Asiaticoside compound and pharmaceutical use thereof

Abstract

Provided are an asiaticoside compound and a pharmaceutical use thereof. The asiaticoside compound comprises a compound having a structural formula as shown in formula (I), a pharmaceutically acceptable salt or ester thereof, or a solvate thereof. The compound of formula (I) of the present invention has anti-inflammatory and anti-ulcerative activities, and can thus be used in the preparation of a drug for preventing or treating inflammatory diseases or ulcerative diseases.

Description

Centella asiatica saponins and their medicinal uses Technical Field

[0001] This invention relates to the field of biomedicine, and in particular to asiatic acid saponins and their medicinal uses. Background Technology

[0002] Inflammation is a common pathological mechanism in acute and chronic diseases, leading to tissue damage, fibrosis, and even organ failure, which can be life-threatening. Furthermore, persistent chronic low-grade inflammation can promote inflammation-cancer transformation, leading to tumor development and progression. In other words, inflammation is the root cause of almost all diseases. Traditional anti-inflammatory drugs mainly include steroidal anti-inflammatory drugs (such as dexamethasone and budesonide), nonsteroidal anti-inflammatory drugs (such as indomethacin and diclofenac), and immunosuppressants (such as cyclosporine A and hydroxychloroquine). However, these drugs have limited efficacy against inflammatory and autoimmune diseases such as systemic lupus erythematosus, and long-term use can lead to adverse reactions. For example, steroidal anti-inflammatory drugs can cause osteoporosis, hyperglycemia, obesity, and hypertension; nonsteroidal anti-inflammatory drugs can cause gastrointestinal ulcers, bleeding, and abnormal kidney function; and immunosuppressants can cause bone marrow suppression and infection. Emerging therapies for inflammatory and autoimmune diseases include inflammatory cytokine antibodies (such as adalimumab, ustekinumab, and gusejinumab), cytokine receptor antibodies (such as dupilumab and tocilizumab), and small molecule targeted drugs (such as utpatinib, criborone, and deuterocelexitinib). However, the clinical indications for existing biologics and small molecule targeted drugs for inflammatory and autoimmune diseases remain limited (e.g., limited or no efficacy in systemic lupus erythematosus), and biologics are expensive and can cause adverse reactions such as infections. Therefore, there is an urgent clinical need to develop safer and more effective anti-inflammatory drugs, especially small molecule anti-inflammatory drugs with lower treatment costs.

[0003] Peptic ulcer, primarily referring to chronic ulcers occurring in the stomach and duodenum, is a common and frequently occurring disease that severely impacts patients' quality of life. Severe ulcers can lead to upper gastrointestinal bleeding (with a mortality rate as high as 8%–13.7% for massive bleeding). The main causes of peptic ulcers are excessive gastric acid secretion, Helicobacter pylori infection, and drug-induced gastric mucosal damage (such as long-term use of low-dose aspirin). Currently, the first-line drugs for treating peptic ulcers in clinical practice are mainly potassium-competitive acid blockers (vonoprazan) and proton pump inhibitors (omeprazole and lansoprazole). However, acid-suppressing drugs such as vonoprazan and omeprazole only inhibit gastric acid secretion, but their repair effect on gastrointestinal mucosal damage is very limited, leading to a long treatment course and some adverse reactions. For example, a typical course of treatment with vonoprazan fumarate tablets is 4-8 weeks, with side effects including diarrhea, constipation, drug hypersensitivity reactions (including anaphylactic shock), drug-induced dermatitis, urticaria, jaundice, and erythema multiforme. A course of treatment with omeprazole is typically 1-2 months, with common adverse reactions including headache, abdominal pain, nausea, diarrhea, and constipation. Furthermore, as a CYP2C19 inhibitor, omeprazole significantly inhibits the antithrombotic efficacy of clopidogrel (FDA black box warning). It is well known that long-term use of low-dose aspirin (to prevent cardiovascular and cerebrovascular thrombotic events) can lead to gastrointestinal ulcers and bleeding. Currently, there is a lack of safe and effective gastrointestinal protective drugs for long-term use. Due to the safety risks of long-term use and potential drug-drug interactions, neither vonoprazan nor omeprazole is suitable for long-term use to prevent gastrointestinal damage caused by aspirin. In conclusion, there is an unmet clinical need for the prevention and treatment of peptic ulcers, and there is an urgent need to develop new drugs for peptic ulcers.

[0004] 2α,3β,23α-Trihydroxy-arbutane-12(13)-en-28-acid (asiatic acid) is an arbutane-type pentacyclic triterpenoid saponin, mainly found in Centella asiatica. Centella asiatica is an internationally recognized herb with wound-healing properties, and its main active ingredients include asiaticoside, hydroxyasiaticoside, asiatic acid, and hydroxyasiatic acid. Asiaticoside and hydroxyasiaticoside have a wide range of pharmacological effects, such as neuroprotection, cardioprotection, hepatoprotection, anti-inflammation, antioxidant, anti-allergy, antidepressant, anti-anxiety, antifibrotic, antibacterial, anti-arthritis, wound-healing, antitumor, and immunomodulatory activities. Med. Sci. Monit. 2020, 26, 924; Med. Sci. Monit. 2019, 25, 1355; Toxicol. Res. 2021, 37, 261). Asiaticoside can be metabolized in vivo to produce asiatic acid and related saponin metabolites (Nat. Prod. Rep. 2011, 28, 543-593), but the material basis and mechanism of action of asiaticoside in vivo are currently unclear. Preparations of Centella asiatica extract containing asiaticoside and hydroxyasiaticoside as the main active ingredients are clinically used to promote wound healing, reduce scarring (e.g., asiaticoside tablets and asiaticoside cream), and treat diabetic foot ulcers (e.g., "Subiyi" cream).

[0005] Several obstacles have been encountered when attempting to develop asiaticoside or hydroxyasiaticoside monomers as natural medicines. For example, the proportions of saponins and impurity profiles vary significantly among Centella asiatica plants from different origins, making it difficult to establish quality standards. Furthermore, asiaticoside and / or hydroxyasiaticoside are difficult to purify to high purity (e.g., >98%), making large-scale preparation of high-purity asiaticoside or hydroxyasiaticoside monomers extremely costly and difficult to industrialize. On the other hand, natural asiatic acid saponins (such as asiaticoside) are primarily found in Centella asiatica, while other types of asiatic acid saponins are very rare, and their activity is unknown. Summary of the Invention

[0006] Purpose of the invention: To address the problems existing in the prior art, the present invention provides a novel asiatic acid saponin compound with anti-inflammatory and anti-ulcer activities.

[0007] Another object of the present invention is to provide the use of asiatic acid saponins or pharmaceutically acceptable salts, esters or solvates thereof in the preparation of medicaments for the prevention or treatment of inflammatory or ulcerative diseases.

[0008] Technical solution: To achieve the above objectives, the present invention provides asiatic acid saponin compounds of the following formula (I) or their pharmaceutically acceptable salts, esters, or solvates:

[0009] Among them, R 1 Selected from H, β-D-glucopyranosyl, β-D-galactopyranosyl, β-D-xylanosyl, α-L-arabinopyranosyl, α-L-rhamnosyl, α-L-rhamnosyl-(1→2)-α-L-arabinopyranosyl, α-L-rhamnosyl-(1→3)-α-L-arabinopyranosyl, β-D-xylanosyl-(1→2)-α-L-arabinopyranosyl, β-D-xylanosyl-(1→3)-α-L-arabinopyranosyl, β-D-glucopyranosyl-(1→2)-α-L-arabinopyranosyl, or β-D-glucopyranosyl-(1→3)-α-L-arabinopyranosyl;

[0010] R 2 Selected from H, β-D-glucopyranosyl, β-D-galactopyranosyl, β-D-xylanosyl, α-L-arabinopyranosyl, α-L-rhamnosyl, α-L-rhamnosyl-(1→2)-α-L-arabinopyranosyl, α-L-rhamnosyl-(1→3)-α-L-arabinopyranosyl, β-D-xylanosyl-(1→2)-α-L-arabinopyranosyl, β-D-xylanosyl-(1→3)-α-L-arabinopyranosyl, β-D-glucopyranosyl-(1→2)-α-L-arabinopyranosyl, or β-D-glucopyranosyl-(1→3)-α-L-arabinopyranosyl;

[0011] R 1 and R 2 They are not both H.

