Tetrahydroanthraquinone from prismatomeria tetrandra, and preparation method, pharmaceutical composition and application thereof

Abstract

The invention discloses tetrahydroanthraquinone compounds as well as a preparation method, a pharmaceutical composition and application thereof, and particularly discloses 13 tetrahydroanthraquinone compounds (1-13) found from prismatomeria tetrandra, and a preparation method of the compounds through phytochemistry. The invention relates to a pharmaceutical composition containing tetrahydroanthraquinone and application thereof in treatment of pneumonia and pulmonary fibrosis.

Description

Technical Field

[0001] The invention belongs to the field of medicine, and specifically relates to tetrahydroanthraquinone compounds extracted and separated from yellow roots, a preparation method and use of the compounds in the preparation of prevention and / or treatment of pneumonia and pulmonary fibrosis. Background Art

[0002] Prismatomeris connata YZRuan is a plant of the genus Prismatomeris in the Rubiaceae family. It is mainly distributed in Guangdong, Guangxi, Hainan and other places. Its root is mostly used as medicine, which has the effects of cooling blood and stopping bleeding, strengthening tendons and bones, removing dampness and jaundice, removing blood stasis and promoting new growth. It can be used to treat traumatic injuries, rheumatic bone pain, leukemia, hepatitis, bleeding gums, thalassemia, aplastic anemia, etc.

[0003] Pneumonia refers to inflammation of the terminal airways, alveoli and lung interstitium, which can be caused by pathogenic microorganisms, physical and chemical factors, immune damage, allergies and drugs. Before the use of antibiotics, bacterial pneumonia posed a great threat to the health of children and the elderly. The emergence and development of antibiotics once significantly reduced the mortality rate of pneumonia, but in recent years, despite the use of powerful antibiotics and effective vaccines, the overall mortality rate of pneumonia has not decreased, and has even increased. The common folk drug Huanggen is boiled in water for oral administration for the prevention and treatment of diseases such as pneumonia and pulmonary fibrosis, but the compounds that play a role in it have not been clear before. Therefore, compounds 1-13 with clear chemical structures and significant anti-inflammatory activity isolated and purified from the traditional Chinese medicine Huanggen have excellent clinical development prospects for the prevention and treatment of pneumonia and pulmonary fibrosis. Summary of the invention

[0004] The inventors first discovered that the root extract of the yellow root has anti-inflammatory activity, and 13 new tetrahydroanthraquinone compounds were extracted and separated from the yellow root extract. Pharmacological evaluation showed that they had good anti-inflammatory effects.

[0005] The technical problem solved by the invention is to provide 13 new tetrahydroanthraquinone compounds.

[0006] Another technical problem solved by the present invention is to provide 13 new preparation methods of tetrahydroanthraquinones.

[0007] Another technical problem solved by the present invention is to provide a pharmaceutical composition, in which 13 tetrahydroanthraquinones have obvious anti-inflammatory activity and can be used to treat pneumonia and pulmonary fibrosis.

[0008] Specifically, the structural formula of compound 1-13 of the present invention is as follows:

[0009]

[0010] The present invention provides 13 new methods for preparing tetrahydroanthraquinones, which are as follows:

[0011] The dried root of the yellow root weighs 50 kg. After being crushed, it is refluxed and extracted with 80% ethanol for 3 times, each time for 2 hours, to obtain an 80% ethanol total extract of the yellow root. After reduced pressure concentration, about 2500 g of extract is obtained. This extract is dispersed in 3L of warm water and then extracted with ethyl acetate. The extraction is performed three times to obtain 132 g of ethyl acetate extraction part. The ethyl acetate extraction part is subjected to multiple column chromatography (forward and reverse silica gel, gel), and finally purified by HPLC to obtain 13 new tetrahydroanthraquinones.

[0012] Another aspect of the present invention also relates to a pharmaceutical composition with the compound of the present invention as an active ingredient. The pharmaceutical composition can be prepared according to methods known in the art. Any dosage form suitable for human or animal use can be prepared by combining the compound of the present invention with one or more pharmaceutically acceptable solid or liquid excipients and or adjuvants. The content of the compound of the present invention in its pharmaceutical composition is usually 0.1-95% by weight.

[0013] The compound of the present invention or the pharmaceutical composition containing it can be administered in a unit dosage form, and the administration route can be enteral or parenteral, such as oral, intravenous, intramuscular, subcutaneous, nasal, oral mucosa, eyes, lungs and respiratory tract, skin, vagina, rectum, etc.

