for protection. Stir at room temperature for 15 - 18 h to form a yellow solution. After monitoring the completion of the reaction, filter the reaction solution. Wash the filter cake with 50 - 60 mL of anhydrous ether and dissolve it in 120 - 180 mL of water. Heat under reflux for 0.5 - 2 h, then cool the solution to 0 °C and filter. Vacuum dry the filter cake to obtain product 6;

[0017] (6) Dissolve product 6 in N,N-dimethylformamide, heat to 55 - 65 °C, and successively add boron trifluoride diethyl etherate and an anhydrous N,N-dimethylformamide solution of methanesulfonyl chloride. Continue to heat to 75 - 95 °C and maintain for 2 - 3 h. After monitoring the completion of the reaction, add 1.5 - 1.8 L of cold water to the reaction solution to form a yellow precipitate. Filter, wash the filter cake with water and dry to obtain a mixture of product 7 and isomeric product 7-1;

[0018] (7) Dissolve the mixture of product 7 and isomeric product 7-1 in cyclohexanol, add potassium carbonate, degas and then fill with inert gas for protection. Stir at 180 - 200 °C to form a black solution. After monitoring that the reaction is complete, 98% of product 7 and 2% of isomeric product 7-1 are produced. Cool the mixed solution, add an appropriate amount of water, and extract with ethyl acetate 3 - 4 times, 30 - 50 mL each time. Combine the extracted organic phases. Wash the combined organic phases with 10 - 20 mL of brine, then dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain the crude product. Purify the crude product by silica gel column chromatography using petroleum ether and ethyl acetate to obtain product 7;

[0019] (8) Dissolve product 7 and (2E)-3-(4-hydroxy-3-methoxyphenyl)-2-acrylic acid in acetonitrile, add a mixture of cerium(III) chloride heptahydrate and sodium iodide, heat the mixture to 75 - 95 °C and stir for 72 - 75 h to form a black solution. After monitoring that the reaction is complete, cool the black solution, add 100 - 200 mL of water, and extract with ethyl acetate 3 - 4 times, 30 - 50 mL each time. Combine the extracted organic phases, wash with 10 - 20 mL of brine, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain the crude product. Purify the crude product by silica gel chromatography using petroleum ether and ethyl acetate, and then purify by elution with water and acetonitrile through a C18 reverse-phase column to obtain a yellow solid, namely flavonoid 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl)isoflavone.

[0020] In a preferred embodiment, the molar ratio of the raw material 2,4,6-trihydroxybenzaldehyde compound, dichloromethane, N,N-diisopropylethylamine compound, and bromomethyl methyl ether in step (1) is: 1:45 - 50:5 - 8:4 - 6.

[0021] In a preferred embodiment, the volume ratio of petroleum ether to ethyl acetate in steps (1) - (3) is 1:0 - 4, the volume ratio of dichloromethane to methanol in step (4) is 1 - 10:0 - 1, and the volume ratio of petroleum ether to ethyl acetate in steps (7) - (8) is 1 - 10:1.

[0022] In a preferred embodiment, the molar ratio of the raw material product 2, dichloromethane, m-chloroperoxybenzoic acid, sodium hydroxide solution, and methanol in step (2) is 1:26 - 30:1 - 3:3 - 4:28 - 30.

[0023] In a preferred embodiment, the molar ratio of the raw material product 3, N,N-dimethylformamide, cesium carbonate, and iodomethane in step (3) is 1:33 - 35:8 - 10:1 - 2.

[0024] In a preferred embodiment, the molar ratio of raw material product 4, methanol, and concentrated hydrochloric acid in step (4) is 1:130 - 140:7 - 9.

[0025] In a preferred embodiment, the molar ratio of raw material product 5, 4-hydroxyphenylacetonitrile, ether, and zinc chloride in step (5) is 1:1 - 1.5:30 - 35:1 - 1.3.

[0026] In a preferred embodiment, the molar ratio of raw material product 6, N,N-dimethylformamide, boron trifluoride diethyl etherate, and an anhydrous N,N-dimethylformamide solution of methanesulfonyl chloride in step (6) is 1:28 - 31:5 - 8:10 - 15. The anhydrous N,N-dimethylformamide solution of methanesulfonyl chloride is obtained by dissolving methanesulfonyl chloride in anhydrous N,N-dimethylformamide, where the molar ratio of methanesulfonyl chloride to anhydrous N,N-dimethylformamide is 1:2 - 2.5.

[0027] In a preferred embodiment, the molar ratio of the mixture of product 7 and isomeric product 7-1, cyclohexanol, and potassium carbonate in step (7) is 1:40 - 45:1 - 3. The molar ratio of raw material product 7, (2E)-3-(4-hydroxy-3-methoxyphenyl)-2-acrylic acid, acetonitrile, cerium chloride heptahydrate, and sodium iodide in step (8) is 1:1.0 - 2.0:210 - 227:0.0008 - 0.002:0.3 - 0.6.

