Cycloheptenone adduct of flavane and stilbene compound

Abstract

The invention relates to a cycloheptenone adduct of flavane and a stilbene compound as shown in a formula (I), and a preparation method and an anti-cerebral ischemia application of the cycloheptenone adduct. The compound provided by the invention is simple in preparation method and shows better neuroprotective activity. # imgabs0 #

Description

Technical Field

[0001] The invention belongs to the field of medicine, and specifically relates to a cycloheptenone adduct of a flavanoid and a stilbene compound, a synthesis method thereof and a use thereof in treating ischemic cerebrovascular diseases. Background Art

[0002] Ischemic cerebrovascular disease (ICVD) is a general term for ischemic cerebrovascular diseases of varying degrees, also known as cerebral ischemic disease. The mortality rate of the disease during the acute attack period is high, and the disability rate is extremely high. [1] At present, there is no clinically effective treatment, and drug treatment has poor long-term effects and is almost unable to reduce neuronal damage. [2] Therefore, the development of highly effective and low-toxic drugs to resist cerebral ischemia injury has always been a hot topic in the international medical community.

[0003] Plant polyphenols are a general term for di(tri)hydroxyphenyl structural fragments and their polymers in plants. Studies have shown that this type of natural product has good antioxidant capacity, and its neuroprotective effect has received extensive attention and research. [3] . Flavanes (such as catechins and epicatechins) and diphenylethylenes (such as resveratrol and piceatannol) are two representative types of plant polyphenols. Due to the easy polymerization of plant polyphenols, the two can also react to form adducts. This type of adduct has various connection methods and can form a complex structural skeleton. Pharmacological research results show that it has neuroprotective, antioxidant, and anti-tumor activities. This type of compound is present in small amounts in plants, resulting in high extraction and separation costs. Therefore, how to construct a chemical synthesis method for this type of component, especially a biomimetic synthesis method using flavanones and diphenylethylene as substrates, is of great significance for the development of new drugs. Summary of the invention

[0004] The object of the present invention is to provide a 5 / 6 / 7 tricyclic dimer compound in which flavanoids and stilbene compounds are connected via a 2-cyclohepten-1-one skeleton.

[0005] Another object of the present invention is to provide a method for preparing the compound.

[0006] Another object of the present invention is to provide a pharmaceutical composition comprising an effective dose of an adduct formed by connecting a class of flavanoids and a stilbene compound via a 2-cycloheptene-1-one skeleton and a pharmaceutically acceptable carrier and / or excipient.

[0007] Another object of the present invention is to provide a use of an adduct formed by connecting a type of flavanoid with a stilbene compound via a 2-cyclohepten-1-one skeleton in the preparation of a drug for treating and / or preventing ischemic cerebrovascular disease.

[0008] The present invention provides the following technical solutions:

[0009] The first aspect of the present invention provides a 5 / 6 / 7 tricyclic adduct of a flavanoid and a stilbene compound connected via a 2-cycloheptene-1-one skeleton, characterized in that it has a structure shown in general formula (I):

[0010]

[0011] in,

[0012] R A Selected from: hydrogen, hydroxyl,

[0013] R B Selected from hydrogen, hydroxy, alkoxy, halogen, cyano;

[0014] a is selected from integers of 0-5; b is selected from integers of 0-4; c is selected from integers of 0-5; d is selected from integers of 0-3.

[0015] Optionally, the halogen is selected from F, Cl, Br, I;

[0016] Optionally, the alkyl group in the “alkyl group”, “haloalkyl group” or “alkylsulfonyl group” is C 1 -C 10 Straight or branched chain alkyl, preferably C 1 -C 7 Straight or branched chain alkyl, preferably C 1 -C 5 The straight chain or branched alkyl group is preferably selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl, isopentyl, 1-ethylpropyl, neopentyl, n-hexyl, 1-methylpentyl, 2-methylpentyl, 3-methylpentyl, isohexyl, 1,1-dimethylbutyl, 2,2-dimethylbutyl, 3,3-dimethylbutyl, 1,2-dimethylbutyl, 1,3-dimethylbutyl, 2,3-dimethylbutyl, 2-ethylbutyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 2,2-dimethylpentyl, 3,3-dimethylpentyl, 2,3-dimethylpentyl, 2,4-dimethylpentyl, 3-ethylpentyl or 2,2,3-trimethylbutyl.

[0017] Optionally, a is selected from an integer of 0-3; Optionally, a is 0, 1, 2 or 3;

[0018] Optionally, b is selected from an integer of 2-3; Optionally, b is 2 or 3;

[0019] Optionally, c is selected from an integer of 0-3; Optionally, c is 0, 1, 2 or 3;

[0020] Optionally, d is 0, 1, 2 or 3.

[0021] Optionally, the compound of the structure represented by formula (I) or a pharmaceutically acceptable salt thereof is selected from the following compounds:

[0022]

[0023] The second aspect of the present invention provides a method for synthesizing a 5 / 6 / 7 tri-cycloadduct of a flavanoid and a stilbene compound connected via a 2-cyclohepten-1-one skeleton, the method comprising the following steps:

[0024] (1) Hydrobromic acid catalyzes the depolymerization to form a flavanone monomer derivative with a thio group connected at the 4 position,

[0025]

[0026] (2) Silver trifluoroacetate catalyzed substitution reaction to synthesize an intermediate in which the 4-position of flavanone is connected to the 2-position of stilbene.

[0027]

[0028] (3) Synthesis of a class of 5 / 6 / 7 tricyclic cycloadducts of flavanoids and stilbenes connected via a 2-cyclohepten-1-one skeleton catalyzed by trivalent iron or divalent copper.

[0029]

[0030] In various types, R A , R B , a, b, c, and d are defined as above.