[0014] The dosage form can be a liquid dosage form, a solid dosage form or a semisolid dosage form. Liquid dosage forms can be solutions (including true solutions and colloidal solutions), emulsions (including o / w types, w / o types and multiple emulsions), suspensions, injections (including water injections, powder injections and infusions), eye drops, nasal drops, lotions and liniments, etc.; solid dosage forms can be tablets (including ordinary tablets, enteric-coated tablets, lozenges, dispersible tablets, chewable tablets, effervescent tablets, orally disintegrating tablets), capsules (including hard capsules, soft capsules, enteric-coated capsules), granules, powders, micropills, dropping pills, suppositories, films, patches, aerosols (powders), sprays, etc.; semisolid dosage forms can be ointments, gels, pastes, etc.

[0015] 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.

[0016] In order to prepare the compound of the present invention into tablets, various excipients known in the art can be widely used, including diluents, binders, wetting agents, disintegrants, lubricants, and glidants. The diluent may be starch, dextrin, sucrose, glucose, lactose, mannose, sorbitol, xylitol, microcrystalline cellulose, calcium sulfate, calcium hydrogen phosphate, calcium carbonate, etc.; the wetting agent may be water, ethanol, isopropanol, etc.; the binder may be starch slurry, dextrin, syrup, honey, glucose solution, microcrystalline cellulose, acacia slurry, gelatin slurry, sodium carboxymethyl cellulose, methyl cellulose, hydroxypropyl methyl cellulose, ethyl cellulose, acrylic resin, carbomer, polyvinyl pyrrolidone, polyethylene glycol, etc.; the disintegrant may be dry starch, microcrystalline cellulose, low-substituted hydroxypropyl cellulose, cross-linked polyvinyl pyrrolidone, cross-linked sodium carboxymethyl cellulose, sodium carboxymethyl starch, sodium bicarbonate and citric acid, polyoxyethylene sorbitan fatty acid ester, sodium dodecyl sulfate, etc.; the lubricant and glidant may be talc, silicon dioxide, stearate, tartaric acid, liquid paraffin, polyethylene glycol, etc.

[0017] 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.

[0018] In order to prepare the dosing unit into a capsule, the active ingredient compound of the present invention can be mixed with a diluent and a glidant, and the mixture can be directly placed in a hard capsule or a soft capsule. The active ingredient compound of the present invention can also be first made into granules or pellets with a diluent, a binder, and a disintegrant, and then placed in a hard capsule or a soft capsule. Various diluents, binders, wetting agents, disintegrants, and glidants used to prepare tablets of the compound of the present invention can also be used to prepare capsules of the compound of the present invention.

[0019] To prepare the compound of the present invention into an injection, water, ethanol, isopropanol, propylene glycol or a mixture thereof can be used as a solvent and an appropriate amount of a solubilizer, a cosolvent, a pH regulator, and an osmotic pressure regulator commonly used in the art can be added. The solubilizer or cosolvent can be poloxamer, lecithin, hydroxypropyl-β-cyclodextrin, etc. The pH regulator can be phosphate, acetate, hydrochloric acid, sodium hydroxide, etc.; the osmotic pressure regulator can be sodium chloride, mannitol, glucose, phosphate, acetate, etc. When preparing a lyophilized powder injection, mannitol, glucose, etc. can also be added as a support.

[0020] Furthermore, if necessary, colorants, preservatives, perfumes, flavoring agents or other additives may be added to the pharmaceutical preparations.

[0021] 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.

[0022] The dosage of the pharmaceutical composition of the compound of the present invention can vary widely according to the nature and severity of the disease to be prevented or treated, the individual conditions of the patient or animal, the route of administration and the dosage form. Generally speaking, the suitable dosage range of the compound of the present invention per day is 0.001-150 mg / kg body weight, preferably 0.1-100 mg / kg body weight, more preferably 1-60 mg / kg body weight, and most preferably 2-30 mg / kg body weight. The above dosage can be administered in one dosage unit or divided into several dosage units, depending on the doctor's clinical experience and the dosage regimen including the use of other treatment means.

[0023] The compound or composition of the present invention can be taken alone or in combination with other therapeutic drugs or symptomatic drugs. When the compound of the present invention has a synergistic effect with other therapeutic drugs, its dosage should be adjusted according to the actual situation.

