Compound extracted from medicine watermelon, pharmaceutical composition of compound and application of compound in liver protection

Abstract

The invention belongs to the field of medicine, and relates to a compound with a novel structure in a medicine, a preparation method of the compound and application of the compound in a medicine with liver protection activity, and the compound has the liver protection activity and can be used for preparing the medicine with the liver protection activity.

Description

Technical Field

[0001] This invention belongs to the field of medicine, specifically relating to the hepatoprotective uses of a class of compounds isolated from medicinal watermelon and their application in the preparation of hepatoprotective drugs. Background Technology

[0002] Liver diseases are a major category of diseases that seriously threaten human health worldwide, including hepatitis caused by various factors, such as fatty liver, alcoholic, viral, and autoimmune hepatitis, as well as liver fibrosis and liver cancer. In recent years, with urbanization, changing lifestyles and dietary structures, environmental pollution, overeating, excessive alcohol consumption, and the abuse of food additives have led to a continuous increase in the incidence of liver diseases. These diseases, including hepatitis, fatty liver, liver fibrosis, cirrhosis, and even liver cancer, are increasingly threatening human health. Therefore, the development of highly effective liver-protective drugs with minimal side effects has attracted widespread attention. Meanwhile, research on the hepatoprotective activity of natural drugs has shown broad application prospects. Thus, developing liver disease treatments with better activity and therapeutic effects from natural products is a problem that researchers urgently need to solve.

[0003] *Citrullus colocynthis* (L.) Schrad., a plant belonging to the genus *Citrullus* in the family Cucurbitaceae, is a medicinal herb originating from the arid regions of North Africa, widespread throughout the Sahara Desert. It is also found in Morocco, Egypt, Sudan, Iran, India, Pakistan, and Afghanistan. During the Song Dynasty, it was transported to Xinjiang via the ancient Silk Road and has been used ever since. Currently, in my country, it is distributed in Northwest, Northeast, and North China, as well as Henan, Hubei, and Xinjiang. It is widely cultivated in Changji, Hotan, and Yecheng to meet clinical needs. In Uyghur medicine, the medicinal part of the watermelon is its dried, ripe fruit, which is considered to have the effects of dispelling phlegm and can be used to treat various headaches, deafness, arthritis, and amenorrhea. Modern pharmacological studies have shown that *Citrullus colocynthis* possesses anti-cancer, hepatoprotective, antioxidant, anti-diabetic, and antimicrobial activities. The medicinal watermelon contains compounds such as cucurbitacin, alkaloids, flavonoids, steroidal saponins, phenolic acids, and phenylpropanoids. Among them, cucurbitacin compounds are the main components and main active ingredients of the medicinal watermelon, and have a variety of biological activities such as anti-tumor, liver protection, and improving the body's immunity. Summary of the Invention

[0004] The applicant's research revealed the isolation of a novel class of compounds from medicinal watermelons, with the following chemical structures:

[0005] Pharmacological experiments have demonstrated that some compounds can effectively improve the cell survival rate and alleviate cell damage in APAP- or H2O2-induced HepG2 cells, showing good potential for development into hepatoprotective drugs. Currently, there are no research reports, preparation methods, or patent documents regarding the use of these compounds in hepatoprotective drugs.

[0006] The technical problem solved by this invention is to provide a method for preparing compounds isolated and extracted from medicinal watermelon and their application in the preparation of liver-protective drugs.

[0007] To solve the technical problem of this invention, the present invention provides the following technical solution:

[0008] The first aspect of the present invention is to provide a class of compounds or pharmaceutically acceptable salts thereof, characterized in that the structure of the class of compounds is as follows:

[0009]

[0010] The pharmaceutically acceptable salts mentioned above are selected from salts formed by compounds and inorganic or organic bases. The organic bases include methylamine, ethylamine, diethylamine, triethylamine, propylamine, butylamine, octylamine, hexamethylenediamine, ethylenediamine, propylenediamine, butylamine, benzylamine, phenethylamine, o-phenylenediamine, and p-phenylenediamine. The inorganic bases include sodium hydroxide, potassium hydroxide, calcium hydroxide, aluminum hydroxide, lithium hydroxide, magnesium hydroxide, zinc hydroxide, copper hydroxide, iron hydroxide, ammonium hydroxide, sodium bicarbonate, and potassium bicarbonate.

[0011] The second aspect of the present invention is to provide a method for preparing the compound described in the first aspect.

[0012] 50 kg of dried watermelon fruit was extracted twice by reflux with 70% ethanol for 2 hours each time. The extract was concentrated and then extracted with petroleum ether, ethyl acetate and n-butanol. The aqueous fraction was concentrated to obtain an extract. The extract was separated by macroporous resin column chromatography, eluting sequentially with water, 15% ethanol, 30% ethanol, 50% ethanol and 95% ethanol to obtain five fractions. The 50% fraction (400 g) from the macroporous resin was separated by LH-20 gel column chromatography, eluting with a gradient of ethanol-water (0%-100%, v / v) to obtain 13 subfractions (AL). The D fraction (27 g) was separated by silica gel column chromatography, eluting with a gradient of dichloromethane and methanol (0%-100%, v / v). After thin-layer chromatography identification, the fractions were combined to obtain 11 components (Da-Dk). The De-Df fractions (1.50 g) were combined and then analyzed by Flash chromatography. Separation was performed using C18 column chromatography with a 0%-100% methanol gradient to obtain De1-De8, followed by semi-preparative HPLC to obtain compounds 1 and 2; Dk (1.86 g) fraction was separated using Flash C18 column chromatography with a 0%-100% methanol gradient to obtain Dk1-Dk10, followed by semi-preparative HPLC to obtain compounds 3, 4, 11, 13, 14, 21; Dc (1.30 g) fraction was separated using Flash C18 column chromatography with a 0%-100% methanol gradient to obtain Dc1-Dc20, followed by semi-preparative HPLC to obtain compounds 5, 6, 7, 8; Dj (0.75 g) fraction was separated using Flash C18 column chromatography with a 0%-100% methanol gradient to obtain Dj1-Dj8, followed by semi-preparative HPLC to obtain compounds 9, 10, 15, 16, 17, 18; Dg (1.00 g) fraction was separated using Flash C18 column chromatography with a 0%-100% methanol gradient to obtain compounds 9, 10, 15, 16, 17, 18; Compound 12 was obtained by C18 column chromatography and elution with a 0%-100% methanol gradient. Compounds 12 were obtained by semi-preparative HPLC. Compounds 19 and 20 were obtained by combining Dh-Di (0.77 g) and separating them by Flash C18 column chromatography and elution with a 0%-100% methanol gradient.

[0013] A third aspect of the present invention provides a pharmaceutical composition comprising the compound described in the first aspect of the present invention or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier or excipient. The dosage form of the pharmaceutical composition may be a liquid dosage form or a solid dosage form. Liquid dosage forms may be true solutions, colloids, microparticles, emulsions, suspensions, etc. Other dosage forms include tablets, capsules, pellets, aerosols, pills, powders, solutions, suspensions, emulsions, granules, suppositories, lyophilized powder injections, etc. The compounds of the present invention can be formulated into conventional formulations, as well as sustained-release formulations, controlled-release formulations, targeted formulations, etc.