[0031] The further preparation method thereof comprises the following steps:

[0032] The dried rhizome of Rhodiola rosea is used as the original drug, the medicinal material is crushed, and 80% ethanol is heated and refluxed for extraction three times. The extracts are combined and filtered, and the extract is concentrated under reduced pressure to obtain an extract. After water precipitation treatment, the precipitate is filtered, and the filtrate is concentrated under reduced pressure and separated by HP-20 macroporous adsorption resin column chromatography, and eluted with three gradients of pure water, 50% ethanol, and 95% ethanol. The eluate of the 50% ethanol part is concentrated under reduced pressure, and the Rhodiola rosea flavanone polymer part is obtained after freeze-drying (see Chinese patent, CN201010587820.7 for details). The part is dissolved in methanol, and the thiodepolymerization reagent benzyl mercaptan and 48% hydrobromic acid aqueous solution are added, and the reaction is carried out at 60°C for 6h to obtain a benzylthio-substituted flavanone monomer. The benzylthio-substituted flavanone monomer is dissolved in ultra-dry N,N-dimethylformamide, stirred at -10°C, and an excess of different substituted diphenylethylene and silver trifluoroacetate are added successively, and the reaction is stirred for 2h to obtain a flavanone diphenylethylene coupling intermediate. The flavanone stilbene coupling intermediate was dissolved in water, copper sulfate pentahydrate was added and stirred at room temperature for 2 hours to obtain the target product.

[0033] The third aspect of the present invention provides a pharmaceutical composition comprising a pharmaceutically effective dose of a compound having a structure represented by formula (I) or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

[0034] The pharmaceutical composition usually contains 0.1-95% by weight of the compound of formula (I) or its pharmaceutically acceptable salt. The compound of the present invention generally contains 0.1-100 mg in a unit dosage form, and preferably contains 4-150 mg in a unit dosage form.

[0035] The pharmaceutical composition of the compounds of this invention can be prepared according to methods well known in the art. When used for this purpose, if necessary, the compounds of this invention can be combined with one or more solid or liquid pharmaceutical excipients and / or adjuvants to prepare suitable application forms or dosage forms that can be used as human medicine or veterinary medicine.

[0036] The compound of the present invention or the pharmaceutical composition containing the same can be administered in a unit dosage form, and the administration route can be enteral or parenteral, such as oral, intramuscular, subcutaneous, nasal, oral mucosa, skin, peritoneum or rectum.

[0037] The administration route of the compound of the present invention or the pharmaceutical composition containing the compound can be injection, which includes intravenous injection, intramuscular injection, subcutaneous injection, intradermal injection and acupoint injection.

[0038] The dosage form for administration can be a liquid dosage form or a solid dosage form. For example, the liquid dosage form can be a true solution, a colloid, a microparticle dosage form, an emulsion dosage form, or a suspension dosage form. Other dosage forms include tablets, capsules, dripping pills, aerosols, pills, powders, solutions, suspensions, emulsions, granules, suppositories, freeze-dried powder injections, etc. Preferably, the dosage form is selected from tablets, capsules, pills, granules, oral solutions, and suspensions.

[0039] The compound of the present invention can be prepared into common preparations, sustained-release preparations, controlled-release preparations, targeted preparations and various microparticle drug delivery systems.

[0040] For example, in order to formulate a unit dosage form into tablets, various carriers well known in the art can be widely used. Examples of carriers include diluents and absorbents, such as starch, dextrin, calcium sulfate, lactose, mannitol, sucrose, sodium chloride, glucose, urea, calcium carbonate, kaolin, microcrystalline cellulose, aluminum silicate, etc.; wetting agents and binders, such as water, glycerol, polyethylene glycol, ethanol, propanol, starch slurry, dextrin, syrup, honey, glucose solution, acacia slurry, gelatin slurry, sodium carboxymethyl cellulose, shellac, methylcellulose, potassium phosphate, polyvinyl pyrrolidone, etc.; disintegrants, such as dry starch, alginate, agar powder, brown seaweed starch, sodium bicarbonate and citric acid, calcium carbonate, polyoxyethylene sorbitan fatty acid esters, sodium lauryl sulfate, methylcellulose, ethylcellulose, etc.; disintegration inhibitors, such as sucrose, tristearin, cocoa butter, hydrogenated oil, etc.; absorption promoters, such as quaternary ammonium salts, sodium lauryl sulfate, etc.; lubricants, such as talc, silicon dioxide, corn starch, stearate, boric acid, liquid paraffin, polyethylene glycol, etc. The tablets can be further made into coated tablets, such as sugar-coated tablets, film-coated tablets, enteric-coated tablets, or double-layer tablets and multi-layer tablets.

[0041] For example, in order to make the dosing unit into a pill, various carriers known in the art can be widely used. Examples of carriers include diluents and absorbents, such as glucose, lactose, starch, cocoa butter, hydrogenated vegetable oil, polyvinyl pyrrolidone, monostearate glyceride, kaolin, talc, etc.; binders, such as gum arabic, tragacanth gum, gelatin, ethanol, honey, liquid sugar, rice paste or flour paste, etc.; disintegrants, such as agar powder, dry starch, alginate, sodium dodecyl sulfate, methyl cellulose, ethyl cellulose, etc.

[0042] For example, in order to prepare the dosing unit into a capsule, the active ingredient compound of the present invention is mixed with the above-mentioned various carriers, and the resulting mixture is placed in a hard gelatin capsule or a soft capsule. The active ingredient compound of the present invention can also be prepared into a microcapsule, suspended in an aqueous medium to form a suspension, or loaded into a hard capsule or prepared as an injection for use.

[0043] For example, the compound of the present invention is made into an injection preparation, such as a solution, suspension, emulsion, or freeze-dried powder injection. This preparation may be aqueous or non-aqueous, and may contain one or more pharmacodynamically acceptable carriers, diluents, adhesives, lubricants, preservatives, surfactants, or dispersants. For example, the diluent may be selected from water, ethanol, polyethylene glycol, 1,3-propylene glycol, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol lipids, fatty acid esters, and the like. In addition, in order to prepare an isotonic injection, an appropriate amount of sodium chloride, glucose, or glycerol may be added to the injection preparation. In addition, conventional cosolvents, buffers, pH regulators, and the like may also be added. These adjuvants are commonly used in the art.

[0044] In addition, if necessary, colorants, preservatives, perfumes, flavoring agents, sweeteners or other materials may be added to the pharmaceutical preparations.