[0024] The present invention studies the activity of novel tetrahydroanthraquinone compounds in inhibiting cell inflammation by using an in vitro pharmacological model of LPS-induced macrophage NO release. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 --Flowchart of Yellow Root Extraction

[0026] Figure 2 --Separation flow chart of ethyl acetate extract of yellow root

[0027] Figure 3- -0.05μmol / mL monomer and 40μg / mL component drug treatment of RAW264.7 cells for 12 hours

[0028] Figure 4- -0.2μmol / mL monomer drug treatment of RAW264.7 cells for 12 hours Cell proliferation

[0029] Figure 5- - NO release in RAW264.7 cells induced by LPS treated with compounds 1-13 DETAILED DESCRIPTION

[0030] The examples and pharmacological activity experiments further illustrate the present invention but do not constitute any limitation to the present invention. 1. Preparation of compounds 1-13 (see Appendix Figure 1-2 )

[0031] The dried root of the yellow root weighs 50 kg. After being crushed, it is refluxed and extracted with 80% ethanol for 3 times, each time for 2 hours, to obtain 80% ethanol total extract of the yellow root. After reduced pressure concentration, about 2500 g of extract is obtained. This extract is dispersed in 3L of warm water and then extracted with ethyl acetate. The extraction is carried out three times to obtain 132 g of ethyl acetate extraction part.

[0032] Further separation was carried out by dichloroethane / methanol (50:1-5:1) silica gel column chromatography, and 40 g was obtained at the 10:1 elution portion. This portion was subjected to gradient elution by petroleum ether-ethyl acetate (5:1-1:4) on a silica gel column. After elution for 40 column volumes, one component was combined every 5 column volumes to obtain 8 elution components (recorded as Fr.1 to Fr.8).

[0033] Fr.6 (10.2 g) was subjected to medium pressure column chromatography (30% MeOH / H2O-100% MeOH / H2O, gradient elution for 9 hours) and about 500 ml was collected to obtain 45 fractions, named F1 to F45. F14 was subjected to C18 column HPLC (20% MeCN-H2O) to obtain compound 6 (1.8 mg). F16 was subjected to C18 column HPLC (55% MeOH-H2O) to obtain F16-1, and F16-1 was subjected to C18 column HPLC (30% MeCN-H2O) to obtain compound 5 (3.1 mg). F17 was prepared by cellulose column HPLC (60% MeCN-H2O) to obtain F17-1 and F17-2, F17-1 was prepared by C18 column HPLC (35% MeCN-H2O) to obtain compound 4 (5.0 mg), and F17-2 was prepared by C18 column HPLC (40% MeCN-H2O) to obtain compound 3 (3.5 mg). F18 was prepared by cellulose column HPLC (80% MeOH-H2O) to obtain F18-1, F18-2, F18-3, F18-1 was prepared by cellulose column HPLC (60% MeCN-H2O) to obtain compound 9 (2.5 mg), F18-2 was prepared by cellulose column HPLC (50% MeCN-H2O) to obtain compound 10 (2.8 mg), and F18-3 was prepared by C18 column HPLC (50% MeOH-H2O) to obtain compound 11 (3.3 mg). F19 was prepared by cellulose column HPLC (40% MeCN-H2O) to obtain F19-1, and F19-1 was prepared by cellulose column HPLC (80% MeOH-H2O) to obtain compound 12 (3.5 mg). F20 was prepared by C18 column HPLC (50% MeOH-H2O) to obtain F20-1, and F20-1 was prepared by cellulose column HPLC (90% MeOH-H2O) to obtain compound 13 (2.5 mg). F27 was prepared by cellulose column HPLC (85% MeCN-H2O) to obtain compound 8 (4.2 mg). F29 was prepared by C18 column HPLC (55% MeOH-H2O) to obtain compound 1 (1.1 mg). F30 was prepared by C18 column HPLC (60% MeOH-H2O) to obtain F30-1, and F30-1 was prepared by phenyl column HPLC (40% MeCN-H2O) to obtain compound 2 (3.8 mg). F34 was purified by C18 column HPLC (60% MeOH-H2O) to obtain F34-1, and F34-1 was purified by C18 column HPLC (40% MeCN-H2O) to obtain compound 7 (3.3 mg).

[0034] The physicochemical and spectral data of compound 1-14 are as follows:

[0035] The physicochemical and spectral data of compound 1 are as follows:

[0036] Light yellow powder; -46.0(c 0.10MeOH), (+)-HRESI-Ms m / z 395.1699[M+H] + , (calcd.for C 20 H 27 O8, 375.1700). 1 H NMR data (CDCl3, 600MHz), δ H : 4.49 (1H, m, H-1), 3.54 (1H, dd, J = 7.8, 10.8Hz, H-2), 1.82 (1H, m, H-3), 2. 17 (1H, ddd, J = 1.2, 5.4, 19.2Hz, H-4), 2.85 (1H, ddd, J = 3.6, 11.4, 19.2Hz, H-4), 6.98 (1H, s, H-8), 1.19 (3H, d, J = 6.0Hz, H-11), 3.93 (3H, s, H-12), 3. 92(3H,s,H-13), 3.96(3H,s,H-14), 3.01(3H,s,H-15), 3.07(3H,s,H-16).