[0028] In a preferred embodiment, the monitoring method is to use LCMS to monitor whether the reaction is completed.

[0029] Compared with the prior art, the beneficial effects of the present invention are as follows: (1) The present invention provides a method for the total synthesis of natural product flavonoids. This method uses an artificial synthesis method, has a relatively high yield, can replace the method of separating and preparing this compound from plants, and greatly reduces the cost.

[0030] (2) The artificial synthesis method adopted by the present invention is easy to standardize and automate the production process, is suitable for large-scale industrial production. Compared with the traditional plant extraction method, the synthesis process is not affected by factors such as raw material supply and seasonal changes, and can stably provide a large number of high-quality natural product flavonoid products.

[0031] (3) In the synthesis process of the present invention, through multiple purification operations such as silica gel column chromatography and C18 reversed-phase column elution, impurities can be effectively removed to obtain high-purity flavonoids. At the same time, by using LCWS to monitor the reaction process, the reaction situation can be understood in real time, and the reaction conditions can be adjusted in a timely manner to ensure the stability and consistency of product quality.

[0032] (4) In the prior art, the synthesis of compound 7 from compound 6 produces isomer 7-1, and it is difficult to separate isomer 7-1 from compound 7. By heating under alkaline conditions, isomer 7-1 can be converted to compound 7. However, when the equivalent is increased to 10 times, isomer 7-1 cannot be completely converted to compound 7, and it is difficult to obtain compound 7 with high purity. After the condition optimization of the present invention, compound 7-1 is completely converted to compound 7, obtaining a higher yield and higher purity;

[0033] (5) The flavonoid compounds synthesized by the present invention have good anti-inflammatory effects and the characteristic of selectively inhibiting mPGES-1, and can be used for the treatment of various diseases such as inflammatory diseases, Alzheimer's disease, Parkinson's disease, intestinal cancer, liver-related diseases, and myocardial ischemia-reperfusion. The improvement of the synthesis method makes the supply of flavonoid compounds more sufficient, providing strong support for further research in their drug development and clinical applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 It is the mass spectrum of the flavonoid compound 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl)isoflavone synthesized in Example 1 of the present invention;

[0035] Figure 2 It is the hydrogen spectrum of the flavonoid compound 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl)isoflavone synthesized in Example 1 of the present invention;

[0036] Figure 3 It is the chromatogram of the flavonoid compound 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl)isoflavone synthesized in Example 1 of the present invention. DETAILED DESCRIPTION OF THE INVENTION

[0037] In order to enable those skilled in the art to better understand the technical solution, the present invention will be described in detail below in conjunction with embodiments. The description of this part is only exemplary and explanatory, and should not have any limiting effect on the protection scope of the present invention.

[0038] Example 1:

[0039] A synthesis method of a natural product flavonoid compound, comprising the following specific steps:

[0040] (1) Dissolve 40 g of 2,4,6-trihydroxybenzaldehyde in 800 mL of dichloromethane, then add 271.25 mL of N,N-diisopropylethylamine. After degassing the mixture, purge it with nitrogen three times and fill it with nitrogen for protection. Stir at 0 °C for 10 min, and add 105.92 mL of bromo(methoxy)methane. Warm up to room temperature and stir for 16 h to obtain a red solution. After monitoring the completion of the reaction by LCMS, pour the red solution into 200 mL of water and extract it three times with 200 mL of ethyl acetate each time. Combine the extracted organic phases, dry them with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and then dry it to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate mixture with a volume ratio of 1:1 as the eluent to obtain a yellow solid product 2 with a purity of 33.65%;

[0041] (2) Dissolve 61 g of product 2 in 400 mL of dichloromethane, add 64.89 g of m-chloroperbenzoic acid (85% purity), stir at room temperature for 16 h, then add 243.96 mL of 10% sodium hydroxide solution and 250 mL of methanol, and continue to stir at room temperature for 3 h to obtain a reaction solution. After monitoring the completion of the reaction by LCMS, acidify the reaction solution with 2-fold hydrochloric acid solution, then extract it three times with 280 mL of ethyl acetate each time. Combine the extracted organic phases, dry them with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and then dry it to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate mixture with a volume ratio of 1:1 as the eluent to obtain a yellow oily product 3 with a purity of 60.98%;

[0042] (3) Dissolve 35.64 g of product 3 in 350 mL of N,N-dimethylformamide, add 384.68 g of cesium carbonate, degas, purge it with nitrogen three times, and fill it with nitrogen for protection. Then add 12.65 mL of iodomethane, fill it with inert gas for protection, and stir at room temperature for 18 h to form a yellow solution. After monitoring the completion of the reaction by LCMS, pour the yellow solution into 180 mL of water and extract it three times with 200 mL of ethyl acetate each time. Combine the extracted organic phases, wash them with saturated brine, dry them with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and then dry it to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate mixture with a volume ratio of 1:1 as the eluent to obtain a yellow oily product 4 with a purity of 74.83%;