[0014] To formulate the drug delivery unit into tablets, a wide variety of carriers known in the art can be used, such as diluents including starch, dextrin, calcium sulfate, lactose, mannitol, sucrose, etc.; humectants and binders such as water, glycerin, polyethylene glycol, ethanol, gelatin, sodium carboxymethyl cellulose, methylcellulose, polyvinylpyrrolidone, etc.; disintegrants such as starch, calcium carbonate, methylcellulose, sodium dodecyl sulfonate, etc.; and lubricants such as talc, silica, stearates, liquid paraffin, polyethylene glycol, etc. To formulate the drug delivery unit into pills, a wide variety of carriers known in the art can be used, such as diluents including glucose, lactose, starch, hydrogenated vegetable oil, polyvinylpyrrolidone, etc.; binders such as gum arabic, gelatin, ethanol, etc.; and disintegrants such as dried starch, sodium dodecyl sulfonate, methylcellulose, etc. To formulate the drug delivery unit into capsules, the compound of the present invention is mixed with the above-mentioned carriers, and the resulting mixture is placed in hard gelatin capsules or soft capsules; the compound of the present invention can also be formulated into microcapsules and suspended in an aqueous medium to form a suspension for application.

[0015] The fourth aspect of the present invention is to provide the application of the compounds and their derivatives described in the first aspect of the present invention in the preparation of hepatoprotective drugs.

[0016] Some compounds of the present invention and their pharmaceutically acceptable salts possess hepatoprotective effects, effectively improving cell survival rates and alleviating cell damage caused by APAP or H2O2-induced HepG2 cell injury, thus showing good potential for development into hepatoprotective drugs. Therefore, the compounds of the present invention and their pharmaceutically acceptable salts also relate to methods for treating and improving diseases related to liver injury.

[0017] Beneficial technical effects:

[0018] 1. Some of the novel watermelon compounds of the present invention have significant liver-protective effects and can significantly increase the cell survival rate of APAP or H2O2-induced HepG2 cell damage.

[0019] 2. The novel watermelon compound of this invention has a novel structure that has not been reported in the literature and has the potential to be further developed into a liver-protective drug. Detailed Implementation

[0020] The following examples and pharmacological activity experiments are used to further illustrate the present invention, but they do not imply any limitation on the present invention.

[0021] The room temperature described in the embodiments is the conventional room temperature in the art, preferably 15-30°C.

[0022] Experimental results are expressed as mean ± standard error. After parametric or nonparametric variance tests, p < 0.05 was considered statistically significant, and p < 0.01 was considered extremely statistically significant.

[0023] Example 1: Preparation and identification of the monomer Colocynthivaleric acid AU from watermelon.

[0024] 50 kg of dried watermelon fruit was extracted twice by reflux with 70% ethanol for 2 hours each time. The extract was concentrated and then extracted with petroleum ether, ethyl acetate, and n-butanol. The aqueous fraction was concentrated to obtain an extract. The extract was separated by macroporous resin column chromatography, eluting sequentially with water, 15% ethanol, 30% ethanol, 50% ethanol, and 95% ethanol to obtain five fractions. The 50% fraction (400 g) from the macroporous resin was separated by LH-20 gel column chromatography, using a gradient elution with ethanol-water as the solvent (0%-100%, v / v) to obtain 13 subfractions (AL). Fraction D (27.0 g) was separated by silica gel column chromatography, using a gradient elution with dichloromethane and methanol as solvents (0%-100%, v / v). After identification by thin-layer chromatography, the fractions were combined to obtain 11 fractions (Da-Dk). Fractions De-Df (1.50 g) were combined and then analyzed by Flash chromatography. C18 column chromatography separation, elution with a 0%-100% methanol gradient to obtain De1-De8, De3 (200 mg), followed by semi-preparative HPLC separation (35% methanol-water, 3 mL / min) to obtain compound 1 (t). R =15.8min) and compound 2 (t R =19.5min). Dk (1.86g) was separated by Flash C18 column chromatography, eluted with a 0%-100% methanol gradient to obtain Dk1-Dk10. Dk1-Dk2 were combined (220mg) and then separated by semi-preparative HPLC (32% methanol-water, 3mL / min) to obtain compound 3 (t). R =16.9min) and compound 4 (t R =20.4min); Dk6-Dk9 were combined and separated by Flash C18 column chromatography, eluted with a 0%-100% methanol gradient to obtain Dk(6-9)a-Dk(6-9)i, Dk(6-9)b-Dk(6-9)d. The combined Dk(6-9)a-Dk(6-9)i and Dk(6-9)b-Dk(6-9)d were then separated by semi-preparative HPLC (40% methanol-water, 3mL / min) to obtain compound 13 (t). R =13.1min) and compound 14 (t R =14.8min), Dk(6-9)e(250mg) was separated by semi-preparative HPLC (24% acetonitrile-water, 3mL / min) to give compound 11 (t R =10.0 min); Dk(6-9)e(200 mg) was separated by semi-preparative HPLC (26% acetonitrile-water, 3 mL / min) to obtain compound 21 (t R=11.0 min). Dc (1.30 g) was separated by Flash C18 column chromatography, eluted with a 0%-100% methanol gradient to obtain Dc1-Dc20. Dc3 (87 mg) was separated by semi-preparative HPLC (30% methanol-water, 3 mL / min) to obtain compound 5 (t). R =17.0min) and compound 6 (t R =20.9 min); Dc7 (91 mg) was separated by semi-preparative HPLC (40% methanol-water, 3 mL / min) to obtain compound 7 (t R =20.7min) and compound 8 (t R =22.5min). Dj (0.75g) was separated by Flash C18 column chromatography, eluted with a 0%-100% methanol gradient to obtain Dj1-Dj8, Dj4-Dj5 (110mg). The combined Dj1-Dj8 and Dj4-Dj5 were then separated by semi-preparative HPLC (35% methanol-water, 3mL / min) to obtain compound 9 (t). R =20.4min) and compound 10 (t R =22.5min); Dj6 was separated by semi-preparative HPLC (40% methanol-water, 3mL / min) to obtain compound 15 (t R =18.8min), compound 16 (t R =19.2min) and compound 17 (t R =19.7min); Dj7 was separated by semi-preparative HPLC (24% acetonitrile water, 3mL / min) to obtain compound 18 (t R =15.7min). Dg (1.00g) was separated by Flash C18 column chromatography, eluted with a 0%-100% methanol gradient to obtain Dg1-Dg7. Dg3 (210mg) was separated by semi-preparative HPLC (35% methanol-water, 3mL / min) to obtain compound 12 (t). R =8.5min). Dh-Di (0.77g) was combined and separated by Flash C18 column chromatography, eluted with a 0%-100% methanol gradient to obtain D(hi)1-D(hi)11, D(hi)6 (144mg). After semi-preparative HPLC separation (40% methanol-water, 3mL / min), compound 19 (t) was obtained. R =25.3min) and compound 20 (t R =26.7min).

[0025] The spectral information and NMR signal assignments of the above-mentioned new compounds are as follows:

[0026] Colocynthivaleric acid A(1):White amorphous powder;[α]20D-3(c 0.1,MeOH);UV(MeOH)λ max (logε)212(3.55),310(3.67)nm;IRν max 3390,2925,1723,1605,1514,1454,1375,1252,1161,1075,1053,853cm-1; 1 H NMR(500MHz,Methanol-d4)δ H 7.70(2H,d,J=7.0Hz,H-2' / 6'),6.97(1H,d,J=12.5Hz,H-7'),6.75(2H,d,J=8.5Hz,H-3' / 5'),5.90(1H,d,J=13.0Hz,H-8'),4.89(1H,s,H-2),4.77(1H,d,J=8.0Hz,H-1”),4.66(1H,dd,J=9.5,8.0Hz,H-2”),3.79(1H,dd,J=12.0,2.0Hz,H-6”),3.62(1H,dd,J=12.0,5.5Hz,H-6”),3.49(1H,t,J=9.5Hz,H-3”),3.34(1H,d,J=10.0Hz,H-4”),3.26(1H,m,H-5”),2.04(3H,s,3H-8”),1.38(3H,s,H-4),1.38(3H,s,H-4); 13 C NMR(125MHz,Methanol-d4)δ C 171.8(C-7”),171.6(C-1),167.7(C-9'),160.3(C-4'),146.5(C-7'),134.0(C-2' / 6'),127.5(C-1'),115.9(C-3' / 5'),115.8(C-8'),96.8(C-1”),80.1(C-2),78.4(C-3),77.8(C-5”),76.3(C-3”),75.2(C-2”),71.5(C-4”),62.6(C-6”),25.2(C-4),22.4(C-5'),21.1(C-8”).