[0037] 13 C NMR data (CDCl3, 150MHz), δ C : 74.3(C-1), 77.6(C-2), 32.6(C-3), 31.7(C-4), 141.0(C-4a), 154.4(C-5), 144.2(C-6), 158.4(C-7), 104.5(C-8), 136.6(C-8a), 99 .3(C-9), 141.2(C-9a), 182.0(C-10), 120.7(C-10a), 17.5(C-11), 61.6(C-12), 61.3(C-13), 56.5(C-14), 51.9(C-15), 52.3(C-16).

[0038] The physicochemical and spectral data of compound 2 are as follows:

[0039] Light yellow powder; -45.0(c 0.10MeOH), (+)-HRESI-MS m / z 387.1408[M+Na] + , (calcd.for C 19 H 24 O7Na, 387.1414). 1 H NMR data (CDCl3, 400MHz), δH : 5.16 (1H, d, J = 7.6Hz, H-1), 3.77 (1H, dd, J = 7.6, 10.8Hz, H-2), 2.10 (1H, m, H-3), 2.50 (1H, m, H-4), 3.31 (1H, m, H-4), 7. 26(1H,s,H-8), 1.43(3H,d,J=6.4Hz,H-11), 4.31(3H,s,H-12), 4.13(3H,s,H-13), 4.17(3H,s,H-14), 4.10(3H,s,H-15).

[0040] 13 C NMR data (CDCl3, 100MHz), δ C : 74.4(C-1), 78.2(C-2), 32.9(C-3), 31.6(C-4), 129.0(C-4a), 148.2(C-5), 140.3(C-6), 152.7(C-7), 97.0(C-8), 124.3(C-8) a), 145.3(C-9), 116.5(C-9a), 146.2(C-10), 112.7(C-10a), 17.8(C-11), 62.4(C-12), 61.3(C-13), 56.0(C-14), 60.8(C-15).

[0041] The physicochemical and spectral data of compound 3 are as follows:

[0042] Light yellow powder; -30.0(c 0.10MeOH), (+)-HRESI-MS m / z 349.1284[M+H] + , (calcd.for C 18 H 21 O7, 349.1282). 1 H NMR data (CDCl3, 600MHz), δ H : 4.89 (1H, d, J = 4.2Hz, H-1), 3.54 (1H, m, H-2), 2.11 (1H, m, H-3), 2.17 (1H, m, H-4), 2.97 (1H, m, H-4), 7 .47(1H,s,H-8), 1.14(3H,d,J=6.6Hz,H-11), 3.93(3H,s,H-12), 3.96(3H,s,H-13), 4.01(3H,s,H-14).

[0043] 13 C NMR data (CDCl3, 150MHz), δ C: 64.3(C-1), 72.8(C-2), 29.9(C-3), 30.5(C-4), 147.1(C-4a), 154.5(C-5), 148.4(C-6), 157.5(C-7), 106.0(C-8), 129 .3(C-8a), 185.3(C-9), 139.2(C-9a), 183.3(C-10), 119.9(C-10a), 17.0(C-11), 61.5(C-12), 61.7(C-13), 56.6(C-14).

[0044] The physicochemical and spectral data of compound 4 are as follows:

[0045] Light yellow powder; -56.0(c 0.10MeOH), (+)-HRESI-MS m / z 349.1282[M+H] + , (calcd.for C 18 H 21 O7, 349.1282). 1 H NMR data (CDCl3, 400MHz), δ H : 4.69 (1H, ddd, J = 1.2, 3.2, 8.0Hz, H-1), 3.50 (1H, dd, J = 8.0, 11.2Hz, H-2), 1.83 (1H, m, H-3), 2.15 (1H, ddd, J = 3.2, 11.2, 20.0Hz, H-4), 2.95 (1H, ddd, J=1.2, 5.2, 20.0Hz, H-4), 7.44 (1H, s, H-5), 1.20 (3H, d, J=6.4Hz, H-11), 4.00 (3H, s, H-12), 3.95 (3H, s, H-13), 3.92 (3H, s, H-14).