[0043] (4) Dissolve 26.44 g of product 4 in 530 mL of methanol, add 68.78 mL of 37% concentrated hydrochloric acid for mixing. After degassing the mixture, purge it with nitrogen three times and fill it with nitrogen for protection. Stir at room temperature for 3 h to form a yellow solution. After monitoring the completion of the reaction by LCMS, concentrate the reaction solution under reduced pressure and then dry it to obtain an intermediate product. Dissolve it in 150 mL of water and extract it three times with 150 mL of ethyl acetate each time. Combine the extracted organic phases, dry them with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and then dry it to obtain a crude product. Purify the crude product by silica gel column chromatography using a dichloromethane and methanol eluent with a volume ratio of 10:1 to obtain a yellow oily product 5 with a purity of 63.41%;

[0044] (5) Mix 8.5 g of product 5, 7.25 g of 4-hydroxyphenylacetonitrile and 204 mL of ether, then add 7.42 g of zinc chloride to obtain a reaction solution. After degassing the reaction solution, purge it with nitrogen three times. Slowly introduce hydrogen chloride gas into the reaction solution at a temperature below 0 °C for 3 h and fill it with nitrogen for protection. Stir at room temperature for 18 h to form a yellow solution. After monitoring the completion of the reaction by LCMS, filter the reaction solution. Wash the filter cake with 50 mL of anhydrous ether and dissolve it in 120 mL of water. After heating under reflux for 1 h, cool the solution to 0 °C and filter. Vacuum dry the filter cake to obtain a yellow solid product 6 with a purity of 57.53%;

[0045] (6) Dissolve 9.0 g of product 6 in 67 mL of N,N-dimethylformamide, heat it to sixty °C, and successively add 19.5 mL of boron trifluoride etherate and 38 mL of an anhydrous N,N-dimethylformamide solution containing 11 mL of methanesulfonyl chloride. Continue to heat to ninety °C and maintain for 2.5 h. After monitoring the completion of the reaction by LCMS, add 1.5 L of cold water to the reaction solution to form a yellow precipitate. Filter and collect the filter cake, wash it with water and dry it to obtain a mixture of product 7 and isomeric product 7-1 with a purity of 91.94%;

[0046] (7) Dissolve 13 g of the mixture of product 7 and isomeric product 7-1 in 200 mL of cyclohexanol, add 8.97 g of potassium carbonate, degas it and purge it with nitrogen three times, and fill it with nitrogen for protection. Stir at 180 °C for 16 h to form a black solution. After monitoring the completion of the reaction by LCMS, 98% of the target product and 2% of isomeric impurities are formed. After cooling the mixed solution, add 20 mL of water, and then extract it three times with 50 mL of ethyl acetate each time. Combine the extracted organic phases, wash the combined organic phases with 10 mL of brine, then dry it with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and then dry it to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate eluent with a volume ratio of 10:1 to obtain a yellow solid product 7 with a purity of 84.62%;

[0047] (8) Dissolve 5.1 g of product 7 and 6.6 g of (2E)-3-(4-hydroxy-3-methoxyphenyl)-2-acrylic acid in 200 mL of acetonitrile, add 3.16 mg of cerium(III) chloride heptahydrate and 1.27 g of sodium iodide and mix. Heat the mixture to 90 °C and stir for 72 h to form a black solution. After monitoring the completion of the reaction by LCMS, cool the above mixed solution, add 100 mL of water, extract with ethyl acetate 3 times, 30 mL each time. Combine the extracted organic phases, wash the combined organic phases with 10 mL of brine, dry over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel chromatography using a petroleum ether and ethyl acetate eluent with a volume ratio of 10:1, and then purify by elution through a C18 reverse-phase column using water (containing 0.1% NH4HCO3) and acetonitrile (95%) as the eluent to obtain a yellow solid, namely the flavonoid compound 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl)isoflavone, with a purity of 97%.