[0027] Colocynthivaleric acid B(2):White amorphous powder;[α]20D-15(c 0.1,MeOH);UV(MeOH)λmax (logε)228(3.62),300(3.88),314(3.95)nm;IRν max 3421,2928,1732,1633,1604,1516,1444,1375,1255,1170,1073,1033,836cm-1; 1 H NMR(500MHz,Methanol-d4)δ H 7.70(1H,d,J=16.0Hz,H-7'),7.52(2H,d,J=8.5Hz,H-2' / 6'),6.82(2H,d,J=8.5Hz,H-3' / 5'),6.45(1H,d,J=15.5Hz,H-8'),4.99(1H,s,H-2),4.85(1H,d,J=8.0Hz,H-1”),4.68(1H,dd,J=10.0,8.0Hz,H-2”),3.83(1H,d,J=10.5Hz,H-6”),3.64(1H,dd,J=11.5,3.5Hz,H-6”),3.51(1H,m,H-3”),3.34(2H,m,H-5”),2.06(3H,s,H-8”),1.47(3H,s,H-5),1.43(3H,s,H-4); 13 C NMR(125 MHz,Methanol-d4)δ C 171.8(C-7”),171.6(C-1),168.7(C-9'),161.5(C-4'),147.5(C-7'),131.4(C-2' / 6'),127.1(C-1'),116.9(C-3' / 5'),114.6(C-8'),96.8(C-1”),79.9(C-2),78.0(C-5”),76.3(C-3”),75.2(C-2”),71.5(C-4”),62.7(C-6”),25.1(C-4),22.6(C-5),21.1(C-8”).

[0028] Colocynthivaleric acid C(3):White amorphous powder;[α]20 D-18(c 0.1,MeOH);UV(MeOH)λ max (logε)228(3.70),300(3.94),314(4.02)nm;IRν max3385,2925,2856,1705,1633,1605,1515,1443,1373,1348,1262,1204,1169,1074,1034,832 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H 7.69(1H,d,J=16.0 Hz,H-7'),7.50(2H,d,J=8.5 Hz,H-2' / 6'),6.82(2H,d,J=9.0 Hz,H-3' / 5'),6.44(1H,d,J=16.0 Hz,H-8'),5.10(1H,s,H-2),4.66(1H,d,J=8.0 Hz,H-1”),3.81(1H,dd,J=12.0,2.5 Hz,H-6”),3.60(1H,dd,J=12.0,6.0 Hz,H-6”),3.39(1H,t,J=9.0 Hz,H-3”),3.29(1H,dd,J=5.5,2.0 Hz,H-5”),3.25(1H,m,H-4”),3.19(1H,m,H2”),1.50(3H,s,H-5),1.44(3H,s,H-4); 13 C NMR(125 MHz,Methanol-d4)δ C 172.4(C-1),168.7(C-9'),161.5(C-4'),147.6(C-7'),131.4(C-2' / 6'),127.1(C-1'),116.8(C-3' / 5'),114.4(C-8'),98.8(C-1”),78.7(C-2),78.4(C-3),78.0(C-3”),77.9(C-5”),75.1(C-2”),71.5(C-4”),62.9(C-6”),24.6(C-4),23.4(C-5).

[0029] Colocynthivaleric acid D(4):White amorphous powder;[α]20 D-31(c 0.1,MeOH);UV(MeOH)λ max (logε)227(3.81),299(3.94),311(4.00)nm;IRν max 3375,2929,1716,1605,1514,1455,1390,1236,1190,1156,1075,1044,1018,856 cm -1 ;1 H NMR(500 MHz,Methanol-d4)δ H 7.68(2H,d,J=9.0 Hz,H-2' / 6'),6.95(1H,d,J=13.0 Hz,H-7'),6.75(2H,d,J=8.5 Hz,H-3' / 5'),5.88(1H,d,J=12.5 Hz,H-8'),5.01(1H,s,H-2),4.63(1H,d,J=7.5 Hz,H-1”),3.79(1H,dd,J=12.0,2.0 Hz,H-6”),3.60(1H,m,H-6”),3.39(1H,m,H-3”),3.26(2H,m,H-4” / 5”),3.19(1H,dd,J=8.0,1.5 Hz,H-2”),1.41(3H,s,H-4),1.38(3H,s,H-5); 13 C NMR(125 MHz,Methanol-d4)δ C 172.4(C-1),167.7(C-9'),160.3(C-4'),146.4(C-7'),133.9(C-2' / 6'),127.5(C-1'),115.9(C-3' / 5'),115.7(C-8'),98.8(C-1”),78.8(C-2),78.3(C-3),77.9(C-3”),77.8(C-5”),75.1(C-2”),71.5(C-4”),62.8(C-6”),24.7(C-5),23.3(C-4).

[0030] Colocynthivaleric acid E(5):White amorphous powder;[α]20 D-17(c 0.1,MeOH);UV(MeOH)λ max (logε)229(3.75),301(4.01),314(4.09)nm;IRν max 3423,2931,1716,1632,1604,1515,1441,1374,1259,1204,1169,1076,1042,983,832cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H7.69(1H,d,J=15.5 Hz,H-7'),7.51(2H,d,J=8.5 Hz,H-2' / 6'),6.82(2H,d,J=9.0 Hz,H-3' / 5'),6.44(1H,d,J=16.0 Hz,H-8'),5.10(1H,s,H-2),4.97(1H,t,J=10.0 Hz,H-3”),4.75(1H,d,J=8.0 Hz,H-1”),3.81(1H,dd,J=12.0,2.0 Hz,H-6”),3.63(1H,dd,J=12.0,5.5 Hz,H-6”),3.42(1H,m,H-4”),3.38(1H,m,H-5”),3.31(1H,m,H-2”),2.10(3H,s,H-8”),1.50(3H,s,H-4),1.45(3H,s,H-5); 13 C NMR(125 MHz,Methanol-d4)δ C 172.7(C-7”),172.3(C-1),168.7(C-9'),161.5(C-4'),147.6(C-7'),131.4(C-2 / 6'),127.1(C-1'),116.8(C-3' / 5'),114.5(C-8'),98.7(C-1”),78.9(C-3”),78.8(C-2),78.6(C-3),77.6(C-5”),73.3(C-2”),69.6(C-4”),62.6(C-6”),24.5(C-5),23.5(C-4),21.1(C-8”).