[0046] 13 C NMR data (CDCl3, 100MHz), δ C : 72.9(C-1), 77.4(C-2), 32.0(C-3), 31.8(C-4), 146.4(C-4a), 154.4(C-5), 157.5(C-6), 148.5(C-7), 106.1(C-8), 119 .7(C-8a), 182.9(C-9), 139.5(C-9a), 186.2(C-10), 129.4(C-10a), 17.4(C-11), 61.7(C-12), 56.6(C-13), 61.5(C-14).

[0047] The physicochemical and spectral data of compound 5 are as follows:

[0048] Light yellow powder; +15.0(c 0.10MeOH), (+)-HRESI-MS m / z 335.1127[M+H] + , (calcd.for C 17 H 19 O7, 335.1125). 1 H NMR data (DMSO-d6, 400MHz), δ H : 2.51 (1H, mH-1), 2.72 (1H, d, J = 19.2Hz, H-1), 3.60 (1H, d, J = 4.0Hz, H-2), 2.46 (1H, m, H-4), 2.51(1H,m,H-4), 7.21(1H,s,H-5), 1.23(3H,s,H-11), 3.96(3H,s,H-12), 3.82(3H,s,H-13).

[0049] 13 C NMR data (DMSO-d6, 100MHz), δ C :29.7(C-1), 69.2(C-2), 68.6(C-3), 32.6(C-4), 142.2(C-4a), 103.6(C-5), 157.5(C-6), 140.4(C-7), 154.7(C-8 ), 110.4(C-8a), 188.8(C-9), 142.4(C-9a), 183.1(C-10), 127.5(C-10a), 25.7(C-11), 56.4(C-12), 60.3(C-13).

[0050] The physicochemical and spectral data of compound 6 are as follows:

[0051] Light yellow powder; +17.0(c 0.10MeOH), (+)-HRESI-MS m / z 259.0962[M+H] + , (calcd.for C 15 H 15 O4, 259.0965). 1 H NMR data (MeOD, 400MHz), δ H: 2.72 (1H, mH-1), 2.95 (1H, m, H-1), 3.79 (1H, t, J = 4.0Hz, H-2), 2.66 (2H, m, H-4), 8.02 (2H, m, H-5 / 8), 7.74 (2H, m, H-6 / 7), 1.36 (3H, s, H-11).

[0052] 13 C NMR data (MeOD, 100MHz), δ C :30.5(C-1), 71.5(C-2), 70.8(C-3), 34.3(C-4), 143.4(C-4a), 127.0(C-5 / 8), 133.5(C-6 / 7), 134.6(C-8a), 185.9(C-9), 142.8(C-9a), 185.7(C-10), 134.6(C-10a), 25.3(C-11).

[0053] The physicochemical and spectral data of compound 7 are as follows:

[0054] Light yellow powder; -46.0(c 0.10MeOH), (+)-HRESI-MS m / z 319.1173[M+H] + , (calcd.for C 17 H 19 O6, 317.1176). 1 H NMR data (CDCl3, 500MHz), δ H :2.53(1H,mH-1), 3.04(1H,m,H-1), 3.78(1H,m,H-2), 1.94(2H,m,H-3), 2.32(1H,m,H-4), 2.95( 1H, m, H-4), 7.30 (1H, s, H-8), 1.14 (3H, d, J = 7.0Hz, H-11), 4.02 (3H, s, H-12), 4.03 (3H, s, H-13).

[0055] 13 C NMR data (CDCl3, 100MHz), δ C: 31.1(C-1), 70.8(C-2), 33.9(C-3), 29.0(C-4), 143.4(C-4a), 155.7(C-5), 141.4(C-6), 157.9(C-7), 103.9(C-8 ), 111.1(C-8a), 183.4(C-9), 142.6(C-9a), 189.0(C-10), 127.9(C-10a), 17.4(C-11), 61.2(C-12), 56.6(C-13).

[0056] The physicochemical and spectral data of compound 8 are as follows:

[0057] Light yellow powder; +49.0(c 0.10MeOH), (+)-HRESI-MS m / z 319.1174[M+H] + , (calcd.for C 17 H 19 O6, 317.1176). 1 H NMR data (MeOD, 400MHz), δ H : 2.68 (1H, mH-1), 2.89 (1H, t, J=4.0Hz, H-1), 3.78 (1H, m, H-2), 2.61 (1H, m, H-4), 7.85 (1H, d, J=8 .4Hz, H-5), 7.33 (1H, d, J = 8.4Hz, H-6), 1.34 (3H, s, H-11), 3.96 (3H, s, H-12), 3.84 (3H, s, H-13).