[0048] Example 2:

[0049] A method for synthesizing a natural product flavonoid compound, comprising the following specific steps:

[0050] (1) Dissolve 50 g of 2,4,6-trihydroxybenzaldehyde in 998 mL of dichloromethane, then add 383 mL of N,N-diisopropylethylamine. Degas the mixture and purge with nitrogen 4 times, and fill with nitrogen for protection. Stir at -1 °C for 13 min and add 148 mL of bromo(methoxy)methane. Warm to room temperature and stir for 17 h to obtain a red solution. After monitoring the completion of the reaction by LCMS, pour the red solution into 250 mL of water, extract with ethyl acetate 4 times, 230 mL each time. Combine the extracted organic phases, dry over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate eluent with a volume ratio of 1:2 to obtain a yellow solid product 2, with a purity of 31.72%;

[0051] (2) Dissolve 65 g of Product 2 in 404 mL of dichloromethane. Add 83.18 g of m-chloroperbenzoic acid (85% purity). Stir at room temperature for 8 h. Then add 278 mL of 12% sodium hydroxide solution and 270 mL of methanol. Continue to stir at room temperature for 5 h to obtain a reaction solution. After monitoring the completion of the reaction by LCMS, acidify the reaction solution with 3-fold hydrochloric acid solution, and then extract with ethyl acetate 4 times, 230 mL each time. Combine the extracted organic phases, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate eluent with a volume ratio of 1:2 to obtain yellow oily Product 3 with a purity of 62.15%;

[0052] (3) Dissolve 37.36 g of Product 3 in 355 mL of N,N-dimethylformamide. Add 425.97 g of cesium carbonate. After degassing, purge with nitrogen 4 times and charge with nitrogen for protection. Then add 11 mL of methyl iodide and charge with inert gas for protection. Stir at room temperature for 15 h to form a yellow solution. After monitoring the completion of the reaction by LCMS, pour the yellow solution into 200 mL of water and extract with ethyl acetate 4 times, 250 mL each time. Combine the extracted organic phases, wash with saturated brine, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate eluent with a volume ratio of 1:2 to obtain yellow oily Product 4 with a purity of 75.81%;

[0053] (4) Dissolve 33.94 g of Product 4 in 637 mL of methanol. Add 74 mL of 36% concentrated hydrochloric acid and mix. After degassing, purge with nitrogen 4 times and charge with nitrogen for protection. Stir at room temperature for 4 h to form a yellow solution. After monitoring the completion of the reaction by LCMS, concentrate the reaction solution under reduced pressure and dry to obtain an intermediate product. Dissolve it in 180 mL of water and extract with ethyl acetate 4 times, 200 mL each time. Combine the extracted organic phases, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel column chromatography using a dichloromethane and methanol eluent with a volume ratio of 5:1 to obtain yellow oily Product 5 with a purity of 62.52%;

[0054] (5) After mixing 12.15 g of product 5, 12.73 g of 4-hydroxyphenylacetonitrile and 277 mL of diethyl ether, 11.99 g of zinc chloride was added to obtain a reaction solution. The reaction solution was degassed and purged with nitrogen 4 times. Hydrogen chloride gas was slowly introduced into the reaction solution at a temperature below 0 °C for 1 h, and nitrogen was filled for protection. The mixture was stirred at room temperature for 15 h to form a yellow solution. After monitoring the completion of the reaction by LCMS, the reaction solution was filtered. The filter cake was rinsed with 50 mL of anhydrous diethyl ether and then dissolved in 180 mL of water. After heating under reflux for 2 h, the solution was cooled to 0 °C and filtered. The filter cake was dried in vacuo to obtain a yellow solid product 6 with a purity of 58.33%;

[0055] (6) 10.31 g of product 6 was dissolved in 81 mL of N,N-dimethylformamide and heated to 65 °C. 18 mL of boron trifluoride diethyl etherate and 39 mL of an anhydrous N,N-dimethylformamide solution containing 18 mL of methanesulfonyl chloride were added successively. The mixture was further heated to 80 °C and maintained for 3 h. After monitoring the completion of the reaction by LCMS, 1.6 L of cold water was added to the reaction solution to form a yellow precipitate. After filtration, the filter cake was collected, washed with water and dried to obtain a mixture of product 7 and isomeric product 7-1 with a purity of 92.36%;

[0056] (7) 15 g of the mixture of product 7 and isomeric product 7-1 was dissolved in 227 mL of cyclohexanol, and then 12.76 g of potassium carbonate was added. After degassing and purging with nitrogen 4 times, nitrogen was filled for protection. The mixture was stirred at 190 °C for 17 h to form a black solution. After monitoring the completion of the reaction by LCMS, 98% of the target product and 2% of isomeric impurities were formed. After the mixed solution was cooled, 25 mL of water was added, and then extracted with ethyl acetate 4 times, 60 mL each time. The extracted organic phases were combined, and the combined organic phase was washed with 15 mL of brine, then dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure and dried to obtain a crude product. The crude product was purified by silica gel column chromatography using a mixture of petroleum ether and ethyl acetate with a volume ratio of 8:1 as the eluent to obtain a yellow solid product 7 with a purity of 87.02%;

[0057] (8)Dissolve 5.41 g of product 7 and 3.82 g of (2E)-3-(4-hydroxy-3-methoxyphenyl)-2-acrylic acid in 200 mL of acetonitrile. Add 6.02 mg of cerium(III) chloride heptahydrate and 1.08 g of sodium iodide and mix. Heat the mixture to 95 °C and stir for 74 h to form a black solution. After monitoring the completion of the reaction by LCMS, cool the above mixed solution and add 200 mL of water. Extract with ethyl acetate 4 times, 45 mL each time. Combine the extracted organic phases, wash the combined organic phases with 15*-mL of brine, dry over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel chromatography using a petroleum ether and ethyl acetate eluent with a volume ratio of 8:1, and then purify by elution through a C18 reverse-phase column using water (containing 0.1% NH4HCO3) and acetonitrile (90%) as the eluent to obtain a yellow solid, namely the flavonoid compound 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl)isoflavone, with a purity of 98.3%.