[0031] Colocynthivaleric acid F(6):White amorphous powder;[α]20 D-22(c 0.1,MeOH);UV(MeOH)λ max (logε)212(4.84),301(4.95)nm;IRν max 3407,2925,1722,1605,1514,1448,1374,1258,1158,1079,1042,857,841 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H7.68(2H,d,J=9.0 Hz,H-2” / 6”),6.94(1H,d,J=12.5 Hz,H-7'),6.75(2H,d,J=9 Hz,H-3' / 5'),5.88(1H,d,J=12.5 Hz,H-8'),5.00(1H,s,H-2),4.96(1H,t,J=9.5 Hz,H-3”),4.72(1H,d,J=7.5 Hz,H-1”),3.78(1H,dd,J=12.0,2.0 Hz,H-6”),3.62(1H,dd,J=12.0,5.5Hz,H-6”),3.43(1H,t,J=10.0 Hz,H-4”),3.35(1H,m,H-2”),2.10(3H,s,H-8”),1.41(3H,s,H-4),1.38(3H,s,H-5); 13 C NMR(125 MHz,Methanol-d4)δ C 172.7(C-1 / 7”),167.7(C-9'),160.2(C-4'),146.2(C-7'),133.9(C-2' / 6'),127.5(C-1'),115.9(C-3' / 5' / 8'),98.7(C-1”),79.0(C-2),78.9(C-3”),78.5(C-3),77.5(C-5”),73.3(C-2”),69.6(C-4”),62.5(C-6”),24.7(C-5),23.3(C-4),21.1(C-8”).

[0032] Colocynthivaleric acid G(7):White amorphous powder;[α]20 D-17(c 0.1,MeOH);UV(MeOH)λ max (logε)227(3.83),300(3.98)nm,312(4.03);IRν max 3377,2926,2858,1728,1605,1514,1456,1378,1254,1162,1075,1050,908,862,841 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H7.69(2H,d,J=8.5 Hz,H-2' / 6'),6.95(1H,d,J=13.0 Hz,H-7'),6.75(2H,d,J=9.0 Hz,H-3' / 5'),5.90(1H,d,J=12.5 Hz,H-8'),5.14(1H,s,H-2),4.77(1H,d,J=8.0 Hz,H-1”),4.71(1H,dd,J=9.5,8.0 Hz,H-2”),3.75(1H,d,J=12.0 Hz,H-6”),3.53(2H,m,H-3” / 6”),3.29(2H,m,H-4” / 5”),2.08(3H,s,H-8”),1.90(1H,q,J=7.5 Hz,H-4),1.54(1H,q,J=7.0 Hz,H-4),1.38(3H,s,H-6),0.87(3H,t,J=7.5 Hz,H-5); 13 C NMR(125 MHz,Methanol-d4)δ C 172.2(C-1),171.8(C-7”),168.1(C-9'),160.2(C-4'),146.4(C-7'),134.0(C-2' / 6'),127.5(C-1'),116.1(C-3' / 5'),115.9(C-8'),96.5(C-1”),80.5(C-3),78.8(C-2),78.1(C-5”),76.2(C-3”),75.3(C-2”),71.6(C-4”),62.9(C-6”),29.8(C-4),21.2(C-8”),20.3(C-6),7.7(C-5).

[0033] Colocynthivaleric acid H(8):White amorphous powder;[α]20 D-41(c 0.1,MeOH);UV(MeOH)λ max (logε)226(3.77),300(4.00),314(4.07)nm;IRν max 3391,2927,1732,1632,1604,1516,1446,1377,1351,1256,1204,1170,1076,1039,834cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H7.70(1H,d,J=16.0 Hz,H-7'),7.52(2H,d,J=8.5 Hz,H-2' / 6'),6.82(2H,d,J=8.5 Hz,H-3' / 5'),6.46(1H,d,J=16.0 Hz,H-8'),5.23(1H,s,H-2),4.80(1H,d,J=8.0 Hz,H-1”),4.70(1H,dd,J=9.5,7.5 Hz,H-2”),3.79(1H,dd,J=12.0,2.0 Hz,H-6”),3.57(1H,dd,J=12.0,6.0 Hz,H-6”),3.52(1H,t,J=9.0 Hz,H-3”),3.34(1H,m,H-5”),3.28(1H,m,H-4”),2.09(3H,s,H-8”),2.03(1H,m,H-4),1.61(1H,q,J=7.5 Hz,H-4),1.43(3H,s,H-6),0.92(3H,t,J=7.5 Hz,H-5); 13 C NMR(125 MHz,Methanol-d4)δ C 172.1(C-1),171.7(C-7”),169.3(C-9'),161.5(C-4'),147.5(C-7'),131.4(C-2' / 6'),127.1(C-1'),116.8(C-3' / 5'),114.8(C-8'),96.5(C-1”),80.5(C-3),78.6(C-2),78.1(C-5”),76.1(C-3”),75.2(C-2”),71.5(C-4”),63.0(C-6”),30.0(C-4),21.1(C-8”),20.3(C-6),7.8(C-5).

[0034] Colocynthivaleric acid I(9):White amorphous powder;[α]20 D-12(c 0.1,MeOH);UV(MeOH)λ max (logε)226(3.37),300(3.57),313(3.64)nm;IRν max 3377,2925,2856,1705,1632,1604,1515,1445,1381,1349,1261,1204,1170,1076,1034,832,801 cm -1 ; 1 HNMR(500 MHz,Methanol-d4)δH 7.69(1H,d,J=16.0 Hz,H-7'),7.50(2H,d,J=8.5 Hz,H-2' / 6'),6.82(2H,d,J=9.0 Hz,H-3' / 5'),6.43(1H,d,J=16.0 Hz,H-8'),5.24(1H,s,H-2),4.60(1H,d,J=7.5 Hz,H-1”),3.80(1H,dd,J=12.0,2.5 Hz,H-6”),3.58(1H,dd,J=12.0,6.0 Hz,H-6”),3.36(1H,t,J=8.5 Hz,H-3”),3.27(1H,m,H-5”),3.23(1H,m,H-4”),3.18(1H,dd,J=9.0,7.5 Hz,H-2”),1.98(1H,q,J=7.5 Hz,H-4),1.76(1H,q,J=8.0 Hz,H-4),1.45(3H,s,H-6),1.02(3H,t,J=7.5 Hz,H-5); 13 C NMR(125 MHz,Methanol-d4)δ C 172.5(C-1),168.9(C-9'),161.5(C-4'),147.6(C-7'),131.4(C-2' / 6'),127.1(C-1'),116.9(C-3' / 5'),114.5(C-8'),98.6(C-1”),80.5(C-3),78.1(C-3”),78.0(C-2),77.9(C-5”),75.2(C-2”),71.5(C-4”),63.0(C-6”),30.1(C-4),20.8(C-6),8.0(C-5).

[0035] Colocynthivaleric acid J(10):White amorphous powder;[α]20 D-23(c 0.1,MeOH);UV(MeOH)λ max (logε)226(3.84),311(4.01)nm;IRν max 3378,2929,1716,1604,1514,1456,1384,1260,1157,1077,1033,859,839 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H7.68(2H,d,J=9.0 Hz,H-2' / 6'),6.94(1H,d,J=13.0 Hz,H-7'),6.75(2H,d,J=8.5 Hz,H-3' / 5'),5.88(1H,d,J=12.5 Hz,H-8'),5.15(1H,s,H-2),4.57(1H,d,J=7.5 Hz,H-1”),3.77(1H,dd,J=12.0,2.0 Hz,H-6”),3.56(1H,m,H-6”),3.36(1H,t,J=9.0 Hz,H-3”),3.25(2H,m,H-4” / 5”),3.18(1H,dd,J=9.5,8.0 Hz,H-2”),1.85(1H,q,J=7.5 Hz,H-4),1.69(1H,q,J=7.5 Hz,H-4),1.39(3H,s,H-6),0.97(3H,t,J=7.5 Hz,H-5); 13 C NMR(125 MHz,Methanol-d4)δ C 172.8(C-1),167.8(C-9'),160.3(C-4'),146.4(C-7'),134.0(C-2' / 6'),127.5(C-1'),115.9(C-8'),115.8(C-3' / 5'),98.6(C-1”),80.5(C-3),78.1(C-3”),77.8(C-5”),75.2(C-2”),71.6(C-4”),62.9(C-6”),29.8(C-4),20.9(C-6),8.0(C-5).