[0012] In some embodiments, the asiatic acid saponin compound represented by formula (I) or its pharmaceutically acceptable salt, ester, or solvate is selected from any one of those shown in Table 1 below.

[0013] Table 1. Structure and Nomenclature of Compounds

[0014] The compounds of the present invention can also be used as pharmaceutical salts, wherein the salt is a salt formed by the compounds of the present invention with metal ions (including sodium, potassium, calcium, etc.) or pharmaceutically acceptable amines (including ethylenediamine, tromethamine, etc.) or ammonium ions.

[0015] The compounds of the present invention can also be used to form pharmaceutical compositions in the form of esters, prodrugs, or solvates thereof.

[0016] This invention provides the use of the aforementioned asiatic acid saponins in the preparation of medicaments for the prevention or treatment of inflammatory diseases, ulcerative diseases, or tumor diseases.

[0017] The inflammatory diseases mentioned are metabolic diseases, including: insulin resistance, metabolic syndrome, type 1 or type 2 diabetes, hyperlipidemia, obesity, atherosclerosis, myocardial ischemia, myocardial infarction, arrhythmia, coronary heart disease, hypertension, heart failure, myocardial hypertrophy, myocarditis, diabetic complications (including diabetic cardiomyopathy, diabetic nephropathy, retinopathy, neuropathy, diabetic foot ulcers, delayed wound healing, etc.), non-alcoholic fatty liver disease, non-alcoholic steatohepatitis, alcoholic fatty liver disease, cirrhosis, hyperuricemia, gout, osteoporosis, stroke or cerebral infarction, etc.

[0018] The inflammatory diseases mentioned are secondary diseases caused by autoimmune diseases, organ fibrosis, neurodegenerative diseases, or pathogen infections, including: pneumonia, tuberculosis, inflammatory bowel disease (such as Crohn's disease and ulcerative colitis), Behcet's disease, asthma, chronic obstructive pulmonary disease, chronic bronchitis, emphysema, bronchiolitis obliterans, systemic lupus erythematosus, rheumatoid arthritis, spondyloarthritis, osteoarthritis, synovitis, tendinitis, thromboangiitis obliterans, phlebitis, and intermittent claudication. Keloids, psoriasis, ichthyosis, bullous pemphigoid, dermatitis, contact dermatitis, pancreatitis, chronic nephritis, cystitis, meningitis, gastritis, sepsis, pyoderma gangrenosa, uveitis, idiopathic pulmonary fibrosis, cystic fibrosis, Parkinson's disease, Alzheimer's disease, alpha-complexoprotein disease, depression, multiple sclerosis, systemic sclerosis, amyotrophic lateral sclerosis, fibromyalgia syndrome, neuralgia, Down syndrome, Hallewarden-Schiller disease, Huntington's disease, or Wilson's disease, etc.

[0019] The ulcerative diseases mentioned above are peptic ulcers, including: gastric ulcers, duodenal ulcers, reflux esophagitis, or erosive esophagitis, etc.

[0020] The tumor diseases mentioned include: bone cancer, acute myeloid leukemia, chronic myeloid leukemia, acute lymphoblastic leukemia, chronic lymphoblastic leukemia, myeloproliferative disorders, multiple myeloma, myelodysplastic syndrome, Hodgkin's lymphoma, non-Hodgkin's lymphoma, hemangioma, granuloma, xanthoma, meningeal sarcoma, glioma, astrocytoma, medulloblastoma, ependymoma, germ cell tumor (pineal tumor), glioblastoma multiforme, oligodendroglioma, schwannoma, retinoblastoma, neurofibroma, sarcoma, esophageal cancer, gastric cancer, pancreatic cancer, colorectal cancer, colon cancer, rectal cancer, kidney cancer, prostate cancer, lymphoma, testicular cancer, interstitial cell carcinoma, lung cancer, liver cancer, skin cancer, malignant melanoma, or basal cell carcinoma, etc.

[0021] The present invention also provides a pharmaceutical composition for the prevention or treatment of inflammatory or ulcerative diseases, comprising a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt, ester, or solvate thereof as an active ingredient and pharmaceutically acceptable excipients. The excipients, which can be mixed in any way, may vary depending on the dosage form, administration method, etc. Examples of excipients include excipients, binders, disintegrants, lubricants, flavoring agents, fragrances, colorings, or sweeteners.

[0022] The pharmaceutical composition may be a conventional pharmaceutical formulation, such as capsules, powders, tablets, granules, pills, injections, syrups, oral liquids, inhalers, ointments, suppositories, or patches.

[0023] Furthermore, the compounds of the present invention can be used in combination with one or more other types of drugs for the prevention or treatment of inflammatory or ulcerative diseases.

[0024] The dosage of the compounds of the present invention may be appropriately varied according to the patient's age, weight, symptoms, and route of administration. When administered to an adult (approximately 60 kg), the dosage of the compound of formula (I) or its pharmaceutically acceptable salt, ester, or solvation is 1 mg to 1000 mg per dose, preferably 1 mg to 500 mg per dose, more preferably 5 mg to 60 mg per dose, administered 1 to 3 times daily.

[0025] Beneficial effects: Compared with the prior art, the present invention has the following advantages:

[0026] (1) The asiatic acid saponins of the present invention have significant anti-inflammatory effects, and the anti-inflammatory activity of some compounds is significantly better than that of asiatic acid and asiaticoside, and thus they are expected to be developed into new anti-inflammatory drugs.

[0027] (2) The asiatic acid saponins of the present invention have anti-peptic ulcer effects. In particular, some preferred compounds (such as compounds 3, 4, 6, 13, 15, 17, 19, 20, 22, 26, 29, etc.) have significantly better efficacy than omeprazole, a commonly used clinical drug for treating gastric ulcers, at the same dosage. Therefore, they are expected to be developed into safe and effective new anti-peptic ulcer drugs.

[0028] (3) Compared with natural asiaticoside or hydroxyasiaticoside, the asiaticoside saponins of the present invention are easy to prepare and purify, and are inexpensive, and can be effectively used in industrial production. Attached Figure Description

[0029] Figure 1 shows the effect of the saponin compounds of the present invention on the inhibitory effect of LPS-induced inflammatory response in THP1-derived macrophages (ELISA detection, *p<0.05, **p<0.01, ***p<0.001 vs. model group).

[0030] Figure 2 shows the gastric anatomy of a rat model of ethanol-induced gastric ulcer.

[0031] Figure 3 shows the mouse colon of the anti-ulcerative colitis model of compound 6 of the present invention;

[0032] Figure 4 is a statistical diagram of colon length in mice using compound 6 of the present invention to treat ulcerative colitis. ### p<0.001 vs. control group; *p<0.05, **p<0.01, ***p<0.001 vs. model group). Detailed Implementation

[0033] The technical solution of the present invention will be further described below.