[0045] In order to achieve the purpose of medication and enhance the therapeutic effect, the drug or pharmaceutical composition of the present invention can be administered by any known administration method.

[0046] The dosage of the pharmaceutical composition of the compound of the present invention depends on many factors, such as the nature and severity of the disease to be prevented or treated, the sex, age, weight, personality and individual response of the patient or animal, the route of administration, the number of administrations, and the purpose of treatment. Therefore, the therapeutic dose of the present invention can vary widely. Generally speaking, the dosage of the pharmaceutical ingredients in the present invention is well known to those skilled in the art. It can be appropriately adjusted according to the actual amount of the drug contained in the final preparation of the compound composition of the present invention to achieve the requirements of its therapeutically effective amount and complete the prevention or treatment purpose of the present invention. The daily suitable dosage range of the compound of the present invention: the dosage of the compound of the present invention is 0.001-100 mg / kg body weight, preferably 0.1-60 mg / kg body weight, more preferably 1-30 mg / kg body weight, and most preferably 2-15 mg / kg body weight. The compound of the present invention taken by adult patients is 10-500 mg per day, preferably 20-100 mg, which can be taken once or in 2-3 times; the dosage for children is 5-30 mg per kg body weight, preferably 10-20 mg / kg body weight. The above dosage can be administered in a single dosage form or divided into several, such as two, three or four dosage forms, which is limited by the clinical experience of the administering physician and the dosage regimen of the treatment. The compound or composition of the present invention can be taken alone or in combination with other therapeutic drugs or symptomatic drugs.

[0047] The fourth aspect of the present invention provides a compound of formula (I) or a pharmaceutically acceptable salt thereof, or the use of the pharmaceutical composition in the preparation of a drug for preventing and / or treating ischemic cerebrovascular disease. Optionally, the product is selected from medicines and health products.

[0048] Beneficial effects:

[0049] 1. The 5 / 6 / 7 tricyclic adducts of a class of flavanoids and stilbenes connected via a 2-cycloheptene-1-one skeleton of the present invention have the potential to be further developed into new drugs for treating cerebral ischemic diseases.

[0050] 2. The synthesis route of the 5 / 6 / 7 tricyclic adducts of a type of flavanoids and stilbenes connected via a 2-cycloheptene-1-one skeleton is mature and easy to obtain.

[0051] 3. The preparation method of the 5 / 6 / 7 tricyclic adduct of a type of flavanoid and a stilbene compound connected by a 2-cycloheptene-1-one skeleton of the present invention has a simple route, good stereoselectivity and simple separation.

[0052] 4. The 5 / 6 / 7 tricyclic adduct of the flavanone and the diphenylethylene compound connected by the 2-cycloheptene-1-one skeleton has good anti-cerebral ischemic activity, and there is no relevant report on the anti-cerebral ischemic activity of this type of compound. BRIEF DESCRIPTION OF THE DRAWINGS

[0053] Figure 1 is a schematic diagram of the effect of compound 1 on the behavior of rats after middle cerebral artery occlusion injury (mean±SD);

[0054] Figure 2 Schematic diagram of the effect of compound 1 on the infarct volume of rats with middle cerebral artery occlusion injury (mean±SD) DETAILED DESCRIPTION

[0055] The following will be further described in conjunction with specific examples, but the protection scope of the present invention is not limited thereto:

[0056] Example 1 Synthesis of Compound 1, the molecular structure is as follows:

[0057]

[0058] The preparation and detection of compound 1 are as follows:

[0059] 1. Obtaining the flavanoid polymer part of Rhodiola rosea

[0060] The dried rhizome of Rhodiola rosea is used as the primary drug, which is crushed and extracted three times with 80% ethanol by reflux, and the extracts are combined and filtered, and then concentrated under reduced pressure to obtain an extract. After water precipitation treatment, the precipitate is filtered, and the filtrate is concentrated under reduced pressure and separated with HP-20 macroporous adsorption resin, and eluted with three gradients of pure water, 50% ethanol and 95% ethanol. The eluate of 50% ethanol is concentrated under reduced pressure, and the Rhodiola rosea flavanone polymer part is obtained after freeze-drying (see Chinese patent, CN201010587820.7 for details).

[0061] 2. Synthesis of intermediate compound 1-1.

[0062]

[0063] Take a certain amount of Rhodiola rosea flavanoid polymer part, dissolve it in anhydrous methanol at a mass ratio of 1:30, add the corresponding thio reagent at a ratio of 1:0.5 (mass ratio), and add 48% HBr aqueous solution (mass ratio) at a ratio of 1:0.5, react at 60°C for 4h, add distilled water with a mass ratio of 3:1 to the reaction solution to suspend it, extract it with ethyl acetate three times, combine the organic layers, dry the organic layers over anhydrous sodium sulfate, and concentrate under reduced pressure, then perform reverse phase rapid preparative column chromatography separation in sequence, and obtain 4-(S)-(benzylthio)-epicatechin gallate shown in formula 1-1 with an efficient preparative liquid phase of 60% (methanol / water volume ratio), white powder, and a yield of 41.5%.

[0064] HRESIMS(m / z 563.1018[MH]-,calcd,563.1012); 1 H NMR (500MHz, CD 3 OD)δ H :7.45-7.48(m,2H,2″′,6″′-H),7.28-7.33(m,2H,3″′,5″′-H),7.20′7.25(m,1H,4″′-H) ,6.90(s,2H,2″,6″-H),6.89(brs,1H,2′-H),6.67-6.71(m,2H,5′,6′-H),5.94(d,J=2.0H z,1H,8-H),5.93(d,J=2.0Hz,1H,6-H),5.49(brs,1H,2-H),5.38(dd,J=2.5,1.0Hz,1H,3 -H), 4.16 (d, J = 2.0Hz, 1H, 4-H), 4.11 (d, J = 13.5Hz, 1H, SCH), 4.03 (d, J = 13.5Hz, 1H, SCH).

[0065] 3. Synthesis of intermediate compound 2-1.