[0058] 13 C NMR data (MeOD, 100MHz), δ C :30.7(C-1), 71.7(C-2), 70.8(C-3), 34.1(C-4), 142.1(C-4a), 125.3(C-5), 117.0(C-6), 160.1(C-7), 150.0(C-8 ), 126.3(C-8a), 185.5(C-9), 144.1(C-9a), 184.9(C-10), 127.1(C-10a), 25.2(C-11), 56.8(C-12), 61.4(C-13).

[0059] The physicochemical and spectral data of compound 9 are as follows:

[0060] Light yellow powder; -52.0(c 0.10MeOH), (+)-HRESI-Ms m / z 289.1072[M+H] + , (calcd.for C 16 H 17 O5, 289.1071). 1 H NMR data (CDCl3, 600MHz), δ H : 2.49 (1H, mH-1), 3.02 (1H, t, J=4.0Hz, H-1), 3.74 (1H, ddd, J=4.8, 7.2, 12.0Hz, H-2), 1.88 (1H, m, H-3), 2.28 (1H, m, H-4) , 2.92 (1H, m, H-4), 7.24 (1H, d, J = 8.4Hz, H-7), 7.88 (1H, d, J = 8.4Hz, H-8), 1.10 (3H, d, J = 6.6Hz, H-11), 3.96 (3H, s, H-12).

[0061] 13 C NMR data (CDCl3, 150MHz), δ C :31.2(C-1), 71.0(C-2), 34.0(C-3), 29.5(C-4), 142.5(C-4a), 146.1(C-5), 155.2(C-6), 119.6(C-7), 125 .0(C-8), 126.3(C-8a), 183.6(C-9), 142.5(C-9a), 184.1(C-10), 142.5(C-10a), 17.4(C-11), 62.3(C-12).

[0062] The physicochemical and spectral data of compound 10 are as follows:

[0063] Light yellow powder; -81.0(c 0.10MeOH), (+)-HRESI-MS m / z319.1178[M+H] + , (calcd.for C 17 H 19 O6, 319.1176). 1 H NMR data (MeOD, 500MHz), δ H:2.38(1H,mH-1), 2.92(1H,m,H-1), 3.61(1H,m,H-2), 1.82(1H,m,H-3), 2.21(1H,m,H-4), 2.89( 1H, m, H-4), 7.33 (1H, s, H-8), 1.12 (3H, d, J = 7.0Hz, H-11), 3.90 (3H, s, H-12), 3.95 (3H, s, H-13).

[0064] 13 C NMR data (MeOD, 100MHz), δ C :31.7(C-1), 71.4(C-2), 35.3(C-3), 31.2(C-4), 146.0(C-4a), 155.8(C-5), 157.4(C-6), 147.5(C-7), 111.8(C-8 ), 119.2(C-8a), 185.2(C-9), 141.2(C-9a), 184.3(C-10), 146.0(C-10a), 17.8(C-11), 61.8(C-12), 61.5(C-13).

[0065] The physicochemical and spectral data of compound 11 are as follows:

[0066] Light yellow powder; -66.0(c 0.10MeOH), (+)-HRESI-MS m / z 319.1176[M+H] + , (calcd.for C 17 H 19 O6, 319.1176). 1 H NMR data (CDCl3, 400MHz), δ H :2.48(1H,mH-1), 3.01(1H,m,H-1), 3.72(1H,m,H-2), 1.88(1H,m,H-3), 2.25(1H,m,H-4), 2.88( 1H, m, H-4), 7.45 (1H, s, H-5), 1.09 (3H, d, J = 6.5Hz, H-11), 4.03 (3H, s, H-12), 3.95 (3H, s, H-13).

[0067] 13 C NMR data (CDCl3, 100MHz), δ C:31.2(C-1), 71.1(C-2), 34.0(C-3), 29.4(C-4), 141.5(C-4a), 105.8(C-5), 151.1(C-6), 144.3(C-7), 146.9(C-8 ), 119.3(C-8a), 183.3(C-9), 142.8(C-9a), 183.9(C-10), 126.2(C-10a), 17.4(C-11), 56.7(C-12), 61.8(C-13).

[0068] 41.2(C-9a), 184.3(C-10), 146.0(C-10a), 17.8(C-11), 61.8(C-12), 61.5(C-13).