[0034] This invention discloses novel asiatic acid saponin compounds and their pharmaceutical applications as anti-inflammatory and anti-ulcer drugs. Those skilled in the art can refer to the content of this document and appropriately modify the process parameters to achieve the desired results. It is particularly important to note that all similar substitutions and modifications are obvious to those skilled in the art and are considered to be included in this invention. The methods and applications of this invention have been described through preferred embodiments. Those skilled in the art can clearly modify or appropriately change and combine the methods and applications described herein without departing from the content, spirit, and scope of this invention to realize and apply the technology of this invention.

[0035] All chemical reagents used in the experiment were commercially available and had a chemical purity greater than 95%. Solvents used were analytical grade or chemically pure products. The petroleum ether used had a boiling range of 60 to 90°C. Unless otherwise specified, all purchased reagents and solvents were used directly without further processing. The reaction process was monitored by thin-layer chromatography (TLC) on silica gel plates, and detected under UV light at 254 nm and 365 nm, as well as by color development with phosphomolybdic acid and ethanolic sulfuric acid. The final product was purified by silica gel column chromatography (silica gel: 200-300 mesh). 1 1H NMR data were determined using an AVANED AV-300 NMR spectrometer (Analysis and Testing Center, China Pharmaceutical University). The deuterated solvents CDCl3 and DMSO-d6 were purchased from Cambridge Microsystems, USA. Tetramethylsilane (TMS) was used as an internal standard. The chemical shift of CDCl3 was 7.26 ppm, and that of DMSO-d6 was 2.50 ppm. In the reported data, s represents a singlet, d represents a doublet, t represents a doublet, td represents a triplet, q represents a quartet, m represents a multiplet, and br represents a broad peak. (Final product...) 13 CNMR data and high temperature (80°C) 11H NMR data were analyzed using an AVANCE III 400 / 500MHz Digital NMR Spectrometer (Shanghai Institute of Plant Physiology, Shanghai Institutes for Biological Sciences). The chemical shift of CDCl3 was 77.20 ppm, and that of DMSO-d6 was 39.51 ppm. HRMS data analysis of the final products was performed using an Agilent 6200 series TOF or 6500 series Q-TOF LC / MS system in positive electrospray ionization (ESI) mode.

[0036] Example 1

[0037] 2α,3β,23-Trihydroxy-ursane-12(13)-ene-28-O-α-L-rhamnopyranoside (Compound 1)

[0038] L-rhamnose (2.00 g, 12.18 mmol) was dissolved in pyridine (40 mL), and benzoyl chloride (7.8 mL, 67.2 mmol) was slowly added dropwise under an ice-water bath. The mixture was then stirred at room temperature for 5 hours, and the reaction was monitored by TLC. After the reaction was complete, ethyl acetate (30 mL) was added to dilute the reaction solution, followed by washing with water (40 mL), 1N hydrochloric acid aqueous solution (100 mL), saturated NaHCO3 aqueous solution (40 mL), and saturated NaCl aqueous solution (40 mL). The organic layer was dried over anhydrous sodium sulfate, concentrated, and purified by silica gel column chromatography (petroleum ether:ethyl acetate = 5:1) to give intermediate 1-1 (4.6 g, white solid, yield 66%).

[0039] Intermediate 1-1 (4.6 g, 7.9 mmol) was dissolved in DCM (40 mL), and 33% hydrobromic acid solution (5 mL) was added under ice-water bath. The reaction was allowed to proceed overnight at room temperature. After the reaction was complete, the reaction solution was washed successively with water (40 mL), saturated NaHCO3 aqueous solution (40 mL), and saturated NaCl aqueous solution (40 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated to give crude compound 1-2.

[0040] The crude compounds 1-2 were dissolved in acetone / water (25:1, 26 mL), and silver carbonate (2.61 g, 9.48 mmol) was added. The mixture was reacted at room temperature for 1 hour. After the reaction was complete, the mixture was filtered through diatomaceous earth, concentrated, and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 3:1) to give intermediate 1-3 (2.3 g, white solid, two-step yield 62%).

[0041] Compounds 1-3 (2.3 g, 4.8 mmol) were dissolved in anhydrous DCM (10 mL), and trichloroacetonitrile (11.0 g, 51.36 mmol) and DBU (387.2 μL, 2.59 mmol) were added. The reaction was carried out at room temperature and monitored by TLC. After the reaction was complete, the solution was concentrated and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 10:1) to give compounds 1-4 (2.4 g, white bubbly solid, yield 85%).

[0042] Melanoic acid (AA) (5.0 g, 10.23 mmol) was dissolved in anhydrous tetrahydrofuran (50 mL), and acetic anhydride (5.8 mL, 61.26 mmol) was slowly added dropwise with stirring. The reaction was carried out at room temperature for 4 hours. After the reaction was complete, the organic phase was concentrated and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 3:1) to give compounds 1-5 (white solid, 5.5 g, yield 87.4%).

[0043] Compounds 1-5 (500 mg, 0.81 mmol) and 1-4 (809 mg, 1.06 mmol) were dissolved in anhydrous DCM (10 mL), and 100 mg of 4A molecular sieve was added. Trimethylsilyl trifluoromethanesulfonate (45 μL, 0.096 mmol) was added under an ice-water bath, and the mixture was stirred at room temperature for 1 hour. After the reaction was complete, the molecular sieve was removed by filtration, the mixture was concentrated, and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 8:1) to give compound 1-6 (white solid, 562 mg, yield 55%). Subsequently, compounds 1-6 were dissolved in a DCM-MeOH mixed solution (1:2, v / v, 15 mL), and sodium methoxide (158 mg, 2.9 mmol) was added. The mixture was stirred at room temperature for 0.5 hours. After the reaction was complete, DOWEX 50WX2-100 cation exchange resin was added, and the mixture was stirred for 15 minutes. The pH was adjusted to 7, the resin was removed by filtration, the filtrate was concentrated, and purified by silica gel column chromatography (DCM:MeOH = 10:1) to give compound 1 (white solid, 302 mg, yield: 82%). 1 H NMR(300MHz, Methanol-d4)δ5.90(d,J=1.5Hz,1H),5.32(s,1H),3.74(d,J=1.8 Hz,1H),3.72–3.63(m,3H),3.48(dd,J=16.4,10.3Hz,2H),3.39–3.33(m,2H),3 .27(d,J=11.1Hz,1H),2.25(d,J=11.4Hz,1H),1.25(d,J=6.1Hz,3H),1.15(s,3 H),1.05(s,3H),0.98(s,3H),0.90(d,J=6.3Hz,3H),0.82(s,3H),0.70(s,3H).13 C NMR(101MHz,DMSO-d6)δ175.13(C-28),138.38(C-13),125.73(C-12),75.99 ,71.89,71.53,70.98,70.11,67.89,64.35,53.05,48.34,47.44,47.35,46.4 8,42.95,42.21,38.99,38.70,37.73,36.82,32.78,30.42,27.72,24.11,23 .79,23.43,21.38,18.36,17.84,17.54,17.38,17.31,14.25.HRMS(ESI):m / z calcd for [M+Na] + C 36 H 58 O9Na:657.3973; found:657.40028.