[0066]

[0067] 564 mg (1 mmol) of 4-benzylthio epicatechin gallate (Compound 1-1) was weighed into a 100 mL eggplant-shaped bottle, and then 20 mL of N,N-dimethylformamide was added. After stirring and dissolving, 1220 mg (5 mmol) of piceatannol and 1100 mg (5 mmol) of silver trifluoroacetate were added successively at -10°C. After stirring and reacting for 4 hours at -10°C, the reaction was completed by high performance liquid chromatography. 400 mL of distilled water was added to the reaction solution to terminate the reaction, and the mixture was extracted with ethyl acetate (400 mL × 3 times). The organic layers were combined, washed with saturated sodium bicarbonate solution and saturated sodium chloride solution, dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and separated by reverse phase rapid preparative column chromatography (acetonitrile: water = 25:75) to obtain 233.9 mg of the product with a yield of 34.2%.

[0068] HRESIMS (m / z 685.1551 [M+H] + ,calcd,685.1552); 1 H NMR (500 MHz, DMSO-d 6 )δ H :5.51(1H,brs,2-H),5.25(1H,m,3-H),4.59(1H,brs,4-H),5.74(1H,d,J=2.0Hz,6-H),5.97(1H,d,J=2.0Hz, 8-H),6.49(1H,d,J=2.0Hz,2′-H),6.72(1H,overlap,5′-H),6.66(1H,overlap,6′-H),6.88(2H,brs,2″,6″- H),6.08(1H,d,J=2.5Hz,4″′-H),6.31(1H,brs,6″′-H),7.62(1H,d,J=16.0Hz,7″′-H),6.45(1H,d,J=16.0Hz ,8″′-H),6.89(1H,d,J=8.5Hz,10″′-H),6.59(1H,overlap,13″′-H),7.00(1H,dd,J=2.0Hz,8.5Hz,14″′-H); 13 C NMR (125 MHz, DMSO-d 6 )δ C:74.4(C-2),72.7(C-3),34.8(C-4),157.6(C-5),94.8(C-6),157.2(C-7),95.3(C-8),156.3(C-9),101.3(C-10),130.8(C-1 ′),115.8(C-2′),144.8(C-3′),145.0(C-4′),117.4(C-5′),119.1(C-6′),144.5(C-1″),108.8(C-2″,6″),146.4(C-3″,5″), 140.0(C-4″),165.3(C=O),142.5(C-1″′),117.4(C-2″′),156.8(C-3″′),103.0(C-4″′),156.4(C-5″′),105.3(C-6″′),125. 0(C-7″′), 129.7(C-8″′), 129.5(C-9″′), 115.1(C-10″′), 145.2(C-11″′), 145.5(C-12″′), 116.0(C-13″′), 119.5(C-14″′).

[0069] 4. Synthesis of compound 1.

[0070]

[0071] Weigh 1231 mg (1.8 mmol) of compound 2-1 into a 250 mL round-bottom flask, then add 100 mL of distilled water, stir to dissolve, then dropwise add an aqueous solution of copper sulfate pentahydrate [153.4 mg (0.9 mmol) dissolved in 15 mL of distilled water], stir and react for 5 h at room temperature, and monitor the completion of the reaction by high performance liquid chromatography. Add ethyl acetate for extraction (100 mL × 3 times), combine the organic layers, wash with saturated sodium chloride solution, dry over anhydrous sodium sulfate, evaporate to dryness under reduced pressure, separate by reverse phase rapid preparative column chromatography (methanol: water = 25:75), and obtain 910 mg of the product with a yield of 77.1%.

[0072] HRESIMS (m / z 657.1597 [M+H] + ,calcd,657.1530); 1 H NMR (500 MHz, DMSO-d 6 )δ H:5.14(2H,brs,2,3-H),3.81(1H,m,4-H),2.30(1H,d,J=13.5Hz,6a-H),3.31(1H,overlap,6b-H)5.58(1H,brs,8-H ),4.20(1H,td,J=10.0Hz,10.0Hz,10-H),6.80(1H,d,J=2.0Hz,2′-H),6.59(1H,overlap,5′-H),6.66(1H,overlap, 6′-H),6.79(2H,brs,2″,6″-H),5.93(1H,d,J=2.5Hz,4″′-H),5.69(1H,brs,6″′-H),3.81(1H,overlap,7″′-H),3.1 1(1H,m,8″′-H),6.65(1H,overlap,10″′-H),6.72(1H,d,J=8.5Hz,13″′-H),6.56(1H,dd,J=2.0Hz,8.5Hz,14″′-H); 13 C NMR (125 MHz, DMSO-d 6 )δ C :80.9(C-2),71.9(C-3),43.1(C-4),50.5(C-6),197.1(C-7),111.0(C-8),175.9(C-9),46.0(C-10),138.1(C-1′),115. 7(C-2′),145.1(C-3′),145.2(C-4′),117.6(C-5′),119.3(C-6′),145.3(C-1″),108.8(C-2″,6″),145.5(C-3″,5″),138. 1(C-4″),164.3(C=O),143.9(C-1″′),135.7(C-2″′),154.8(C-3″′),100.9(C-4″′),158.2(C-5″′),101.1(C-6″′),55.1( C-7″′), 50.5(C-8″′), 127.9(C-9″′), 117.6(C-10″′), 145.6(C-11″′), 145.5(C-12″′), 118.5(C-13″′), 119.3(C-14″′).

[0073] Example 2 Synthesis of Compound 2, the molecular structure is as follows:

[0074]

[0075] The preparation and detection of compound 2 are as follows

[0076] 1. The acquisition of the flavanone polymer part of Rhodiola rosea, the synthesis of the intermediate compound 1-1, and the synthesis of the intermediate compound 2-1 were the same as in Example 1.

[0077] 2. Synthesis of Intermediate 3-1

[0078]

[0079] Weigh 100 mg of compound 2-1 into a 50 mL round-bottom flask, add 10 mL of distilled water, stir to dissolve, and then replace with argon for protection. Add 10 mg of tannase at room temperature, stir at room temperature for 2 hours, and monitor the reaction completion by HPLC. Directly freeze-dry the reaction solution, separate by reverse phase rapid preparative column chromatography (acetonitrile: water = 25:75), and obtain 69.6 mg of product with a yield of 89.5%.