[0069] The physicochemical and spectral data of compound 12 are as follows:

[0070] Light yellow powder; -48.0(c 0.10MeOH), (+)-HRESI-MS m / z 289.1071[M+H] + , (calcd.for C 16 H 17 O5, 289.1070). 1 H NMR data (MeOD, 500MHz), δ H : 2.36 (1H, mH-1), 2.92 (1H, m, H-1), 3.59 (1H, m, H-2), 1.79 (1H, m, H-3), 2.17 (1H, m, H-4), 2.84 ( 1H, m, H-4), 7.31 (1H, s, H-5), 7.46 (1H, s, H-8), 1.09 (3H, d, J = 7.0Hz, H-11), 3.98 (3H, s, H-12).

[0071] 13 C NMR data (MeOD, 100MHz), δ C : 32.0(C-1), 71.5(C-2), 35.3(C-3), 31.0(C-4), 144.0(C-4a), 113.2(C-5), 153.0(C-6), 153.4(C-7), 109 .1(C-8), 127.1(C-8a), 185.2(C-9), 142.7(C-9a), 185.4(C-10), 128.7(C-10a), 17.8(C-11), 56.7(C-12).

[0072] The physicochemical and spectral data of compound 13 are as follows:

[0073] Light yellow powder; -89.0(c 0.10MeOH), (+)-HRESI-MS m / z 259.0966[M+H] + , (calcd.for C 15 H 15 O4, 259.6905). 1 H NMR data (MeOD, 400MHz), δ H : 2.21 (1H, mH-1), 2.85 (1H, m, H-1), 3.61 (1H, m, H-2), 1.81 (1H, m, H-3), 2.40 (1H, m, H-4), 2.93 (1H, m, H-4), 7.3 5 (1H, d, J = 2.4Hz, H-5), 7.07 (1H, dd, J = 2.48.4Hz, H-7), 7.90 (1H, d, J = 8.4Hz, H-8), 1.09 (3H, d, J = 6.4Hz, H-11).

[0074] 13 C NMR data (MeOD, 100MHz), δ C :30.9(C-1), 71.5(C-2), 35.2(C-3), 31.9(C-4), 144.7(C-4a), 112.9(C-5), 164.1(C-6), 121.3(C-7 ), 129.9(C-8), 125.8(C-8a), 184.9(C-9), 142.7(C-9a), 185.9(C-10), 135.7(C-10a), 17.8(C-11).

[0075] Pharmacological experiments

[0076] 1. Purpose of the experiment

[0077] The LPS-induced macrophage NO release model (inflammatory model) was used to screen new components and new monomer compounds of Xanthoceras radix that inhibit cellular inflammation.

[0078] 2. Experimental Principle

[0079] The NO concentration is determined using a one-step method. NO generates nitrate and nitrite when it meets oxygen and water. The latter two can generate light red azo compounds when they meet nitrate color developer. The NO concentration can be indirectly measured by colorimetry.

[0080] 3. Experimental instruments and reagents

[0081] Experimental instruments: cell culture incubator, clean bench, microplate reader, cell counting plate, cell culture flask.

[0082] Experimental reagents: DMEM / F12 cell culture medium, penicillin-streptomycin double antibody, fetal bovine serum, lipopolysaccharide, CCK8 cell proliferation reagent, nitric oxide assay kit.

[0083] 4. Experimental Procedure

[0084] 4.1 Cell culture

[0085] RAW264.7 cells were cultured in complete medium (20% fetal bovine serum + 2% penicillin-streptomycin) and cultured at 5.0×10 5 The cells were plated at a density of 100 μL / mL in a 96-well plate, with 200 μL per well, and cultured overnight at 37°C in a 5% CO2 environment.

[0086] 4.2 Cell proliferation

[0087] Serum-free medium was used to prepare 40 μg / mL component solution and 0.05 μmol / mL and 0.2 μmol / mL monomer solution as working solution. 200 μL of working solution was used to culture cells for 12 hours in each well. At the same time, blank wells cultured with serum-free medium and zero wells with only serum-free medium and no cells were set. After 12 hours, the medium in the wells was discarded, and CCK8 working solution (blank medium: CCK8 reagent = 10: 1) was added. The wells were cultured at 37°C for 2 hours, and the absorbance was measured at 450nm. The cell proliferation rate was calculated as (measurement well-zero well) / (blank well-zero well) × 100%

[0088] 4.3 Cell stimulation and intervention

[0089] LPS (1 μg / mL) solution, LPS (1 μg / mL) + drug component (40 μg / mL, 18 μg / mL, 8 μg / mL) solution and LPS (1 μg / mL) + drug monomer (0.2 μmol / mL, 0.05 μmol / mL, 0.01 μmol / mL) solution were prepared in serum-free culture medium. Blank group, LPS group and LPS + drug intervention group were set up. 200 μL of cells were cultured in each well for 12 hours. After 12 hours, the cell supernatant was aspirated for subsequent NO concentration determination.