[0044] Example 2

[0045] 2α,3β,23-Trihydroxy-ursane-12(13)-ene-28-O-β-D-galactopyranoside (Compound 2)

[0046] Following the method of Example 1, L-rhamnose was replaced with D-galactose to prepare compound 2 (white solid, 350 mg, yield: 74%): 1 H NMR(300MHz,Methanol-d4)δ5.32(d,J=8.1Hz,1H),5.25(s,1H),3.89(s,1H) ),3.73-3.61(m,4H),3.58(d,J=6.0Hz,1H),3.53(s,1H),3.49(s,1H),3.35( d,J=9.7Hz,1H),3.27(d,J=11.1Hz,1H),2.24(d,J=11.1Hz,1H),1.13(s,3H) ,1.05(s,3H),0.97(s,3H),0.90(d,J=5.8Hz,3H),0.83(s,3H),0.70(s,3H). 13C NMR(126MHz,DMSO-d6)δ175.56(C-28),138.17(C-13),125.33(C-12),95.05, 76.21,76.06,73.86,69.85,68.06,67.91,64.41,60.18,52.83,47.75,47.52 ,46.53,42.96,42.19,38.83,37.73,36.27,30.54,28.00,24.22,23.74,23.5 1,21.48,17.87,17.48,17.45,17.37,14.26.HRMS(ESI):m / zcalcd.for[M+Na] + C 36 H 58 O 10 Na 673.39222, found 673.39209.

[0047] Example 3

[0048] 2α,3β,23-Trihydroxy-ursane-12(13)-ene-28-O-α-L-arabinopyranoside (Compound 3)

[0049] Following the method of Example 1, L-rhamnose was replaced with L-arabinose to prepare compound 3 (white solid, 219 mg, yield: 83%): 1 H NMR (300MHz, Methanol-d4) δ5.39(d,J=5.3Hz,1H,H-1'),5.29(s,1H),3.99-3.82(m,2H),3.76-3.64(m,2H),3.61-3.43(m,3H),3.37(d,J=9.6H z,1H),3.28(d,J=11.2Hz,1H),2.29(d,J=11.2Hz,1H),1.15(s,3H),1.0 6(s,3H),0.99(s,3H),0.91(d,J=5.9Hz,3H),0.84(s,3H),0.71(s,3H). 13C NMR(101MHz,DMSO-d6)δ175.55(C-28),138.37(C-13),125.33(C-12),94.2 0,76.02,71.70,70.05,67.91,66.04,64.37,64.01,52.66,47.94,47.48,4 6.48,42.95,42.17,38.86,38.76,37.73,36.43,32.65,30.53,28.00,23.9 8,23.78,23.49,21.45,17.86,17.45,17.40,17.33,14.27.HRMS(ESI):m / z calcd.for[M+Na] + C 35 H 56 O9Na 643.38165, found 643.38161.

[0050] Example 4

[0051] 2α,3β,23-Trihydroxy-ursane-12(13)-ene-28-O-β-D-xylanoside (Compound 4)

[0052] Following the method of Example 1, L-rhamnose was replaced with D-xylose to prepare compound 4 (white solid, 305 mg, yield: 65%). 1 H NMR(300MHz,Methanol-d4)δ5.31(d,J=7.2Hz,1H,H-1'),5.25(s,1H),3.86(dd,J =11.4,4.9Hz,1H),3.76-3.61(m,1H),3.60-3.44(m,2H),3.44-3.36(m,1H),3.34 (s,1H),3.29-3.26(m,1H),3.24(d,J=3.9Hz,1H),2.25(d,J=11.2Hz,1H),1.13(s ,3H),1.05(s,4H),0.97(s,4H),0.90(d,J=6.2Hz,4H),0.82(s,3H),0.70(s,3H). 13CNMR(126MHz,DMSO-d6)δ175.15(C-28),137.82(C-13),124.84(C-12),94. 68,75.92,75.59,71.83,69.20,67.45,65.99,63.95,52.24,47.42,47.03, 46.04,42.49,41.74,38.36,38.31,37.27,35.94,32.17,30.06,27.56,23. 62,23.26,23.04,20.98,17.39,16.99,16.95,16.82,13.79.HRMS(ESI):m / z calcd.for[M+Na] + C 35 H 56 O9Na 643.38165,found 643.38237.

[0053] Example 5

[0054] 2α,3β,23-Trihydroxy-ursane-12(13)-ene-28-O-β-D-glucopyranoside (Compound 5)

[0055] Following the method of Example 1, L-rhamnose was replaced with D-glucose to prepare compound 5 (white solid, 310 mg, yield: 65%). 1 H NMR (300MHz, Methanol-d4) δ5.34(d,J=7.9Hz,1H,H-1'),5.26(s,1H),3.80(d,J=11.3Hz,1H),3.75-3.64(m,2H),3.51(d,J=11.1Hz,1H),3.42-3. 32(m,5H),3.27(d,J=11.4Hz,2H),2.24(d,J=11.3Hz,1H),1.13(s,3H),1 .05(s,3H),0.97(s,3H),0.90(d,J=6.1Hz,3H),0.84(s,3H),0.70(s,3H). 13C NMR(126MHz,DMSO-d6)δ175.46(C-28),138.18(C-13),125.33(C-12),94.53,7 9.64,78.06,77.13,76.07,72.77,70.01,67.93,64.42,61.15,47.76,47.53,46 .52,42.96,42.21,38.83,37.74,36.29,32.66,30.53,28.04,24.21,23.70,23. 51,21.48,17.86,17.49,17.44,17.34,14.25.HRMS(ESI):m / zcalcd.for[M+Na] + C 36 H 58 O 10 Na 673.39222, found 673.39181.

[0056] Example 6

[0057] 2-O-α-L-pyranoraminosyl-3β,23-dihydroxy-ursane-12(13)-en-28-acid (compound 6)

[0058] Melanoic acid (AA) (5.0 g, 10.2 mmol) was dissolved in N,N-dimethylformamide (50 mL), and benzyl bromide (1.85 mL, 15.3 mmol) and potassium carbonate (2.8 g, 20.5 mmol) were slowly added dropwise with stirring. The reaction was carried out at room temperature for 6 hours, and the reaction was monitored by TLC. After the reaction was complete, the reaction solution was diluted with ethyl acetate (30 mL), and washed successively with water (100 mL) and saturated brine (50 mL). The organic phase was dried over anhydrous sodium sulfate, concentrated, and purified by silica gel column chromatography (DCM:MeOH = 10:1) to give compound 6-1 (white solid, 3.6 g, yield 61%).

[0059] Compound 6-1 (3.59 g, 6.2 mmol) was dissolved in acetonitrile (50 mL), and benzaldehyde dimethyl acetal (1.13 mL, 7.44 mmol) and p-toluenesulfonic acid (118 mg, 0.62 mmol) were added. The mixture was heated under reflux at 80 °C and stirred for 1 hour. The reaction was monitored by TLC. After the reaction was complete, triethylamine was added dropwise to quench the reaction. The organic phase was dried over anhydrous sodium sulfate, concentrated, and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 3:1) to give compound 6-2 (white solid, 4.7 g, yield: 85%).

[0060] Compound 6-2 (500 mg, 0.868 mmol) and compound 1-4 (864 mg, 1.13 mmol) were dissolved in anhydrous DCM (10 mL), and 100 mg of 4A molecular sieve was added. Trimethylsilyl trifluoromethanesulfonate (30 μL, 0.174 mmol) was added under an ice-water bath, and the mixture was stirred at room temperature for 1 hour. After the reaction was complete, the molecular sieve was removed by filtration, the mixture was concentrated, and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 10:1) to give compound 6-3 (white solid, 985 mg, yield 99%).

[0061] Compound 6-3 was dissolved in a DCM-MeOH mixed solution (1:2, v / v, 9 mL), and 10% Pd / C (98 mg) was added. The mixture was stirred overnight at room temperature under a hydrogen atmosphere. After the reaction was complete, the mixture was filtered through diatomaceous earth, the filtrate was concentrated, and the residue was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 3:1) to give compound 6-4 (white solid, 625 mg, yield: 75%).