[0080] HRESIMS (m / z 533.1309 [M+H] + ,calcd,533.1369); 1 H NMR (500 MHz, DMSO-d 6 )δ H :4.68(1H,d,J=5.0Hz,2-H),3.65(1H,m,3-H),4.49(1H,d,J=2.0Hz,4-H),5.83(1H,d,J=2.0Hz,6-H),5.90(1H ,d,J=2.0Hz,8-H),6.57(1H,d,J=2.0Hz,2′-H),6.70(1H,overlap,5′-H),6.41(1H,dd,J=2.0Hz,8.5Hz,6′-H) ,6.27(1H,d,J=2.5Hz,4″′-H),6.29(1H,d,J=2.5Hz,6″′-H),6.57(1H,d,J=16.0Hz,7″′-H),6.34(1H,d,J=16. 0Hz, 8″′-H), 6.53 (1H, overlap, 10″′-H), 6.70 (1H, overlap, 13″′-H), 6.38 (1H, dd, J = 2.0Hz, 8.5Hz, 14″′-H); 13 CNMR (125MHz, DMSO-d 6 )δ C:75.2(C-2),71.5(C-3),37.9(C-4),16.2(C-5),94.8(C-6),156.7(C-7),95.4(C-8),156.9(C-9),102.6 (C-10),130.9(C-1′),115.2(C-2′),145.1(C-3′),145.0(C-4′),117.4(C-5′),118.1(C-6′),139.8(C-1″ ′),116.0(C-2″′),157.3(C-3″′),105.4(C-4″′),157.2(C-5″′),103.4(C-6″′),125.8(C-7″′),124.5(C -8″′), 129.2(C-9″′), 115.4(C-10″′), 145.9(C-11″′), 145.8(C-12″′), 115.0(C-13″′), 119.1(C-14″′).

[0081] 5. Synthesis of Compound 2

[0082]

[0083] Weigh 86 mg (0.16 mmol) of compound 3-1 into a 50 mL round-bottom flask, then add 15 mL of H 2 O, stirred and dissolved, then added dropwise an aqueous solution of copper sulfate pentahydrate [20.0 mg (0.08 mmol) dissolved in 5 mL H 2 O], stirred for 4 hours at room temperature, and the reaction was completed by high performance liquid chromatography. The product was precipitated as a white solid, filtered and washed to obtain 31.2 mg of crude product, and the mother liquor was extracted with ethyl acetate (50 mL × 3 times), and the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, combined with the filter residue, and separated by reverse phase rapid preparative column chromatography (methanol: water = 50:50), to obtain 28.0 mg of product, with a yield of 35.7%.

[0084] HRESIMS (m / z 505.1495 [M+H] + ,calcd,505.1420); 1 H NMR (500 MHz, DMSO-d 6 )δ H:5.44(1H,brs,2-H),4.11(2H,td,J=10.0Hz,10.0Hz,2.0Hz,3-H),3.64(1H,overlap,4-H),2.23(1H,m,6a-H),3.23( 1H,dd,J=13.0Hz,8.0Hz,6b-H),5.67(1H,brs,8-H),3.08(1H,m,10-H),6.78(1H,d,J=2.0Hz,2′-H),6.69(1H,overla p,5′-H),6.68(1H,overlap,6′-H),6.01(1H,d,J=2.5Hz,4″′-H),6.17(1H,brs,6″′-H),3.45(1H,m,7″′-H),3.69(1H ,overlap,8″′-H),6.57(1H,d,J=2.5Hz,10″′-H),6.64(1H,overlap,13″′-H),6.47(1H,dd,J=2.5Hz,8.5Hz,14″′-H); 13 C NMR (125 MHz, DMSO-d 6 )δ C :83.2(C-2),72.5(C-3),45.9(C-4),45.4(C-6),197.5(C-7),102.7(C-8),177.3(C-9),51.0(C-10), 128.7(C-1′),115.6(C-2′),145.5(C-3′),145.8(C-4′),116.2(C-5′),118.8(C-6′),147.8(C-1″′),1 27.8(C-2″′),154.7(C-3″′),101.8(C-4″′),159.0(C-5″′),107.4(C-6″′),56.3(C-7″′),44.7(C-8″ ′), 133.9(C-9″′), 115.9(C-10″′), 145.5(C-11″′), 144.3(C-12″′), 116.2(C-13″′), 120.0(C-14″′).

[0085] Example 3 Synthesis of Compound 3, the molecular structure is as follows:

[0086]

[0087] The preparation and detection of compound 3 are as follows:

[0088] 1. Obtaining the flavanone polymer part of Rhodiola rosea. The synthesis of the intermediate compound 1-1 is the same as in Example 1.

[0089] 2. Synthesis of Compound 3

[0090] Weigh 564 mg (1 mmol) of 4-benzylthio epicatechin gallate (Compound 1-1) into a 100 mL eggplant-shaped bottle, then add 20 mL of N,N-dimethylformamide, stir to dissolve, then add 1300 mg (5 mmol) of 5-[(1E)-2-(3,5-dihydroxyphenyl)vinyl]-1,2,3-pyrogallol and 1100 mg (5 mmol) of silver trifluoroacetate at -10°C in sequence, stir and react at -10°C for 4 h, then add 100 mL of distilled water and 62.4 mg (0.25 mmol) of copper sulfate pentahydrate directly to the reaction solution without monitoring, stir and react at room temperature for 2 h, and monitor the reaction completion by high performance liquid chromatography. The mixture was extracted with ethyl acetate (400 mL × 3 times), and the organic layers were combined and washed with saturated sodium bicarbonate solution and saturated sodium chloride solution, dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and separated by reverse phase flash preparative column chromatography (methanol: water = 25:75) to obtain 75.3 mg of the product with a total yield of 11.2% for two steps.