[0058] Example 3:

[0059] A method for synthesizing a natural product flavonoid compound, comprising the following specific steps:

[0060] (1) Dissolve 95 g of 2,4,6-trihydroxybenzaldehyde in 851 mL of dichloromethane, then add 258 mL of N,N-diisopropylethylamine. Degas the mixture and purge with nitrogen 3 times, and charge with nitrogen for protection. Stir at -2 °C for 15 min and add 111 mL of bromo(methoxy)methane. Warm to room temperature and stir for 18 h to obtain a red solution. After monitoring the completion of the reaction by LCMS, pour the red solution into 240 mL of water. Extract with ethyl acetate 3 times, 250 mL each time. Combine the extracted organic phases, dry over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate eluent with a volume ratio of 1:4 to obtain a yellow solid product 2, with a purity of 31.72%;

[0061] (2) Dissolve 83 g of Product 2 in 541 mL of dichloromethane, add 96.14 g of m-chloroperoxybenzoic acid (85% purity), stir at room temperature for 20 h, then add 337.5 mL of 11% sodium hydroxide solution and 337 mL of methanol, and continue to stir at room temperature for 2.5 h to obtain a reaction solution. After monitoring the completion of the reaction by LCMS, acidify the reaction solution with 2-fold hydrochloric acid solution, then extract it with ethyl acetate three times, 200 mL each time. Combine the extracted organic phases, dry them over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry it to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate eluent with a volume ratio of 1:4 to obtain yellow oily Product 3 with a purity of 62.15%;

[0062] (3) Dissolve 68.4 g of Product 3 in 656 mL of N,N-dimethylformamide, add 768.8 g of cesium carbonate, degas and purge with nitrogen three times, and then charge with nitrogen protection. Then add 26 mL of methyl iodide, charge with inert gas protection, and stir at room temperature for 15 h to form a yellow solution. After monitoring the completion of the reaction by LCMS, pour the yellow solution into 190 mL of water, extract it with ethyl acetate three times, 230 mL each time. Combine the extracted organic phases, wash them with saturated brine, dry them over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry it to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate eluent with a volume ratio of 1:4 to obtain yellow oily Product 4 with a purity of 75.81%;

[0063] (4) Dissolve 46.92 g of Product 4 in 920 mL of methanol, add 95 mL of 38% concentrated hydrochloric acid for mixing, degas the mixture and purge with nitrogen three times, and then charge with nitrogen protection. Stir at room temperature for 3.6 h to form a yellow solution. After monitoring the completion of the reaction by LCMS, concentrate the reaction solution under reduced pressure and dry it to obtain an intermediate product. Dissolve it in 200 mL of water, extract it with ethyl acetate three times, 180 mL each time. Combine the extracted organic phases, dry them over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry it to obtain a crude product. Purify the crude product by silica gel column chromatography using a dichloromethane and methanol eluent with a volume ratio of 6:1 to obtain yellow oily Product 5 with a purity of 62.52%;

[0064] (5) Mix 32.68 g of product 5, 41.73 g of 4-hydroxyphenylacetonitrile and 754 mL of diethyl ether, then add 37.01 g of zinc chloride to obtain a reaction solution. Degas the reaction solution and purge it with nitrogen three times. Slowly introduce hydrogen chloride gas into the reaction solution at a temperature below 0 °C for 5 h, and charge nitrogen for protection. Stir at room temperature for 18 h to form a yellow solution. After monitoring the completion of the reaction by LCMS, filter the reaction solution. Wash the filter cake with 50 mL of anhydrous diethyl ether and dissolve it in 160 mL of water. After heating under reflux for 0.5 h, cool the solution to 0 °C and filter. Vacuum dry the filter cake to obtain yellow solid product 6 with a purity of 58.33%;

[0065] (6) Dissolve 23.6 g of product 6 in 194 mL of N,N-dimethylformamide, heat to 55 °C, and successively add 78 mL of boron trifluoride diethyl etherate and 71 mL of an anhydrous N,N-dimethylformamide solution containing 28 mL of methanesulfonyl chloride. Continue to heat to 95 °C and maintain for 2.5 h. After monitoring the completion of the reaction by LCMS, add 1.8 L of cold water to the reaction solution to form a yellow precipitate. After filtration, collect the filter cake, wash it with water and dry it to obtain a mixture of product 7 and isomeric product 7-1 with a purity of 91.38%;