[0036] Colocynthivaleric acid K(11):White amorphous powder;[α]20 D-8(c 0.1,MeOH);UV(MeOH)λ max (logε)227(3.78),299(3.97),316(4.08)nm;IRν max 3386,2923,1716,1632,1604,1515,1441,1373,1262,1162,1074,1033,834 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H7.69(1H,d,J=16.0 Hz,H-7'),7.51(2H,d,J=9.0 Hz,H-2' / 6'),6.82(2H,d,J=8.5Hz,H-3' / 5'),6.43(1H,d,J=16.0 Hz,H-8'),5.13(1H,s,H-2),4.63(1H,d,J=8.0 Hz,H-1'),3.80(1H,dd,J=12.0,2.0 Hz,H-6”),3.76(3H,s,H-OCH3),3.60(1H,dd,J=12.0,5.5Hz,H-6”),3.37(1H,t,J=8.5 Hz,H-3”),3.27(1H,m,H-5”),3.24(1H,m,H-4”),3.16(1H,dd,J=9.0,7.5 Hz,H-2”),1.48(3H,s,H-5),1.42(3H,s,H-4); 13 C NMR(125 MHz,Methanol-d4)δ C 171.0(C-1),168.7(C-9'),161.7(C-4'),147.8(C-7'),131.5(C-2' / 6'),127.0(C-1'),116.9(C-3' / 5'),114.1(C-8'),98.9(C-1”),79.1(C-9'),78.4(C-3),78.0(C-3”),77.9(C-5”),75.0(C-2”),71.5(C-4”),62.9(C-6”),52.8(C-OCH3),24.3(C-4),23.4(C-5).

[0037] Colocynthivaleric acid L(12):White amorphous powder;[α]20 D-25(c 0.1,MeOH);UV(MeOH)λ max (logε)211(3.87),228(3.92),314(4.21)nm;IRν max 3400,2986,1716,1631,1605,1588,1516,1444,1373,1263,1169,1075,1043,830 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H7.63(1H,d,J=16.0 Hz,H-7),7.49(2H,d,J=9.0Hz,H-2 / 6),6.81(2H,d,J=8.5 Hz,H-3 / 5),6.42(1H,d,J=16.0 Hz,H-8),5.06(1H,s,H-2'),4.65(1H,d,J=8.0Hz,H-1”),4.25(1H,dd,J=11.5,2.0 Hz,H-6”),4.12(1H,dd,J=11.5,5.5 Hz,H-6”),3.47(1H,m,H-5”),3.39(1H,t,J=9.0 Hz,H-3”),3.27(1H,t,J=9.0 Hz,H-4”),3.20(1H,dd,J=9.5,8.0 Hz,H-2”),1.91(3H,s,H-8”),1.46(3H,s,H-5'),1.43(3H,s,H-4'); 13 C NMR(125MHz,Methanol-d4)δ C 172.9(C-1' / 7”),168.4(C-9),161.4(C-4),147.1(C-7),131.3(C-2 / 6),127.1(C-1),116.8(C-3 / 9),114.7(C-8),98.8(C-1”),78.8(C-2'),78.4(C-3'),77.8(C-3”),75.0(C-2”),74.9(C-5”),71.3(C-4”),64.8(C-6”),24.5(C-4'),23.4(C-5'),20.7(C-8”).

[0038] Colocynthivaleric acid M(13):White amorphous powder;[α]20 D-15(c 0.1,MeOH);UV(MeOH)λ max (logε)227(4.24),314(4.15)nm;IRν max 3377,2927,1721,1606,1514,1455,1371,1233,1151,1075,1044,1018,855,830 cm -1 ; 1 H NMR(500MHz,DMSO-d6)δ H7.67(2H,d,J=8.5 Hz,H-2' / 6'),7.32(2H,d,J=9.0 Hz,H-3” / 5”),6.99(2H,d,J=9.0 Hz,H-2” / 6”),6.95(1H,d,J=13.0 Hz,H-7'),6.75(2H,d,J=9.0Hz,H-3' / 5'),5.85(1H,d,J=13.0 Hz,H-8'),5.15(1H,d,J=12.0 Hz,H-7”),5.07(1H,d,J=12.0 Hz,H-7”),4.95(1H,s,H-2),4.87(1H,d,J=7.5 Hz,H-1””),4.45(1H,d,J=8.0 Hz,H-1”'),3.67(1H,d,J=12.0 Hz,H-6””),3.58(1H,d,J=12.0 Hz,H-6”'),3.46(1H,dd,J=12.0,5.5 Hz,H-6””),3.35(1H,m,H-6”'),3.32(1H,m,H-5””),3.24(1H,m,H-H-2”” / 3””),3.15(1H,m,H-3”' / 4””),3.09(1H,m,H-5”'),3.03(1H,m,H-4”'),2.89(1H,td,J=9.0,4.5 Hz,H-2”'),1.26(1H,s,H-4),1.23(1H,s,H-5); 13 C NMR(125MHz,DMSO-d6)δ C 168.3(C-1),165.6(C-9'),159.1(C-4'),157.4(C-1”),145.0(C-7'),132.9(C-2' / 6'),130.0(C-3” / 5”),128.7(C-4”),125.3(C-1'),116.0(C-2” / 6”),115.0(C-3' / 5'),114.3(C-8'),100.2(C-1””),97.4(C-1”'),77.9(C-2),77.0(C-5””),76.7(C-3””),76.6(C-3”'),76.5(C-5”'),76.3(C-3),73.4(C-2”'),73.2(C-2””),70.1(C-4”'),69.6(C-4””),66.2(C-7”),61.2(C-6”'),60.6(C-6””),23.8(C-5),22.5(C-4).

[0039] Colocynthivaleric acid N(14):White amorphous powder;[α]20 D-25(c 0.1,MeOH);UV(MeOH)λ max (logε)227(4.04),301(4.05),316(4.14)nm;IRν max 3377,2921,1715,1631,1605,1515,1444,1373,1352,1235,1162,1075,1044,832 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H 7.68(1H,d,J=15.5 Hz,H-7'),7.50(2H,d,J=9.0Hz,H-2' / 6'),7.34(2H,d,J=9.0 Hz,H-3” / 5”),7.08(2H,d,J=9.0 Hz,H-2” / 6”),6.82(2H,d,J=8.5 Hz,H-3' / 5'),6.42(1H,d,J=15.5 Hz,H-8'),5.21(1H,d,J=12.0 Hz,H-7”),5.12(1H,s,H-2),5.11(1H,d,J=13.5 Hz,H-7”),4.90(1H,d,H-1””),4.59(1H,d,J=8.0 Hz,H-1”'),3.89(1H,dd,J=12.0,2.0 Hz,H-6””),3.79(1H,dd,J=12.0,2.0Hz,H-6”'),3.70(1H,dd,J=12.0,5.5 Hz,H-6””),3.58(1H,dd,J=12.0,5.5 Hz,H-6”'),3.45(3H,m,H-2”' / 3”” / 5””),3.39(1H,m,H-5”'),3.36(1H,m,H-4”'),3.25(2H,m,H-3”' / 4””),3.13(1H,dd,J=9.0,7.5 Hz,H-2””),1.43(3H,s,H-4),1.36(3H,s,H-5); 13 C NMR(125 MHz,Methanol-d4)δ C170.3(C-1),168.8(C-9'),161.9(C-4'),159.3(C-1”),148.0(C-7'),131.5(C-2' / 6'),131.3(C-3” / 5”),130.6(C-4”),126.8(C-1'),117.6(C-2” / 6”),117.0(C-3' / 5'),114.0(C-8'),102.1(C-1””),98.8(C-1”'),79.2(C-2),78.1(C-5”'),77.9(C-3”” / 5””),77.8(C-3”'),75.0(C-2””),74.9(C-2”'),71.5(C-4””),71.3(C-4”'),67.9(C-7”),62.9(C-6”'),62.4(C-6””),24.3(C-4),23.5(C-5).