[0091] HRESIMS (m / z 673.1550 [M+H] + ,calcd,673.1552); 1 H NMR (500 MHz, DMSO-d 6 )δ H :5.13(1H,overlap,2-H),5.13(1H,overlap,3-H),4.18(1H,t,J=9.0Hz,4-H),2.30(1H,d,6a-H),3.29(1H, dd,J=13.0Hz,7.5Hz,6b-H),5.72(1H,brs,8-H),3.01(1H,t,J=9.0Hz,10-H),6.79(1H,overlap,2′-H),6.6 4(1H,J=8.0Hz,2.0Hz,5′-H),6.59(1H,J=8.0Hz,6′-H),6.79(2H,overlap,4″,6″-H),5.58(1H,brs,6″′-H) ,3.76(1H,m,7″′-H),3.81(1H,overlap,8″′-H),6.21(2H,brs,10″′,14″′-H),5.93(1H,J=8.0Hz,12″′-H); 13 C NMR (125 MHz, DMSO-d 6 )δ C:84.4(C-2),79.6(C-3),46.0(C-4),50.5(C-6),197.1(C-7),101.0(C-8),175.9(C-9),55.0(C-10),127.9(C-1′),114.5(C-2 ′),145.2(C-3′),145.2(C-4′),116.2(C-5′),115.0(C-6′),164.3(OC=O),120.0(C-1″),108.9(C-2″),145.5(C-3″),138.2(C- 4″),116.2(C-5″),108.9(C-6″),135.0(C-1″′),119.3(C-2″′),145.1(C-3″′),131.6(C-4″′),147.8(C-5″′),111.0(C-6″′), 56.8(C-7″′), 45.0(C-8″′), 145.7(C-9″′), 106.3(C-10″′), 154.8(C-11″′), 101.2(C-12″′), 158.3(C-13″′), 106.3(C-14″′).

[0092] Example 4 Synthesis of Compound 4, the molecular structure is as follows:

[0093]

[0094] The preparation and detection of compound 4 are as follows:

[0095] 1. Obtaining the flavanone polymer part of Rhodiola rosea. The synthesis of the intermediate compound 1-1 is the same as in Example 1.

[0096] 2. Synthesis of intermediate compound 4-1.

[0097]

[0098] Weigh 564 mg (1 mmol) of 4-benzylthio epicatechin gallate (Compound 1-1) into a 100 mL eggplant-shaped bottle, then add 20 mL of N,N-dimethylformamide, stir to dissolve, then add 1140 mg (5 mmol) of resveratrol and 1100 mg (5 mmol) of silver trifluoroacetate at -10°C, stir at -10°C for 4 hours, and the reaction is complete by high performance liquid chromatography. Add 400 mL of distilled water to the reaction solution to terminate the reaction, extract with ethyl acetate (400 mL × 3 times), combine the organic layers, wash with saturated sodium bicarbonate solution and saturated sodium chloride solution, dry with anhydrous sodium sulfate, evaporate to dryness under reduced pressure, separate by reverse phase rapid preparative column chromatography (acetonitrile: water = 25:75), and obtain 278.6 mg of the product with a yield of 41.7%.

[0099] HRESIMS (m / z 669.1602 [M+H] + , calcd, 669.1603); 1 1H NMR (500 MHz, DMSO-d 6 ) δ H : 5.51 (1H, brs, 2-H), 5.24 (1H, m, 3-H), 4.58 (1H, brs, 4-H), 5.84 (1H, d, J = 2.0 Hz, 6-H), 5.99 (1H, d, J = 2.0 Hz, 8-H), 6.49 (1H, overlap, 2′-H), 6.74 (1H, overlap, 5′-H), 6.66 (1H, dd, J = 8.0 Hz, 2.0 Hz, 6′-H), 6.90 (2H, brs, 2″, 6″-H), 6.49 (1H, d, J = 2.5 Hz, 4″′-H), 6.61 (1H, brs, 6″′-H), 7.68 (1H, d, J = 15.0 Hz, 7″′-H), 6.69 (1H, d, J = 15.0 Hz, 8″′-H), 7.43 (2H, d, J = 8.5 Hz, 10″′, 14″′-H), 6.77 (2H, d, J = 8.5 Hz, 11″′, 13″′-H); 13 13C NMR (125 MHz, DMSO-d 6 ) δ C : 73.9 (C-2), 72.5 (C-3), 36.0 (C-4), 156.9 (C-5), 94.5 (C-6), 157.2 (C-7), 95.9 (C-8), 155.9 (C-9), 101.5 (C-10), 130.3 (C-1′), 114.0 (C-2′), 144.6 (C-3′), 144.9 (C-4′), 115.2 (C-5′), 117.1 (C-6′), 119.5 (C-1″), 108.7 (C-2″, 6″), 145.5 (C-3″, 5″), 139.9 (C-4″), 165.5 (C=O), 138.9 (C-1″′), 117.4 (C-2″′), 156.4 (C-3″′), 103.2 (C-4″′), 156.6 (C-5″′), 105.2 (C-6″′), 125.0 (C-7″′), 128.6 (C-8″′), 129.5 (C-9″′), 128.0 (C-10″′), 115.6 (C-11″′), 157.3 (C-12″′), 115.6 (C-13″′), 128.0 (C-14″′).

[0100] 3. Synthesis of Compound 4.

[0101]

[0102] Weigh 1202 mg (1.8 mmol) of compound 4-1 into a 250 mL round-bottom flask, then add 100 mL of distilled water, stir to dissolve, then dropwise add an aqueous solution of ferric chloride hexahydrate [243.3 mg (0.9 mmol) dissolved in 15 mL of distilled water], stir and react at 35 ° C for 22 h, and the reaction is complete by high performance liquid chromatography. Add ethyl acetate for extraction (100 mL × 3 times), combine the organic layers, wash with saturated sodium chloride solution, dry over anhydrous sodium sulfate, evaporate to dryness under reduced pressure, and separate by reverse phase rapid preparative column chromatography (methanol: water = 25:75), to obtain 186.6 mg of the product, with a yield of 16.2%.