[0066] (7) Dissolve 36.9 g of the mixture of product 7 and isomeric product 7-1 in 520 mL of cyclohexanol, then add 25.84 g of potassium carbonate. Degas and purge with nitrogen three times, and charge nitrogen for protection. Stir at 200 °C for 18 h to form a black solution. After monitoring the completion of the reaction by LCMS, 98% of the target product and 2% of isomeric impurities are formed. After cooling the mixed solution, add 30 mL of water, and extract with ethyl acetate three times, 80 mL each time. Combine the extracted organic phases, wash the combined organic phases with 13 mL of brine, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel column chromatography using a petroleum ether and ethyl acetate eluent with a volume ratio of 6:1 to obtain yellow solid product 7 with a purity of 79.02%;

[0067] (8) Dissolve 10.17 g of product 7 and 7.53 g of (2E)-3-(4-hydroxy-3-methoxyphenyl)-2-acrylic acid in 400 mL of acetonitrile, add 10.05 mg of cerium(III) chloride heptahydrate and 1.95 g of sodium iodide and mix. Heat the mixture to 75 °C and stir for 74 h to form a black solution. After monitoring the completion of the reaction by LCMS, cool the above mixed solution, add 150 mL of water, extract with ethyl acetate three times, 50 mL each time. Combine the extracted organic phases, wash the combined organic phases with 20 mL of brine, dry over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel chromatography using petroleum ether and ethyl acetate in a volume ratio of 9:1 as the eluent, and then purify by elution through a C18 reverse-phase column using water (containing 0.1% NH4HCO3) and acetonitrile (80%) as the eluent to obtain a yellow solid, namely the flavonoid 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl)isoflavone, with a purity of 96.21%.

[0068] Perform mass spectrometry analysis on the flavonoid obtained in Example 1 using electrospray ionization mass spectrometry (ESI), as Figure 1 shown. The known mass spectrometry data of this flavonoid are: Mass Spectrum (ESI) m / z = 451.1 (M+H) + , indicating that a protonated ion is formed during the ionization process of this flavonoid, and the molecular weight of the compound is 451.1. To further determine the chemical environment of hydrogen atoms and the structural information in the compound, use nuclear magnetic resonance (NMR) technology to measure the compound and obtain a 1H NMR spectrum, as Figure 2 shown. The 1H NMR data are: 1H-NMR (400 MHz, DMSO-d6) = δ 13.24 (s, 1H), 10.21 (s, 1H), 9.57 (s, 1H), 8.66 (s, 1H), 8.39 (s, 1H), 7.49 - 7.28 (m, 2H), 6.92 - 6.85 (m, 1H), 6.84 - 6.78 (m, 2H), 6.68 - 6.61 (m, 2H), 4.70 (q, J = 7.2 Hz, 1H), 3.78 (s, 3H), 3.71 (s, 3H), 1.68 (d, J = 7.2 Hz, 3H).

[0069] To further determine the drug quality of the flavonoid obtained in Example 1, use a chromatograph to measure the compound and obtain a chromatogram, as Figure 3 shown. The peak area of the main component at 5.594 min accounts for 97.19% of the total peak area, indicating that the purity of this flavonoid is relatively high, and the areas of other peaks are small, indicating that the impurity content is low. The specific chromatographic data are shown in the following table.

[0070] Table 1

[0071]

[0072] To verify the optimization effect of the conditions for separating and purifying the mixture of product 7 and isomeric product 7-1 in step (7) of the present invention, a comparative experiment was conducted. The experimental procedure was as follows: A mixture of starting compounds 7 and 7-1 (0.33 mmol, 1 eq), in which the content of product 7 was 60%, was added to a 10 mL glass reaction flask equipped with magnetic stirring. Then, different bases / acids (0.50 mmol, 1.5 eq) and different solvents (1 mL) were added. After three nitrogen replacements, the reaction was heated in an oil bath or by microwave for 18 h. The reaction progress and the isomer ratio in the reaction solution were monitored by LCMS, and the results are shown in Table 2.

[0073] Table 2

[0074]

[0075]

[0076] To further verify the optimization effect of the reaction conditions between product 7 and compound (2E)-3-(4-hydroxy-3-methoxyphenyl)-2-acrylic acid in step (8) of the present invention, a comparative experiment was conducted. The experimental procedure was as follows: Starting compound 7 (0.17 mmol, 1 eq) and compound (2E)-3-(4-hydroxy-3-methoxyphenyl)-2-acrylic acid (0.34 mmol, 2 eq) were added to a 10 mL glass reaction flask equipped with magnetic stirring. Then, a catalyst and an additive (when present) were added, and a solvent (1 mL) was added. After three nitrogen replacements, the reaction was heated in an oil bath. The reaction progress was monitored by LCMS, and the experimental results are shown in Table 3.