[0062] Compound 6-4 was dissolved in a DCM-MeOH mixed solution (1:2, v / v, 15 mL), and sodium methoxide (178 mg, 3.3 mmol) was added. The mixture was stirred at room temperature for 0.5 hours. After the reaction was complete, DOWEX 50WX2-100 cation exchange resin was added, and the mixture was stirred for 15 minutes. The pH was adjusted to 7, the resin was removed by filtration, the filtrate was concentrated, and purified by silica gel column chromatography (DCM:MeOH = 10:1) to obtain compound 6 (white solid, 388 mg, yield: 93%). 1 H NMR (300MHz, DMSO-d6) δ11.94(s,1H),5.14(s,1H),4.69(s,1H),4.61(d,J=3.2Hz,1H),4.5 5(d,J=5.5Hz,1H),4.44(d,2H),4.11(d,J=3.5Hz,1H),3.74-3.66(m,1H),3.63-3.49(m,3H ),3.44-3.36(m,2H),3.20-3.14(m,1H),3.10-3.00(m,1H),2.11(d,J=11.3Hz,1H),1.08(d ,J=2.8Hz,3H),1.05(s,3H),0.92(s,6H),0.82(d,J=6.0Hz,3H),0.75(s,3H),0.56(s,3H). 13C NMR(126MHz,DMSO-d6)δ178.80(C-28),138.71(C-13),124.95(C-12),97.29,73.5 9,73.38,72.69,71.65,70.95,68.70,65.39,64.21,52.88,47.38,47.30,46.23,4 3.29,43.18,42.25,38.98,38.91,37.54,36.79,32.61,30.68,27.98,24.30,23.7 5,23.48,21.56,18.44,17.90,17.47,17.44,17.18,15.64,14.33.HRMS(ESI):m / z calcd.for[M+Na] + C 36 H 58 O9Na 657.3973, found 657.39915.

[0063] Example 7

[0064] 2-O-β-D-glucopyranosyl-3β,23-dihydroxy-ursane-12(13)-en-28-acid (compound 7)

[0065] Following the methods of Examples 1 and 6, L-rhamnose was replaced with D-glucose to prepare compound 7 (white solid, 515 mg, yield: 99%). 1 H NMR (300MHz, DMSO-d6) δ11.94(s,1H),5.16(s,1H),4.94(s,3H),4.58-4.40(m,2H),4. 23(d,J=7.7Hz,1H,H-1'),4.12(s,1H),3.75-3.54(m,2H),3.49-3.36(m,2H),3.31(s,1 H),3.20-3.11(m,2H),3.10-3.00(m,2H),2.96-2.84(m,1H),2.12(d,J=11.4Hz,1H),1 .06(s,3H),0.93(s,3H),0.92(s,3H),0.82(d,J=6.1Hz,3H),0.75(s,3H),0.56(s,3H). 13C NMR(126MHz,DMSO-d6)δ178.78(C-28),138.74(C-13),124.93(C-12),103.13,8 0.31,77.14,76.94,73.86,73.63,70.46,65.39,63.97,61.46,52.90,47.41,47. 32,45.91,45.00,43.12,42.27,38.97,38.90,37.64,36.79,32.62,30.68,27.97 ,23.78,23.54,21.55,17.71,17.45,17.40,17.21,15.64,14.54.HRMS(ESI):m / z calcd.for[M+Na] + C 36 H 58 O 10 Na 673.39222, found 673.39107.

[0066] Example 8

[0067] 2-O-α-L-pyranarabinoyl-3β,23-dihydroxy-ursane-12(13)-en-28-acid (compound 8)

[0068] Following the methods of Examples 1 and 6, L-rhamnose was replaced with L-arabinose to prepare compound 8 (white solid, 215 mg, yield: 92%). 1 H NMR(300MHz,DMSO-d6)δ11.93(s,1H),5.14(s,1H),4.89(s,1H),4.64(s,1H), 4.57-4.42(m,2H),4.21(d,J=4.4Hz,1H,H-1'),4.05(s,1H),3.77-3.52(m,3H) ,3.49-3.34(m,4H),3.28-3.21(m,1H),3.11-2.98(m,1H),2.11(d,J=11.3Hz,1 H),1.05(s,3H),0.92(s,6H),0.81(d,J=6.2Hz,3H),0.74(s,3H),0.55(s,3H). 13C NMR(101MHz,DMSO-d6)δ178.95(C-28),138.79(C-13),124.85(C-12),101.9 0,77.32,73.48,72.97,70.90,67.81,65.63,64.00,52.90,47.42,47.31,46. 00,44.33,43.03,42.26,38.98,38.90,37.68,36.81,32.63,30.69,27.97,24 .31,23.77,23.53,21.56,17.72,17.48,17.44,17.24,14.38.HRMS(ESI):m / z calcd.for[M+Na] + C 35 H 56 O9Na643.38165, found 643.38202.

[0069] Example 9

[0070] 2-O-β-D-galactopyranosyl-3β,23-dihydroxy-ursane-12(13)-en-28-acid (compound 9)

[0071] Following the methods of Examples 1 and 6, L-rhamnose was replaced with D-galactose to prepare compound 9 (white solid, 76 mg, yield: 42%). 1 H NMR(300MHz,DMSO-d6)δ11.94(s,1H),5.15(s,1H),4.80(s,1H),4.73(s,1H) ,4.60(s,1H),4.51-4.38(m,2H),4.18(d,J=6.4Hz,1H,H-1'),4.14(s,1H),3. 68-3.38(m,6H),3.30-3.25(m,3H),3.13-2.99(m,1H),2.11(d,J=10.9Hz,1H ),1.05(s,3H),0.92(s,6H),0.82(d,J=5.5Hz,3H),0.75(s,3H),0.55(s,3H). 13C NMR(126MHz,DMSO-d6)δ178.81(C-28),138.75(C-13),124.91(C-12),103.77, 80.22,75.66,73.75,73.63,71.05,68.53,63.95,60.74,52.90,47.42,47.32, 45.92,45.07,43.10,42.27,38.97,38.90,37.66,36.79,32.63,30.68,27.96, 24.30,23.78,23.54,21.55,17.70,17.44,17.41,17.22,14.51.HRMS(ESI):m / z calcd.for[M+Na] + C 36 H 58 O 10 Na 673.39222, found 673.39142.

[0072] Example 10

[0073] 2-O-β-D-xylanopyranosyl-3β,23-dihydroxy-ursane-12(13)-en-28-acid (compound 10)

[0074] Following the methods of Examples 1 and 6, L-rhamnose was replaced with D-xylose to prepare compound 10 (white solid, 318 mg, yield: 74%). 1 H NMR (300MHz, DMSO-d6) δ11.92(s,1H),5.14(s,1H),4.97(d,J=4.2Hz,3H),4.46(t,J=5.0Hz, 1H),4.20(d,J=7.5Hz,1H,H-1'),3.93(s,1H),3.70(dd,J=11.0,4.9Hz,1H),3.60(s,1H),3.4 2-3.35(m,1H),3.29(s,2H),3.08(dd,J=14.4,7.0Hz,3H),2.99-2.89(m,1H),2.11(d,J=11.1 Hz,1H),1.05(d,J=4.9Hz,3H),0.92(s,6H),0.81(d,J=6.0Hz,3H),0.74(s,3H),0.55(s,3H). 13C NMR(101MHz,DMSO-d6)δ178.77(C-28),138.76(C-13),124.90(C-12),10 2.82,76.85,73.48,73.33,66.18,63.95,52.88,47.40,47.31,45.93,44. 60,43.05,42.25,38.97,38.88,37.67,36.78,32.61,30.67,27.96,24.28 ,23.78,23.54,21.55,17.71,17.48,17.40,17.22,14.40.HRMS(ESI):m / z calcd.for[M+Na] + C 35 H 56 O9Na 643.38165, found 643.38269.