[0040] Colocynthivaleric acid O(15):White amorphous powder;[α]20 D-29(c 0.1,MeOH);UV(MeOH)λ max (logε)226(4.05),301(4.02),316(4.11)nm;IRν max 3390,2923,2850,1716,1631,1604,1515,1446,1373,1261,1236,1162,1075,1042,832,802 cm -1 ; 1 H NMR(500MHz,Methanol-d4)δ H7.63(1H,d,J=16.0 Hz,H-7'),7.49(2H,d,J=9.0 Hz,H-2' / 6'),7.34(2H,d,J=8.5 Hz,H-3” / 5”),7.08(2H,d,J=8.5 Hz,H-2” / 6”),6.82(2H,d,J=9.0Hz,H-3' / 5'),6.40(1H,d,J=16.0 Hz,H-8'),5.18(1H,d,J=12.0 Hz,H-7”),5.11(1H,d,J=12.0 Hz,H-7”),5.08(1H,s,H-2),4.90(1H,d,H-1””),4.58(1H,d,J=7.5 Hz,H-1”'),4.22(1H,dd,J=12.0,2.5 Hz,H-6”'),4.09(1H,dd,J=11.5,6.0 Hz,H-6”'),3.89(1H,dd,J=12.0,2.0 Hz,H-6””),3.70(1H,dd,J=12.0,5.5 Hz,H-6””),3.45(4H,m,H-2”' / 5”' / 3”” / 5””),3.40(1H,m,H-4””),3.35(1H,t,J=9.0 Hz,H-3”'),3.26(1H,t,J=9.5 Hz,H-4”'),3.15(1H,dd,J=9.0,7.5 Hz,H-2””),1.87(3H,s,H-8”'),1.39(3H,s,H-4),1.35(3H,s,H-5); 13 C NMR(125 MHz,Methanol-d4)δ C 172.9(C-7”'),170.3(C-1),168.4(C-9'),161.5(C-4'),159.3(C-1”),147.5(C-7'),131.4(C-2' / 6'),130.7(C-4”),127.0(C-1'),117.6(C-2” / 6”),116.9(C-3' / 5'),114.3(C-8'),102.2(C-1””),98.7(C-1”'),79.0(C-2),78.4(C-3),78.1(C-5””),77.9(C-3””),77.8(C-3”'),74.9(C-2”'),74.8(C-5”'),71.4(C-4”'),71.3(C-4””),67.9(C-7”),64.8(C-6”'),62.4(C-6””),23.8(C-5),23.6(C-4),20.7(C-8”').

[0041] Colocynthivaleric acid P(16):White amorphous powder;[α]20 D 8(c 0.1,MeOH);UV(MeOH)λ max (logε)227(4.18),315(4.08)nm;IRν max 3378,2925,2854,1732,1606,1515,1457,1375,1236,1152,1074,1047,862,830 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H 7.67(2H,d,J=9.0 Hz,H-2' / 6'),7.32(2H,d,J=8.5 Hz,H-3” / 5”),7.04(2H,d,J=8.5Hz,H-2” / 6”),6.96(1H,d,J=13.0 Hz,H-7'),6.75(2H,d,J=8.5 Hz,H-3' / 5'),5.85(1H,d,J=13.0 Hz,H-8'),5.17(1H,d,J=12.0 Hz,H-7”),5.10(1H,d,J=12.0 Hz,H-7”),4.89(d,1H),4.88(1H,s,H-2),4.70(1H,d,J=8.0 Hz,H-1”'),4.64(1H,dd,J=9.5,8.0 Hz,H-2”'),3.88(1H,dd,J=12.0,2.0 Hz,H-6””),3.78(1H,dd,J=12.0,2.5 Hz,H-6”'),3.70(1H,dd,J=12.0,5.0 Hz,H-6””),3.61(1H,dd,J=12.0,5.5Hz,H-3”'),3.49(1H,d,J=9.5Hz,H-3”'),3.46(1H,m,H-2””),3.45(1H,m,H-5””),3.43(1H,m,H-3””),3.39(1H,m,H-4””),3.35(1H,d,J=9.5 Hz,H-4”'),3.23(1H,m,H-5”'),2.04(3H,s,H-8”'),1.33(3H,s,H-4),1.30(3H,s,H-5); 13 C NMR(125MHz,Methanol-d4)δ C171.8(C-7”'),169.6(C-1'),167.6(C-9'),160.4(C-4'),159.2(C-1”),147.0(C-7'),134.1(C-2' / 6'),131.3(C-3” / 5”),130.7(C-4”),127.4(C-1'),117.6(C-2” / 6”),116.0(C-3' / 5'),115.4(C-8'),102.1(C-1””),96.7(C-1”'),80.2(C-2),78.6(C-3),78.1(C-5””),77.9(C-5”'),77.8(C-3””),76.2(C-3”'),75.1(C-2”'),74.9(C-2””),71.4(C-4”'),71.3(C-4””),67.9(C-7”),62.6(C-6”'),62.4(C-6””),24.8(C-5),22.6(C-4),21.2(C-8”').

[0042] Colocynthivaleric acid Q(17):White amorphous powder;[α]20 D-16(c 0.1,MeOH);UV(MeOH)λ max (logε)226(4.13),276(3.70),313(4.00)nm;IRν max 3376,2925,2856,1722,1606,1514,1373,1236,1151,1075,1043,830 cm -1 ; 1 H NMR(500MHz,Methanol-d4)δ H7.64(2H,d,J=8.5 Hz,H-2' / 5'),7.34(2H,d,J=8.5 Hz,H-3” / 5”),7.06(2H,d,J=8.5Hz,H-2” / 6”),6.93(1H,d,J=13.0 Hz,H-7'),6.74(2H,d,J=9.0 Hz,H-3' / 5'),5.82(1H,d,J=13.0 Hz,H-8'),5.19(1H,d,J=12.0 Hz,H-7”),5.11(1H,d,J=12.0 Hz,H-7”),5.04(1H,s,H-2),4.90(1H,overlapped,H-1””),4.55(1H,d,J=8.0 Hz,H-1”'),4.26(1H,dd,J=12.0,2.5 Hz,H-6”'),4.08(1H,dd,J=11.5,6.5 Hz,H-6”'),3.89(1H,dd,J=12.5,2.5Hz,H-6””),3.70(1H,dd,J=12.0,5.5 Hz,H-6””),3.47(1H,m,H-3””),3.45(2H,m,H-5”' / 5””),3.41(2H,m,H-2”” / H-4””),3.35(1H,t,J=9.0 Hz,H-3”'),3.23(1H,m,H-4”'),3.15(1H,dd,J=9.5,8.0 Hz,H-2”'),1.95(3H,s,H-8”'),1.29(3H,s,H-4),1.28(3H,s,H-5); 13 CNMR(125 MHz,Methanol-d4)δ C172.9(C-7”'),170.4(C-1),167.4(C-9'),160.3(C-4'),159.2(C-1”),146.5(C-7'),134.0(C-2' / 6'),131.4(C-3” / 5”),130.6(C-4”),127.4(C-1'),117.6(C-2” / 6”),115.9(C-3' / 5'),115.5(C-8'),102.1(C-1””),98.8(C-1”'),79.3(C-2),78.3(C-3),78.1(C-5””),77.9(C-3””),77.8(C-3”' / 3””),74.9(C-5”' / 2””),,74.8(C-2””),71.6(C-4”'),71.3(C-4””),67.9(C-7”),65.0(C-6”'),62.4(C-6””),23.9(C-5),23.5(C-4),20.8(C-8”').