[0077] Table 3

[0078]

[0079]

[0080] It should be noted that in this article, the terms: including, containing and any other variants are intended to cover non-exclusive inclusion, so that a process, method, article or device including a series of elements not only includes those elements, but also includes other elements not explicitly listed, or also includes elements inherent in such a process, method, article or device. Specific examples are used in this article to elaborate on the principles and implementation manners of the technical solution of the present invention. The description of the above examples is only used to help understand the method and its core idea of the present invention. The above is only the preferred implementation manner of the present invention. It should be pointed out that due to the limited nature of written expression and objectively infinite specific structures, for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements, refinements or changes can be made, or the above technical features can be combined in an appropriate manner; these improvements, refinements, changes or combinations, or directly applying the concept and technical solution of the present invention to other occasions without improvement, shall all be regarded as the protection scope of the present invention.

Claims

1. A synthetic method of a natural product flavonoid compound, characterized in that, The synthesis method proceeds according to the following route: The synthesis method includes the following specific steps: (1) Dissolve the 2,4,6-trihydroxybenzaldehyde compound in dichloromethane, then add the N,N-diisopropylethylamine compound. After degassing, purge with nitrogen 3 - 4 times, and charge with inert gas for protection. Stir at a temperature of -2 - 0 °C for 10 - 15 min, and add bromo(methoxy)methane. Stir at room temperature for 15 - 18 h to obtain a red solution. After monitoring the completion of the reaction, pour the red solution into 200 - 250 mL of water, extract with ethyl acetate 3 - 4 times, 200 - 250 mL each time. Combine the extracted organic phases, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and then dry to obtain a crude product. Purify the crude product by silica gel column chromatography using petroleum ether and ethyl acetate as eluents to obtain Product 2; (2) Dissolve Product 2 in dichloromethane, add m-chloroperoxybenzoic acid, stir at room temperature for 5 - 20 h, then add 10 - 12% sodium hydroxide solution and methanol, and stir at room temperature to obtain a reaction solution. After monitoring the completion of the reaction, acidify the reaction solution with 2 - 3 times the amount of hydrochloric acid solution, then extract with ethyl acetate 3 - 4 times, 200 - 280 mL each time. Combine the extracted organic phases, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and then dry to obtain a crude product. Purify the crude product by silica gel column chromatography using petroleum ether and ethyl acetate as eluents to obtain Product 3; (3) Dissolve Product 3 in N,N-dimethylformamide, add cesium carbonate, degas, purge with nitrogen 3 - 4 times, and charge with nitrogen for protection. Then add methyl iodide, charge with inert gas for protection, and stir at room temperature for 15 - 18 h to form a yellow solution. After monitoring the completion of the reaction, pour the yellow solution into 180 - 200 mL of water, extract with ethyl acetate 3 - 4 times, 200 - 250 mL each time. Combine the extracted organic phases, wash with saturated brine, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and then dry to obtain a crude product. Purify the crude product by silica gel column chromatography using petroleum ether and ethyl acetate to obtain Product 4; (4) Dissolve Product 4 in methanol, add 36 - 38% concentrated hydrochloric acid for mixing, degas, purge with nitrogen 3 - 4 times, and charge with inert gas for protection. Stir at room temperature for 3 - 4 h to form a yellow solution. After monitoring the completion of the reaction, concentrate the yellow solution under reduced pressure and then dry to obtain an intermediate product. Dissolve the intermediate product in 150 - 200 mL of water, then extract with ethyl acetate 3 - 4 times, 150 - 200 mL each time. Combine the extracted organic phases, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and then dry to obtain a crude product. Purify the crude product by silica gel column chromatography using dichloromethane and methanol to obtain Product 5; (5) Mix product 5 with 4-hydroxyphenylacetonitrile and diethyl ether, add zinc chloride to obtain a reaction solution. Degas the reaction solution and purge it with nitrogen 3 - 4 times. Slowly introduce hydrogen chloride gas into the reaction solution at a temperature below 0 °C for 1 - 5 h, and charge an inert gas for protection. Stir at room temperature for 15 - 18 h to form a yellow solution. After monitoring the completion of the reaction, filter the reaction solution. Wash the filter cake with 50 - 60 mL of anhydrous diethyl ether and then dissolve it in 120 - 180 mL of water. Heat under reflux for 0.5 - 2 h, then cool the solution to 0 °C, filter, and vacuum dry the filter cake to obtain product 6; (6) Dissolve product 6 in N,N-dimethylformamide, heat to 55 - 65 °C, and successively add anhydrous N,N-dimethylformamide solutions of boron trifluoride diethyl etherate and methanesulfonyl chloride. Continue to heat to 75 - 95 °C and maintain for 2 - 3 h. After monitoring the completion of the reaction, add 1.5 - 1.8 L of cold water to the reaction solution to form a yellow precipitate. Filter and wash and dry the filter cake with water to obtain a mixture of product 7 and isomeric product 7-1; (7) Dissolve the mixture of product 7 and isomeric product 7-1 in cyclohexanol, add potassium carbonate, degas and charge an inert gas for protection, and stir at 180 - 200 °C to form a black solution. After monitoring the completion of the reaction, 98% of product 7 and 2% of isomeric product 7-1 are produced. Cool the mixed solution and add an appropriate amount of water, then extract with ethyl acetate 3 - 4 times, 30 - 50 mL each time. Combine the extracted organic phases, wash the combined organic phases with 10 - 20 mL of brine, then dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel column chromatography using petroleum ether and ethyl acetate to obtain product 7; (8) Dissolve product 7 and (2E)-3-(4-hydroxy-3-methoxyphenyl)-2-acrylic acid in acetonitrile, add a mixture of cerium(III) chloride heptahydrate and sodium iodide, heat the mixture to 75 - 95 °C and stir for 72 - 75 h to form a black solution. After monitoring the completion of the reaction, cool the black solution and add 100 - 200 mL of water, then extract with ethyl acetate 3 - 4 times, 30 - 50 mL each time. Combine the extracted organic phases, wash with 10 - 20 mL of brine, dry with anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure and dry to obtain a crude product. Purify the crude product by silica gel chromatography using petroleum ether and ethyl acetate, and then purify it by elution with water and acetonitrile through a C18 reversed-phase column to obtain a yellow solid, namely flavonoid 6-methoxy-4′,5,7-trihydroxy-8-(1″-(3″′-methoxy-4″′-hydroxyphenyl)ethyl)isoflavone.