[0075] Example 11

[0076] 2-O-α-L-pyranoraminosyl-3β,23-dihydroxy-ursane-12(13)-ene-28-O-β-D-glucopyranoside

[0077] (Compound 11)

[0078] Following the methods of Examples 1 and 6, L-rhamnose was replaced with D-glucose to obtain compound 11-2 (white frothy solid, 272 mg, yield: 64%). Compound 11-2 (272 mg, 0.4 mmol) and compound 6-4 (300 mg, 0.32 mmol) were dissolved in dichloromethane:water (33 mL, 10:1). Potassium carbonate (88 mg, 0.64 mmol) and tetrabutylammonium bromide (41 mg, 0.13 mmol) were added under stirring. The reaction was carried out at room temperature for 24 hours under argon protection, and the reaction was monitored by TLC. After the reaction was complete, the reaction solution was washed successively with water (20 mL) and saturated NaCl aqueous solution (20 mL). The organic layer was dried over anhydrous sodium sulfate, concentrated, and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 1:1) to obtain compound 11-3 (white solid, 327 mg, yield: 68%).

[0079] Following the method of Example 1, compounds 1-6 were replaced with compounds 11-3, and compound 11 (white solid, 151 mg, yield: 88%) was obtained according to the method of Example 1: 1H NMR(300MHz, Methanol-d4)δ5.32(d,J=7.9Hz,1H,H-1”),5.24(s,1H),4.82( s,1H,H-1'),3.90-3.62(m,6H),3.55-3.45(m,3H),3.39-3.31(m,4H),3.24( d,J=11.2Hz,2H),2.22(d,J=11.1Hz,1H),1.24(d,J=6.1Hz,3H),1.11(s,3H) ,1.02(s,3H),0.95(s,3H),0.88(d,J=6.2Hz,3H),0.82(s,3H),0.68(s,3H). 13 C NMR (126MHz, DMSO-d6) δ174.97,137.65,124.83,96.85,94.05,77.60,76.66,73.1 6,72.91,72.30,72.20,71.18,70.46,69.53,68.24,63.71,60.66,52.40,47.28,4 6.98,45.78,42.81,41.75,38.38,38.33,37.03,35.79,32.13,30.06,27.57,23.7 4,23.19,23.05,20.99,17.97,17.38,16.97,16.85,16.80,13.89.HRMS(ESI):m / z calcd.for[M+Na] + C 42 H 68 O 14 Na 819.45013, found 819.44993.

[0080] Example 12

[0081] 2-O-α-L-arabinopyranosyl-3β,23-dihydroxy-ursane-12(13)-ene-28-O-β-D-glucopyranoside (compound 12)

[0082] Following the method of Example 11, L-rhamnose was replaced with L-arabinose to obtain compound 12 (white solid, 151 mg, yield: 88%). 1H NMR(300MHz,Methanol-d4)δ5.34(d,J=7.8Hz,1H,H-1”),5.26(s,1H),4.28(d,J=5.3Hz,1H, H-1'),3.89(d,J=12.3Hz,1H),3.85-3.74(m,3H),3.68(dd,J=11.6,3.8Hz,1H),3.59(d,J=12 .4Hz,1H),3.56-3.43(m,5H),3.42-3.33(m,3H),3.27(d,J=11.6Hz,1H),2.24(d,J=11.2Hz, 1H),1.13(s,3H),1.04(s,3H),0.97(s,3H),0.89(d,J=6.1Hz,3H),0.84(s,3H),0.70(s,3H). 13 C NMR(126MHz,DMSO-d6)δ175.46(C-28),138.18(C-13),125.25(C-12),101.87,94.52, 78.06,77.27,77.12,73.50,72.97,72.76,70.89,69.97,67.82,65.65,61.11,52.89, 47.76,47.49,46.05,44.42,43.04,42.23,38.85,38.79,37.65,36.26,32.61,30.53, 28.03,24.20,23.69,23.58,21.46,17.68,17.45,17.32,17.29,14.39.HRMS(ESI):m / z calcd.for[M+Na] + C 41 H 66 O 14 Na 805.43448, found 805.43420.

[0083] Example 13

[0084] 2-O-β-D-glucopyranosyl-3β,23-dihydroxy-ursane-12(13)-ene-28-O-β-D-glucopyranoside (compound 13)

[0085] Following the method of Example 11, L-rhamnose was replaced with D-glucose to obtain compound 13 (white solid, 301 mg, yield: 99%). 1H NMR(300MHz, Methanol-d4)δ5.34(d,J=7.9Hz,1H,H-1”),5.26(s,1H),4.34(d,J=7.7Hz, 1H,H-1'),3.90-3.74(m,3H),3.72-3.59(m,2H),3.58-3.44(m,4H),3.35(d,J=7.1Hz,5H) ,3.26(d,J=4.0Hz,1H),3.22-3.13(m,1H),2.24(d,J=11.2Hz,1H),1.18(t,J=7.0Hz,3H) ,1.13(s,3H),1.04(s,3H),0.97(s,3H),0.89(d,J=6.3Hz,3H),0.84(s,3H),0.71(s,3H). 13 C NMR(126MHz,DMSO-d6)δ175.46(C-28),138.15(C-13),125.28(C-12),103.08,94.5 3,80.22,78.07,77.14,76.93,73.85,73.65,72.77,70.46,69.98,63.95,61.45,61. 12,52.89,47.77,47.49,45.09,43.12,42.25,38.85,38.81,37.60,36.27,32.62,30 .54,28.03,24.22,23.69,23.60,21.47,17.67,17.43,17.28,14.56.HRMS(ESI):m / z calcd.for[M+Na] + C 42 H 68 O 15 Na 835.44504, found 835.44539.

[0086] Example 14

[0087] 2-O-β-D-xylanopyranosyl-3β,23-dihydroxy-ursane-12(13)-ene-28-O-β-D-glucopyranoside (compound 14)

[0088] Following the method of Example 11, L-rhamnose was replaced with D-xylose to obtain compound 14 (white solid, 772 mg, yield: 93%). 1H NMR (300MHz, Methanol-d4) δ5.34 (d, J = 7.9 Hz, 1H, H-1”), 5.26 (s, 1H), 4.31 (d, J = 7. 5Hz,1H,H-1'),3.88(dd,J=11.2,5.3Hz,1H),3.83-3.77(m,2H),3.71-3.65(m,1H),3 .55-3.44(m,4H),3.42-3.33(m,4H),3.29-3.15(m,3H),2.24(d,J=10.9Hz,1H),1.1 3(s,3H),1.04(s,3H),0.97(s,3H),0.89(d,J=6.2Hz,3H),0.84(s,3H),0.69(s,3H). 13 C NMR(126MHz,DMSO-d6)δ175.45(C-28),138.18(C-13),125.25(C-12),102.75,94.5 2,78.30,78.06,77.13,76.86,73.47,73.36,72.76,69.97,66.18,63.97,61.13,52. 89,47.77,47.49,45.99,44.69,43.06,42.24,38.85,38.80,37.65,36.26,32.61,30 .54,28.04,24.21,23.70,23.60,21.47,17.67,17.45,17.30,14.41.HRMS(ESI):m / z calcd.for[M+Na] + C 41 H 66 O 14 Na 805.43448, found 805.43498.