[0043] Colocynthivaleric acid R(18):White amorphous powder;[α]20 D-58(c 0.1,MeOH);UV(MeOH)λ max (logε)227(4.21),302(4.19),316(4.28)nm;IRν max 3377,2925,1733,1604,1515,1382,1235,1166,1074,1044,834 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H7.69(1H,d,J=16.0 Hz,H-7'),7.51(2H,d,J=9.0 Hz,H-2' / 6'),7.34(2H,d,J=8.5 Hz,H-3” / 5”),7.08(2H,d,J=8.5 Hz,H-2” / 6”),6.82(2H,d,J=9.0 Hz,H-3' / 5'),6.43(1H,d,J=16.0 Hz,H-8'),5.21(1H,s,H-2),5.18(1H,d,J=12.0 Hz,H-7”),5.09(1H,d,J=12.0 Hz,H-7”),4.90(1H,overlapped,H-1””),4.75(1H,d,J=8.0 Hz,H-1”'),4.69(1H,t,J=7.5Hz,H-2”'),3.89(1H,dd,J=12.0,2.0 Hz,H-6””),3.77(1H,dd,J=12.0,2.0 Hz,H-6”'),3.70(1H,dd,J=12.0,5.5 Hz,H-6””),3.55(1H,dd,J=12.0,5.5 Hz,H-6”'),3.50(1H,m,H-3”'),3.46(2H,m,H-2”” / 5””),3.43(1H,m,H-3””),3.40(1H,m,H-4””),3.29(2H,m,H-4”' / 5”'),2.07(3H,s,H-8”),1.95(1H,q,J=7.0 Hz,H-5),1.57(1H,q,J=7.0 Hz,H-5),1.36(3H,s,H-4),0.89(3H,t,J=7.5 Hz,H-6); 13 C NMR(125 MHz,Methanol-d4)δ C171.7(C-7”'),169.9(C-1),169.0(C-9'),161.6(C-4'),159.2(C-1”),147.9(C-7'),131.5(C-2' / 6'),131.2(C-3” / 5”),130.7(C-4”),127.0(C-1'),117.6(C-2” / 6”),116.9(C-3' / 5'),114.3(C-8'),102.2(C-1””),96.4(C-1”'),80.8(C-3),78.1(C-5””),77.9(C-5”' / 3””),76.1(C-3”'),75.2(C-2”'),74.9(C-2””),71.5(C-4”'),71.3(C-4””),67.9(C-7”),62.9(C-6”'),62.4(C-6””),30.1(C-5),21.2(C-8”'),20.3(C-4),7.9(C-6).

[0044] Colocynthivaleric acid S(19):White amorphous powder;[α]20 D-36(c 0.1,MeOH);UV(MeOH)λ max (logε)226(4.16),300(4.12),316(4.22)nm;IRν max 3396,2925,1716,1631,1604,1515,1442,1374,1238,1167,1076,1043,834 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H7.68(1H,d,J=16.0 Hz,H-7'),7.50(2H,d,J=9.0 Hz,H-2' / 6'),7.34(2H,d,J=8.5Hz,H-3” / 5”),7.08(2H,d,J=9.0 Hz,H-2” / 6”),6.82(2H,d,J=8.5 Hz,H-3' / 5'),6.43(1H,d,J=15.5 Hz,H-8'),5.22(1H,d,J=12.0 Hz,H-7”),5.11(1H,s,H-2),5.10(1H,d,J=12.0 Hz,H-7”),4.95(1H,d,J=9.0 Hz,H-3”'),4.66(1H,d,J=7.5 Hz,H-1”'),3.89(1H,dd,J=12.0,2.0 Hz,H-6””),3.79(1H,dd,J=12.0,2.5 Hz,H-6”'),3.70(1H,dd,J=12.0,5.5 Hz,H-6””),3.61(1H,dd,J=12.0,5.5 Hz,H-6”'),3.45(3H,m,H-2”” / 4”” / 5””),3.41(2H,m,H-4”' / 3””),3.35(1H,m,H-5”'),3.25(1H,dd,J=9.5,7.5 Hz,H-2”'),2.10(3H,s,H-8”'),1.43(3H,s,H-5),1.38(3H,s,H-4); 13 C NMR(125 MHz,Methanol-d4)δ C 172.7(C-7”'),170.3(C-1),168.7(C-9'),161.6(C-4'),159.3(C-1”),147.9(C-7'),131.5(C-2' / 6'),131.4(C-3” / 5”),130.6(C-4”),127.0(C-1'),117.6(C-2” / 6”),116.9(C-3' / 5'),114.2(C-8'),102.1(C-1””),98.7(C-1”'),79.2(C-2),78.9(C-3),78.8(C-3”'),78.1(C-5””),77.9(C-3””),77.5(C-5”'),74.9(C-2””),73.3(C-2”'),71.3(C-4””),69.6(C-4”'),67.9(C-7”),62.6(C-6”'),62.4(C-6””),24.3(C-4),23.5(C-5),21.2(C-8”').

[0045] Colocynthivaleric acid T(20):White amorphous powder;[α]20 D-6(c 0.1,MeOH);UV(MeOH)λ max (logε)226(3.96),276(3.51),314(3.84)nm;IRν max 3377,2923,2856,1723,1606,1514,1374,1260,1237,1151,1075,1043,802 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H 7.65(2H,d,J=8.5 Hz,H-2' / 6'),7.34(2H,d,J=8.5 Hz,H-3” / 5”),7.06(2H,d,J=9.0 Hz,H-2” / 6”),6.96(1H,d,J=13.0 Hz,H-7'),6.74(2H,d,J=8.5 Hz,H-3' / 5'),5.85(1H,d,J=12.5 Hz,H-8'),5.22(1H,d,J=12.0 Hz,H-7”),5.11(1H,d,J=12.0 Hz,H-7”),5.02(1H,s,H-2),4.93(1H,overlapped,J=9.5Hz,H-3”'),4.90(1H,overlapped,H-1””),4.61(1H,d,J=7.5 Hz,H-1”'),3.89(1H,dd,J=12.0,2.0 Hz,H-6””),3.76(1H,dd,J=12.0,2.5 Hz,H-6”'),3.70(1H,dd,J=12.0,5.0Hz,H-6””),3.60(1H,dd,J=12.0,5.0 Hz,H-6”'),3.45(3H,m,H-2”” / 3”” / 5””),3.40(2H,m,H-4”' / 4””),3.28(1H,m,H-5”'),3.24(1H,dd,J=9.5,7.5 Hz,H-2”'),2.11(3H,s,H-8”'),1.34(3H,s,H-4),1.32(3H,s,H-5); 13 CNMR(125 MHz,Methanol-d4)δ C172.7(C-7”'),170.3(C-1),167.7(C-9'),160.3(C-4'),159.3(C-1”),146.8(C-7'),134.0(C-2' / 6')),131.4(C-3” / 5”),130.6(C-4”),127.5(C-1'),117.6(C-2” / 6”),115.9(C-3' / 5'),115.4(C-8'),102.1(C-1””),98.7(C-1”'),79.4(C-2),78.9(C-3”'),78.6(C-3),78.1(C-2””),77.9(C-5””),77.5(C-5”'),74.9(C-3””),73.3(C-2”'),71.3(C-4””),69.6(C-4”'),68.0(C-7”),62.5(C-6”'),62.4(C-6””),24.4(C-5),23.3(C-4),21.2(C-8”').