2. The synthetic method of a natural product flavonoid compound according to claim 1, wherein In the step (1), the molar ratio of the raw materials 2,4,6-trihydroxybenzaldehyde compound, dichloromethane, N,N-diisopropylethylamine compound, and bromomethyl methyl ether is: 1:45 - 50:5 - 8:4 - 6.

3. The synthesis method of a natural product flavonoid compound according to claim 1, characterized in that, The volume ratio of petroleum ether to ethyl acetate in the steps (1)-(3) is 1:0-4, the volume ratio of dichloromethane to methanol in the step (4) is 1-10:0-1, and the volume ratio of petroleum ether to ethyl acetate in the steps (7)-(8) is 1-10:

1.

4. A method for synthesizing a natural product flavonoid compound according to claim 1, characterized in that, In the step (2), the molar ratio of raw material product 2, dichloromethane, m-chloroperbenzoic acid, sodium hydroxide solution, and methanol is 1:26-30:1-3:3-4:28-30.

5. A method for synthesizing a natural product flavonoid compound according to claim 1, characterized in that, In the step (3), the molar ratio of raw material product 3, N,N-dimethylformamide, cesium carbonate, and methyl iodide is 1:33-35:8-10:1-2.

6. The synthesis method of a natural product flavonoid compound according to claim 1, characterized in that, In the step (4), the molar ratio of raw material product 4, methanol, and concentrated hydrochloric acid is 1:130-140:7-9.

7. A method for synthesizing a natural product flavonoid compound according to claim 1, characterized in that, In the step (5), the molar ratio of raw material product 5, 4-hydroxyphenylacetonitrile, diethyl ether, and zinc chloride is 1:1-1.5:30-35:1-1.

3.

8. A method for synthesizing a natural product flavonoid compound according to claim 1, characterized in that, In the step (6), the molar ratio of raw material product 6, N,N-dimethylformamide, boron trifluoride diethyl etherate, and an anhydrous N,N-dimethylformamide solution of methanesulfonyl chloride is 1:28-31:5-8:10-15. The anhydrous N,N-dimethylformamide solution of methanesulfonyl chloride is obtained by dissolving methanesulfonyl chloride in anhydrous N,N-dimethylformamide, and the molar ratio of methanesulfonyl chloride to anhydrous N,N-dimethylformamide is 1:2-2.

5.

9. A method for synthesizing a natural product flavonoid compound according to claim 1, characterized in that, In the step (7), the molar ratio of the mixture of product 7 and isomeric product 7-1, cyclohexanol, and potassium carbonate is 1:40-45:1-3. In the step (8), the molar ratio of raw material product 7, (2E)-3-(4-hydroxy-3-methoxyphenyl)-2-acrylic acid, acetonitrile, cerium chloride heptahydrate, and sodium iodide is 1:1.0-2.0:210-227:0.0008-0.002:0.3-0.

6.

10. A method for synthesizing a natural product flavonoid compound according to claim 1, characterized in that, The monitoring method is to monitor whether the reaction is completed by LCMS.

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