[0103] HRESIMS (m / z 641.1654 [M+H] + ,calcd,641.1640); 1 H NMR (500 MHz, DMSO-d 6 )δ H :5.15(2H,brs,2,3-H),3.82(1H,s,4-H),2.29(1H,d,J=13.5Hz,6a-H),3.33(1H,overlap,6b-H)5.58(1H,brs ,8-H),4.21(1H,td,J=10.0Hz,2.0Hz,10-H),6.83(1H,d,J=2.0Hz,2′-H),6.58(1H,overlap,5′-H),6.66(1H,o verlap,6′-H),6.78(2H,brs,2″,6″-H),5.92(1H,d,J=2.0Hz,4″′-H),5.65(1H,d,J=2.0Hz,6″′-H),3.82(1H,o verlap,7″′-H),3.19(1H,m,8″′-H),7.08(2H,d,J=8.5Hz,10″′,14″′-H),6.76(2H,d,J=8.5Hz,11″′,13″′-H); 13 C NMR (125 MHz, DMSO-d 6 )δ C:80.9(C-2),72.1(C-3),44.7(C-4),46.1(C-6),197.1(C-7),101.0(C-8),176.0(C-9),46.0(C-10),127.9(C-1′), 114.7(C-2′),145.1(C-3′),145.6(C-4′),114.9(C-5′),117.6(C-6′),119.4(C-1″),108.85(C-2″,6″),145.2(C-3 ″,5″),145.02(C-4″),164.3(C=O),128.8(C-1″′),119.6(C-2″),154.8(C-3″),100.9(C-4″′),156.0(C-5″′),110. 9(C-6″′), 50.4(C-7″′), 43.3(C-8″′), 135.0(C-9″′), 128.8(C-10″′,14″′), 115.5(C-11″′,13″′), 158.2(C-12″′).

[0104] Experimental Example 1 In vitro screening model: Screening of protective activity against neuronal cell damage induced by glutamate or hydrogen peroxide

[0105] L-Glutamate is a major excitatory neurotransmitter in the central nervous system, distributed in the cerebral cortex and hippocampal neurons. When the glutamate content is elevated, it becomes a neurotoxin, inducing severe neuronal damage. Overactivation of neurons mediated by glutamate or overstimulation of glutamate can lead to cell death. Hydrogen peroxide is a strong oxidant that causes lipid peroxidation by producing hydroxyl radicals, amplifying free radical reactions and causing damage to cells. The neuronal cell injury model induced by glutamate or hydrogen peroxide has become an effective model for studying neuronal cell injury, which is helpful to explore the neuroprotective effect and mechanism of cycloheptenone and its synthetic intermediates of a class of flavanoid and diphenylethylene adducts in the present invention.

[0106] 1 Materials and methods

[0107] 1.1 Materials

[0108] Fetal calf serum (FCS) and DMEM sugar-free culture medium were Invitrogen products; trypsin was purchased from Gibco; centrifuge tubes, 96-well cell culture plates, cell culture dishes, and cell culture flasks were Corning products; cover slips were DIAMOND products; and dimethyl sulfoxide was a Sinopharm product. Other reagents were purchased from Sigma.

[0109] 1.2 Methods

[0110] PC-12 cells were collected at 1.5×10 4 The density of cells / mL was 100 μL per well in a 96-well culture plate. The experiment was divided into a blank group, a model group, a drug-added group, and a positive drug group. Before oxygen and glucose deprivation, the incubator was set to 94% N 2 , 5% CO 2 , 1% O 2 , 37℃ for 1h. The original cell culture medium was discarded, and an equal amount of preheated sugar-free DMEM culture medium was added, and the cells were placed in an oxygen-deficient incubator for 2h. Then, under normal culture conditions, 10μM of compound or edaravone was added to the drug group and positive drug group, and an equal amount of culture medium was added to the model group. The changes in cell morphology and survival rate were observed at different time points of oxygen-glucose deprivation for 2h and reoxygenation and resugarization.

[0111] PC-12 cells were collected at 1.5×10 4 The density of cells / mL was 100 μL per well in a 96-well culture plate. The experiment was divided into a blank group, a model group, a drug-added group, and a positive drug group. The final concentration of 120 μM H 2 O 2 , 8mM glutamate for 2h, then the blank group was cultured with complete medium, the positive drug group was added with 10μM edaravone, and the drug-added group was added with 10μM of the test compound. After incubation for 24h, 10μL CCK-8 was added to each well, and the absorbance at 490nm was measured using a microplate reader after 60min to calculate the survival rate.

[0112] 2 Results

[0113] In vitro pharmacological experiments showed that most of these compounds and their intermediates have neuroprotective activity. Among them, the adduct 1 of flavanone with 2-cycloheptene-1-one structure and diphenylethylene has a good protective effect on glutamate and hydrogen peroxide-induced cell damage, and the cell survival rate was increased by 15.73% and 31.64%, respectively. The survival rate was increased = (survival rate of the drug administration group - survival rate of the model group) / (survival rate of the model group). The structures and pharmacological activity results of some of the more active compounds are shown in the following table.

[0114]

[0115] Table 1. In vitro neuroprotective activity screening results

[0116]

[0117] Experimental Example 2 In vivo screening model: Study on the preventive and therapeutic effects on cerebral ischemia-reperfusion injury

[0118] The middle cerebral artery occlusion model is a classic animal model for studying ischemic stroke. This model uses a suture inserted into the skull via the internal carotid artery to block the blood supply to the contralateral anterior cerebral communicating artery and the middle cerebral artery. This experiment has the advantages of no craniotomy, definite effect, and accurate control of ischemia and reperfusion time.

[0119] 1 Materials and methods

[0120] 1.1 Materials

[0121] (1) Test product: Compound 1. Purity ≥ 90%, yellow solid powder, 560 mg in total, easily soluble in dimethyl sulfoxide (DMSO).

[0122] (2) Reagents: 2% TTC staining solution, purchased from Shanghai Yuanye Biotechnology Co., Ltd.

[0123] (3) Animals: Male SD rats weighing 280-300 g, SPF grade, purchased from Sibeifu (Beijing) Biotechnology Co., Ltd.

[0124] (4) Equipment: rat operating table, surgical instruments, nylon thread plug (diameter 0.24 mm), sutures, etc.