[0046] Colocynthivaleric acid U(21):White amorphous powder;[α]20 D-49(c 0.1,MeOH);UV(MeOH)λ max (logε)226(4.16),301(4.12),316(4.22)nm;IRν max 3384,2925,1718,1631,1604,1515,1459,1380,1350,1233,1168,1076,1043,833 cm -1 ; 1 H NMR(500 MHz,Methanol-d4)δ H7.68(1H,d,J=16.0 Hz,H-7”),7.50(2H,d,J=8.5Hz,H-2' / 6'),7.35(2H,d,J=9.0 Hz,H-3” / 5”),7.09(2H,d,J=9.0 Hz,H-2” / 6”),6.82(2H,d,J=9.0 Hz,H-3” / 5”),6.42(1H,d,J=16.0 Hz,H-8”),5.24(1H,s,H-2),5.22(1H,d,J=12.0 Hz,H-7”),5.08(1H,d,J=12.0 Hz,H-7”),4.90(1H,overlapped,H-1””),4.53(,1H,d,J=7.5 Hz,H-1”'),3.90(1H,dd,J=12.0,2.0 Hz,H-6””),3.78(1H,dd,J=12.0,2.0 Hz,H-6”'),3.70(1H,dd,J=12.0,5.5 Hz,H-6”'),3.56(1H,dd,J=12.0,5.5 Hz,H-6”'),3.45(3H,m,H-5”' / 2”” / 3””),3.39(1H,m,H-4””),3.35(1H,m,H-3”'),3.24(2H,m,H-4”' / 5””),3.13(1H,m,H-2”'),1.92(1H,q,J=7.5 Hz,H-5),1.69(1H,dd,J=7.5Hz,H-5),1.39(3H,s,H-4),0.97(3H,d,J=7.5Hz,H-6); 13 C NMR(125MHz,Methanol-d4)δ C 170.5(C-1),168.8(C-9'),161.6(C-4'),159.3(C-1”),147.9(C-7'),131.5(C-2' / 6'),131.3(C-3” / 5”),130.7(C-4”),127.0(C-1'),117.6(C-2” / 6”),116.9(C-3' / 5'),114.2(C-8'),102.2(C-1””),98.6(C-1”'),80.6(C-3),78.1(C-3”' / 5”'),77.9(C-3””),77.8(C-5””),77.7(C-2),75.1(C-2”'),74.9(C-2””),71.6(C-4”'),71.3(C-4””),67.9(C-7”),63.0(C-6”'),62.5(C-6””),30.3(C-5),20.7(C-4),7.9(C-6).

[0047] Pharmacological experiments

[0048] Example 1: Protective effect of Colocynthivaleric acid AU from watermelon on hepatocyte damage in human HepG2 liver cancer cells.

[0049] The in vitro liver protection models for the test compounds were established using APAP and H2O2 models, with human hepatocellular carcinoma HepG2 cells as the experimental cells. Cells were grown in DMEM medium (containing 100 U / mL penicillin and 100 μg / mL streptomycin) with 10% fetal bovine serum at 37°C, 5% CO2, and saturated humidity. Cells were passaged using digestion with 0.25% trypsin and 0.02% EDTA.

[0050] In the APAP model, the MTT assay was used. HepG2 cells were seeded in 96-well cell culture plates and cultured for 24 h. Then, 10 μM of the test compound and 8 mM APAP were added. A positive control group (bicyclool), a solvent blank control group, and a model group were also included. Cells were cultured for another 24 h. The culture medium was discarded, and 100 μL of MTT (0.5 mg / mL) solution was added to each well. Cells were cultured for another 4 h, and the MTT solution was discarded. 150 μL of DMSO was added to each well, and the mixture was shaken. The absorbance was measured at 570 nm using a microplate reader. Cell viability (%) = (average OD of the drug group / average OD of the solvent control group) × 100%.

[0051] In the H2O2 model, HepG2 cells were seeded in 96-well cell culture plates and cultured for 24 h. Then, 10 μM of the test compound was added. A positive control group (bicyclool), a solvent blank control group, and a model group were also included. After 12 h of pre-incubation with the drug, except for the solvent control group which received an equal volume of culture medium, all other groups received 8 μL / well of 10 mM H2O2 solution (final H2O2 concentration 400 μM) and continued incubation for another 4 h. The culture medium was discarded, and 100 μL of MTT (0.5 mg / ml) solution was added to each well. Cells were cultured for another 4 h, and the MTT solution was discarded. Then, 150 μL of DMSO was added to each well, and the mixture was shaken. The absorbance was measured at 570 nm using a microplate reader.

[0052] Experimental results showed that in the APAP model, compounds 4, 5, 6, 14, 18, and 21 significantly improved the survival rate of APAP-treated HepG2 cells, and compounds 3, 11, and 15 also showed certain hepatocellular damage protective activity; in the H2O2 model, compounds 3, 5, 9, 10, 11, and 17 significantly improved the survival rate of H2O2-treated HepG2 cells.

[0053] Table 1: Protective effect of compound 1-21 against APAP-induced HepG2 cell damage

[0054]

[0055] ***P < 0.001, compared with the blank control group; # P < 0.05 ## P < 0.01, ### P < 0.001, compared with the APAP model group.

[0056] Table 2: Protective effect of compound 1-21 against H2O2-induced HepG2 cell damage

[0057]

[0058]

[0059] ***P < 0.001, compared with the blank control group; # P < 0.05 ## P < 0.01, ### P < 0.001, compared with the APAP model group.

Claims

1. A class of compounds or pharmaceutically acceptable salts thereof, characterized in that, The structure of this type of compound is as follows:

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that, The pharmaceutically acceptable salts are selected from salts formed by compounds and inorganic or organic bases.

3. The compound according to claim 2 or a pharmaceutically acceptable salt thereof, characterized in that, The organic bases include methylamine, ethylamine, diethylamine, triethylamine, propylamine, butylamine, octylamine, hexamethylenediamine, ethylenediamine, propylenediamine, butylamine, benzylamine, phenethylamine, o-phenylenediamine, and p-phenylenediamine. The inorganic bases include sodium hydroxide, potassium hydroxide, calcium hydroxide, aluminum hydroxide, lithium hydroxide, magnesium hydroxide, zinc hydroxide, copper hydroxide, iron hydroxide, ammonium hydroxide, sodium bicarbonate, and potassium bicarbonate.

4. The method for preparing the compound of claim 1, characterized in that, The preparation method is as follows: the medicinal watermelon is extracted by reflux with 70% ethanol, and the concentrated extract is purified by organic solvent extraction, macroporous resin column chromatography, gel column chromatography, silica gel column chromatography and preparative HPLC to obtain compound 1-21.

5. A pharmaceutical composition, characterized in that, The pharmaceutical composition comprises the compound of any one of claims 1-3 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier or excipient.

6. The pharmaceutical composition according to claim 5, characterized in that, The dosage forms of the pharmaceutical composition include tablets, capsules, pills, granules, oral liquids, or suspensions.

7. Use of the compound according to any one of claims 1-3 in the preparation of a medicament with hepatoprotective function.

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