[0089] Example 15

[0090] 2-O-β-D-galactopyranosyl-3β,23-dihydroxy-ursane-12(13)-ene-28-O-β-D-glucopyranoside (compound 15)

[0091] Following the method of Example 11, L-rhamnose was replaced with D-galactose to obtain compound 15 (white solid, 150 mg, yield: 86%). 1H NMR(300MHz, Methanol-d4)δ5.32(d,J=7.9Hz,1H,H-1”),5.24(s,1H),4.28(d,J=5.7Hz,1H,H-1’),3.90-3.72(m,5H),3.70-3.62(m,2H),3.57-3.44(m ,6H),3.39-3.32(m,2H),3.23(d,1H),2.22(d,J=12.0Hz,1H),1.11(s,3H), 1.02(s,3H),0.95(s,3H),0.87(d,J=6.0Hz,3H),0.82(s,3H),0.68(s,3H). 13 C NMR(126MHz,DMSO-d6)δ174.97(C-23),137.70(C-13),124.80(C-12),103.33,94.05,79. 81,77.61,76.67,75.20,73.25,73.19,72.30,70.56,69.52,68.09,63.46,60.66,60.31, 52.42,47.30,47.03,45.47,44.70,42.63,41.78,38.38,38.33,37.15,35.79,32.15,30. 07,27.57,23.74,23.23,23.12,21.00,17.18,16.88(d,J=16.6Hz),14.08.HRMS(ESI):m / z calcd.for[M+Na] + C 42 H 68 O 15 Na 835.44504, found 835.44575.

[0092] Example 16

[0093] 2-O-α-L-pyranoraminosyl-3β,23-dihydroxy-ursane-12(13)-ene-28-O-α-L-arabinopyranoside (compound 16)

[0094] Following the method of Example 11, D-glucose was replaced with L-arabinose to obtain compound 16 (white solid, 117 mg, yield: 73%). 1H NMR(300MHz,Methanol-d4)δ5.41(d,J=5.6Hz,1H),5.31(s,1H),3.96-3.81 (m,4H),3.85-3.62(m,5H),3.64-3.47(m,4H),3.41(t,J=9.2Hz,2H),3.27(d ,J=10.3Hz,1H),2.30(d,J=11.2Hz,1H),1.28(d,J=6.2Hz,3H),1.16(s,3H) ,1.06(s,3H),1.00(s,3H),0.92(d,J=6.2Hz,3H),0.85(s,3H),0.73(s,3H). 13 C NMR (126MHz, DMSO-d6) δ175.55,138.32,125.30,97.32,94.22,73.60,73.36,72 .69,71.73,71.64,70.95,70.05,68.70,66.06,64.17,64.04,52.70,47.95,47.4 0,46.24,43.29,42.20,38.87,38.76,37.50,36.42,32.60,30.55,28.02,23.98 ,23.73,23.52,21.44,18.45,17.85,17.41,17.31,17.23,14.36.HRMS(ESI):m / z calcd.for[M+Na] + C 41 H 66 O 13 Na 789.43956, found 789.43956.

[0095] Example 17

[0096] 2-O-β-D-galactopyranosyl-3β,23-dihydroxy-ursane-12(13)-ene-28-O-α-L-arabinopyranoside (Compound 17)

[0097] Following the method of Example 15, D-glucose was replaced with L-arabinose to obtain compound 17 (white solid, 220 mg, yield: 88%). 1H NMR(300MHz, Methanol-d4)δ5.40(d,J=5.3Hz,1H,H-1”),5.30(s,1H),4.33(d,J=6.1Hz,1H,H-1’),3.93-3.64(m,8H),3.63-3.48(m,6H) ,3.27(d,J=11.7Hz,1H),2.30(d,J=11.0Hz,1H),1.16(s,3H),1.05(s,3H),0.99(s,3H),0.91(d,J=6.2Hz,3H),0.85(s,3H),0.73(s,3H). 13 C NMR(126MHz,DMSO-d6)δ175.55,138.37,125.26,103.72,94.22,80.12,75.66,7 3.74,73.64,71.73,71.03,70.05,68.53,66.06,64.05,63.93,60.74,52.71,47. 96,47.45,45.94,45.12,43.09,42.22,38.86,38.76,37.62,36.42,32.63,30.55 ,28.01,23.99,23.77,23.58,21.44,17.65,17.39,17.26,14.53.HRMS(ESI):m / z calcd.for[M+Na] + C 41 H 66 O 14 Na 805.43448, found 805.43531.

[0098] Example 18

[0099] 2-O-β-D-xylanopyranosyl-3β,23-dihydroxy-ursane-12(13)-ene-28-O-α-L-arabinopyranoside (compound 18)

[0100] Following the method of Example 14, D-glucose was replaced with L-arabinose to obtain compound 18 (white solid, 200 mg, yield: 64%). 1H NMR (300MHz, Methanol-d4) δ5.37(d,J=5.5Hz,1H,H-1”),5.27(s,1H),4.30(d,J=7.6Hz,1H,H-1’),3.93-3.45(m,10H),3.35(s,1H) ,3.28-3.13(m,3H),2.27(d,J=11.2Hz,1H),1.13(s,3H),1.03(s,3H),0.96(s,3H),0.89(d,J=6.1Hz,3H),0.82(s,3H),0.69(s,3H). 13 C NMR(126MHz,DMSO-d6)δ175.07,137.92,124.77,102.29,93.75,77.83,76.40,7 3.02,72.87,71.25,69.58,69.50,65.72,65.58,63.57,63.49,52.25,47.49,46. 96,45.50,44.18,42.59,41.75,38.40,38.28,37.18,35.95,32.14,30.08,27.55 ,23.51,23.30,23.12,20.97,17.20,16.95,16.82,16.79,13.95.HRMS(ESI):m / z calcd.for[M+Na] + C 40 H 64 O 13 Na 775.42391, found 775.42372.

[0101] Example 19

[0102] 2-O-α-L-arabinopyranosyl-3β,23-dihydroxy-ursane-12(13)-ene-28-O-α-L-arabinopyranoside (compound 19)

[0103] Following the method of Example 12, D-glucose was replaced with L-arabinose to obtain compound 19 (white solid, 200 mg, yield: 64%). 1H NMR (300MHz, Methanol-d4) δ5.37(d,J=5.7Hz,1H,H-1”),5.28(s,1H),4.28(d,J=6.6Hz,1H,H-1’),3.94-3.46(m,13H),3.27(d, J=11.3Hz,1H),2.27(d,J=11.3Hz,1H),1.13(s,3H),1.03(s,3H),0.97(s,3H),0.89(d,J=6.3Hz,3H),0.83(s,3H),0.69(s,3H). 13 C NMR(126MHz,DMSO-d6)δ175.09,137.92,124.78,101.43,93.76,76.80,73.0 2,72.53,71.27,70.44,69.59,67.37,65.60,65.20,63.57,52.25,47.50,46. 98,45.57,43.94,42.58,41.75,38.40,38.30,37.20,35.96,32.16,30.09,27 .55,23.52,23.30,23.12,20.98,17.22,16.96,16.82,13.94.HRMS(ESI):m / z calcd.for[M+Na] + C 40 H 64 O 13 Na 775.42391, found 775.42422.

[0104] Example 20

[0105] 2-O-β-D-glucopyranosyl-3β,23-dihydroxy-ursane-12(13)-ene-28-O-α-L-arabinopyranoside (Compound 20)

[0106] Following the method of Example 13, D-glucose was replaced with L-arabinose to obtain compound 20 (white solid, 260 mg, yield: 74%). 1H NMR(300MHz, Methanol-d4)δ5.39(d,J=5.5Hz,1H,H-1”),5.29(s,1H),4.35(d,J=7.8Hz,1H,H-1’),3.96-3.79(m,5H),3.72-3.47(m,6H),3.45- 3.35(m,2H),3.30-3.11(m,2H),2.28(d,J=11.6Hz,1H),1.15(s,3H),1. 04(s,3H),0.98(s,3H),0.90(d,J=6.2Hz,3H),0.84(s,3H),0.72(s,3H). 13 C NMR (126MHz, DMSO-d6) δ175.10,137.90,124.82,102.61,93.77,79.70,76.69,76.48,73. 39,73.18,71.28,69.99,69.59,65.61,63.60,63.50,60.98,52.26,47.51,46.98,45.48, 44.59,42.67,41.81,41.77,38.41,38.31,37.15,35.97,32.17,30.10,27.55,23.54,23. 31,23.13,21.10,20.99,17.22,17.00,16.94,16.82,16.79,16.75,14.11.HRMS(ESI):m / z calcd.for[M+Na] + C 41 H 66 O 14 Na805.43448, found 805.43533.