[0125] 1.2 Methods

[0126] Middle cerebral artery occlusion model

[0127] The experimental subjects were SD (280-300g) male rats. After the rats were anesthetized with isoflurane, the skin and subcutaneous tissue were cut along the midline of the neck to expose the ECA, CCA and ICA blood vessels. The surgical thread was used to ligate the proximal end of the CCA and the distal end of the ECA respectively. At the same time, the surgical thread was used to pass through the ECA and tie a slipknot near the bifurcation of the ICA and ECA, and the ICA was clamped with an artery clamp. Then, an incision was made between the two ligatures of the ECA with microscissors. The thread plug (diameter 0.36±0.02mm) was gently inserted from the ECA into the inner cavity of the ICA to a position of about 18mm, blocking the contralateral blood flow of the middle cerebral artery to form ischemia. After 90 minutes of ischemia, the thread plug was removed and the neck was sutured. The sham group (Sham group) did not perform the plug insertion operation, and other surgical steps were the same. After the rats woke up, neurological function scores were performed, and rats without neurological deficits and subarachnoid hemorrhage symptoms were removed. The remaining rats were randomly divided into two groups, namely the model group (Vehicle group) and the drug-treated group (Compound 1 group). The drug-treated group was gavaged with Compound 1 (30 mg / kg), and the sham operation and model groups were gavaged with the same volume of normal saline. The rats were placed on a warming pad throughout the ischemic stage to maintain a body temperature of 37°C.

[0128] 2 Neurological deficit scores in rats

[0129] On the 1st, 3rd and 7th day after the middle cerebral artery occlusion model was established, the Longa scoring method was used to score the neurological deficit of the rats, and the degree of neurological deficit was divided into 5 levels: rats with 0 and 4 points were excluded for unsuccessful modeling, and 1-3 points were considered successful models. Throughout the experiment, the experimenter used a single-blind method to avoid random errors. The results are shown in Figure 1 shown.

[0130] 3TTC staining

[0131] After the rats were anesthetized, they were quickly decapitated and the brains were removed. The brain tissues were placed in a brain mold for coronal sectioning, with each slice being 2 mm thick, for a total of 6 slices. The brain slices were placed in a 2% TTC staining solution and incubated at 37°C in the dark for 10-15 minutes. The brain slices were taken out, fixed with 4% paraformaldehyde and photographed for recording. Active brain tissue contains dehydrogenase, which can reduce TTC to red, so it is stained red. In the infarcted brain tissue, the dehydrogenase activity disappears, so it is not stained. Therefore, the slices are stained in two colors, with normal brain tissue being red and the infarcted area being white. The images were processed with Image J software to calculate the infarct volume.

[0132] 2 Experimental results

[0133] (1) The results of neurological deficit scoring in rats are as follows Figure 1 As shown, compared with the sham operation group, the rats in the model group had obvious neurological damage. After continuous administration for one week, compound 1 could effectively restore the neurological function of the rats;

[0134] (2) TTC staining results Figure 2 As shown, compound 1 can significantly reduce the volume of cerebral infarction, indicating that it has good anti-ischemic stroke activity.

[0135] The above embodiments are not limitations of the present invention, and the present invention is not limited to the above embodiments. As long as the requirements of the present invention are met, they belong to the protection scope of the present invention.

Claims

1. A compound of formula (I) or a pharmaceutically acceptable salt thereof, characterized in that: in, R A Selected from: hydrogen, hydroxyl, R B Selected from hydrogen, hydroxy, alkoxy, halogen, cyano; a is selected from integers of 0-5; b is selected from integers of 0-4; c is selected from integers of 0-5; d is selected from integers of 0-3.

2. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: The halogen is selected from F, Cl, Br, I; Optionally, the alkyl group in the "alkyl", "haloalkyl" or "alkylsulfonyl" is C1-C 10 A straight chain or branched alkyl group, preferably a C1-C7 straight chain or branched alkyl group, preferably a C1-C5 straight chain or branched alkyl group, preferably selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl, isopentyl, 1-ethylpropyl, neopentyl, n-hexyl, 1-methylpentyl, 2-methylpentyl, 3-methylpentyl ... hexyl, 1,1-dimethylbutyl, 2,2-dimethylbutyl, 3,3-dimethylbutyl, 1,2-dimethylbutyl, 1,3-dimethylbutyl, 2,3-dimethylbutyl, 2-ethylbutyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 2,2-dimethylpentyl, 3,3-dimethylpentyl, 2,3-dimethylpentyl, 2,4-dimethylpentyl, 3-ethylpentyl or 2,2,3-trimethylbutyl.

3. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: a is an integer selected from 0-3; optionally, a is 0, 1, 2 or 3; Optionally, b is selected from an integer of 2-3; Optionally, b is 2 or 3; Optionally, c is selected from an integer of 0-3; Optionally, c is 0, 1, 2 or 3; Optionally, d is 0, 1, 2 or 3.

4. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: Selected from the following compounds:

5. A method for preparing a compound of formula (I) or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4, characterized in that: The following steps are involved: (1) Hydrobromic acid catalyzes the depolymerization to form a flavanone monomer derivative with a thio group connected at the 4 position, (2) Silver trifluoroacetate catalyzed substitution reaction to synthesize an intermediate in which the 4-position of flavanone is connected to the 2-position of stilbene. (3) Synthesis of a class of 5 / 6 / 7 tricyclic cycloadducts of flavanoids and stilbenes connected via a 2-cyclohepten-1-one skeleton catalyzed by trivalent iron or divalent copper In various types, R A , R B , a, b, c, d are as defined in any one of claims 1-4.

6. A pharmaceutical composition, characterized in that Containing an effective dose of a compound of the structure represented by formula (I) according to any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

7. The pharmaceutical composition according to claim 6, characterized in that The dosage form of the pharmaceutical composition is selected from tablets, capsules, dripping pills, aerosols, pills, powders, solutions, suspensions, emulsions, granules, suppositories, and lyophilized powder injections; preferably selected from tablets, capsules, pills, granules, oral solutions, and suspensions.

8. Use of the compound of formula (I) or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4, or the pharmaceutical composition according to claim 6 or 7 in the preparation of a product for preventing and / or treating ischemic cerebrovascular disease. Optionally, the product is selected from medicines and health products.

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