[0090] 4.4 NO determination

[0091] Take 100 μL of cell supernatant, add 200 μL of NO reagent 1, mix well, add 100 μL of reagent 2, vortex thoroughly mix, let stand for 10 minutes, centrifuge at 4000 rpm for 15 minutes, and take 160 μL of supernatant for testing. Prepare NO colorimetric reagent according to the ratio of reagent 3: reagent 4: reagent 5 = 2.5: 1: 1, set blank well (add 160 μL of double distilled water), standard well (add 160 μL of 20 μmol / L sodium nitrite standard solution) and measurement well, add 80 μL of colorimetric reagent to each well, mix well, let stand for 15 minutes, measure absorbance at 550 nm, and calculate NO content (μmol / L) = (measurement well - blank well) / (standard well - blank well) × standard well concentration × 4

[0092] 5. Experimental Results

[0093] like Figure 3 The results showed that, except for the significant difference in cell proliferation rate between the 0.05 μmol / mL compound 6 treatment group and the blank group, no obvious cell proliferation inhibition was observed in the other drug treatment groups. The cell proliferation rate of the 0.05 μmol / mL compound 6 12h treatment group was 90.4487% (>90%), and this concentration was considered to be a subsequent drug intervention concentration.

[0094] like Figure 4 The results showed that, except for the significant difference in cell proliferation rate between the 0.2 μmol / mL compound 6 treatment group and the blank group, no obvious cell proliferation inhibition was observed in the other drug treatment groups. The cell proliferation rate of the 0.2 μmol / mL compound 6 12h treatment group was 64.3556% (<90%), and it was considered that this concentration could not be used as a subsequent drug intervention concentration.

[0095] Results of different drugs inhibiting LPS-induced NO release in RAW264.7 cells

[0096] Table 1 Results of different drugs inhibiting LPS-induced NO release in RAW264.7 cells

[0097]

[0098] like Figure 5The results showed that after 1 μg / mL LPS induced RAW264.7 cells for 12 hours, the NO release in the model group increased significantly, and the 0.2 μmol / mL treatment group of compound 1, compound 3, compound 9, and compound 10 decreased significantly, and the 0.2 μmol / mL and 0.05 μmol / mL treatment groups of compound 2, compound 4, and compound 8 decreased significantly in a dose-dependent manner, and the 0.2 μmol / mL, 0.05 μmol / mL, and 0.01 μmol / mL treatment groups of compound 5 and compound 7 decreased significantly in a dose-dependent manner, and the 0.05 μmol / mL and 0.01 μmol / mL treatment groups of compound 6 decreased significantly, and no dose-dependency was observed. The 0.2 μmol / mL treatment group of compound 11 decreased significantly, and the 0.2 μmol / mL and 0.05 μmol / mL treatment groups of compound 12 and compound 13 decreased significantly in a dose-dependent manner.

[0099] 6. Conclusion and discussion

[0100] NO has a wide range of biological effects in the body, including relaxing vascular smooth muscle, inhibiting platelet aggregation, mediating cytotoxic effects and immune regulation. NO produced by macrophages is closely related to cellular inflammation and immune activity. Abnormal production of NO by macrophages can lead to the occurrence and development of a variety of diseases, such as cancer, lupus erythematosus, rheumatoid arthritis, pneumonia, COPD, organ fibrosis, etc.

[0101] The high-dose groups of compound 1-13 monomer drugs can significantly reduce LPS-induced NO release. In terms of the inhibition of NO release, compound 6 has the strongest activity, and compound 5, compound 7, compound 12, compound 13, compound 4, compound 2, and compound 8 can all inhibit LPS-induced NO release.

Claims

1. A tetrahydroanthraquinone compound or a pharmaceutically acceptable salt thereof as shown in Formula 1-13 2. A pharmaceutical composition, characterized in that Contains at least one compound as claimed in claim 1 or a pharmaceutically acceptable salt thereof and a pharmacologically acceptable carrier.

3. Use of the compound of claim 1 or a pharmaceutically acceptable salt thereof in the preparation of a drug for preventing and / or treating pneumonia.

4. Use of the compound of claim 1 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for preventing and / or treating pulmonary fibrosis.

5. Use of the compound of claim 1 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for preventing and / or treating inflammation-related diseases.

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