[0107] Example 21

[0108] 2-O-α-L-pyranoramose-(1→6)-β-D-pyranoglucopyranosyl-3β,23-dihydroxy-ursane-12(13)-en-28-acid (compound 21)

[0109] D-glucose (20 g, 111.02 mmol), sodium acetate (18 g, 133.22 mmol), and DMAP (1.36 g, 11.1 mmol) were added to 150 mL of acetic anhydride, and the mixture was stirred at 60 °C for 6 hours. After the reaction was complete, the solution was poured into ice water and stirred for 10 minutes. After the white solid was completely precipitated, the mixture was filtered and the filter cake was dried to obtain crude acetylated glucose 21-1. The crude product was dissolved in anhydrous DCM (120 mL), and boron trifluoride ether (7.76 mL, 133.22 mmol) and thiophenol (6.32 mL, 133.22 mmol) were added. The mixture was stirred for more than 48 hours. After the reaction was complete, 100 mL of saturated NaHCO3 aqueous solution was added to quench the reaction. The mixture was separated, and the organic layer was washed successively with water (500 mL) and saturated NaCl aqueous solution (500 mL). The mixture was dried over anhydrous sodium sulfate, concentrated, and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 4:1) to obtain intermediate 21-2. Intermediate 21-2 was dissolved in a DCM-MeOH mixed solution (1:2, v / v, 300 mL), sodium methoxide (12 g, 555.1 mmol) was added, and the mixture was stirred for 0.5 hours. After the reaction was complete, cation exchange resin was added to adjust the pH to 7, and the mixture was filtered. The filtrate was concentrated and purified by silica gel column chromatography (DCM:MeOH = 10:1) to obtain intermediate 21-3 (8.5 g, white solid, yield: 28%).

[0110] Intermediate 21-3 (1.90 g, 6.99 mmol) was dissolved in anhydrous pyridine (50 mL), and tert-butyldimethylchlorosilane (1.26 g, 8.39 mmol) and DMAP (0.09 g, 0.74 mmol) were added. The mixture was stirred at 70 °C for 24 hours. After the reaction was complete, it was cooled to room temperature. Intermediate 21-4 was generated during the reaction. No further treatment was required. Acetic anhydride (5 mL, 27.96 mmol) was added directly, and the mixture was stirred for 5 hours. After the reaction was complete, water (50 mL) was added to quench the reaction. DCM (50 mL) was added to separate the layers. The organic layer was washed successively with water (50 × 2 mL), 1N hydrochloric acid aqueous solution until the solution was weakly acidic, washed with saturated NaHCO3 aqueous solution until neutral, and then washed with saturated NaCl aqueous solution (50 mL). The solution was dried over anhydrous sodium sulfate, filtered, and concentrated to obtain crude intermediate 21-5. Intermediate 21-5 was dissolved in anhydrous DCM (50 mL), and boron trifluoride diethyl ether (1.75 mL, 13.98 mmol) was added. The mixture was stirred for half an hour. After the reaction was complete, saturated NaHCO3 aqueous solution (2 mL) was added to quench the reaction. The organic layer was washed once with saturated NaCl aqueous solution (20 mL), concentrated, and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 2:1) to give compound 21-6 (2.64 g, white solid, yield 95%).

[0111] Intermediate 21-6 (400 mg, 1.0 mmol) and compound 1-4 (745 mg, 1.2 mmol) were dissolved in anhydrous DCM (20 mL), and 4A molecular sieve (400 mg) was added. Trimethylsilyl trifluoromethanesulfonate (44.5 μL, 0.2 mmol) was added under ice-water bath conditions. The mixture was reacted at room temperature for half an hour. After the reaction was complete, the molecular sieve was removed by filtration, the mixture was concentrated, and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 4:1) to give compound 21-7 (white bubbly solid, 700 mg, yield: 82%).

[0112] Compound 21-7 was dissolved in an acetone-water mixture (15 mL, 10:1 v / v), and trichloroisocyanuric acid (189.85 mg, 0.82 mmol) was added. The mixture was stirred at room temperature for 1 hour. After the reaction was complete, the acetone was evaporated, and DCM (10 mL) was added. The mixture was separated, and the organic layer was washed with saturated NaHCO3 aqueous solution (20 mL) and saturated NaCl aqueous solution (20 mL), respectively. The mixture was concentrated and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 1:1) to give intermediate 21-8 (white bubbly solid, 530 mg, yield: 81%).

[0113] Intermediate 21-8 (530 mg, 0.69 mmol) was dissolved in DCM, and trichloroacetonitrile (744 μL, 7.42 mmol) and DBU (56 μL, 0.37 mmol) were added. The mixture was stirred at room temperature for 1 hour. After the reaction was complete, the solvent was evaporated, and the solution was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 4:1) to give compound 21-9 (white solid, 560 mg, yield: 89%).

[0114] Following the method of Example 6, intermediates 1-4 were replaced with 21-9 to obtain compound 21 (white solid, 150 mg, yield: 79%). 1H NMR (300MHz, DMSO-d6) δ11.94(s,1H),5.15(s,1H),5.09(d,J=5.3Hz,1H),5.03(d,J=4.7Hz,1H),4.98(d, J=4.6Hz,1H),4.67-4.60(m,2H),4.53(s,1H),4.45-4.33(m,2H),4.24(d,J=7.6Hz,1H,H-1'),4.13(s,1H ),3.81(d,J=8.7Hz,1H),3.69-3.37(m,6H),3.33-3.26(m,1H),3.23-2.89(m,6H),2.11(d,J=11.4Hz,1H) ,1.12(d,J=6.2Hz,3H),1.05(s,3H),0.95-0.89(m,6H),0.81(d,J=6.1Hz,3H),0.75(s,3H),0.56(s,3H). 13 C NMR(126MHz,DMSO-d6)δ178.76,138.69,124.96,102.92,101.34,80.10,76.88,75 .42,73.71,73.51,72.60,71.18,70.69,68.81,67.64,63.94,52.88,47.42,47.32, 45.85,45.05,43.17,42.26,38.97,38.89,37.63,36.78,32.62,30.67,27.96,24. 29,23.78,23.53,21.55,18.38,17.71,17.44,17.40,17.21,14.46.HRMS(ESI):m / z calcd.for[M+Na] + C 42 H 68 O 14 Na 819.45013, found 819.45153.

[0115] Example 22

[0116] 2-O-β-D-xylanopyranosyl-(1→6)-β-D-glucopyranosyl-3β,23-dihydroxy-ursane-12(13)-en-28-acid (compound 22)

[0117] Following the methods of Examples 1 and 21, L-rhamnose in Example 1 was replaced with D-xylose, and then, following the method of Example 21, compound 22 (white solid, 98 mg, yield: 77%) was obtained. 1H NMR (300MHz, DMSO-d6) δ11.95(s,1H),5.28-4.89(m,7H),4.76(d,J=1.9Hz,1H),4.46(t,J=4.9H z,1H),4.31(d,J=7.5Hz,1H,H-1′),4.13(d,J=7.4Hz,1H,H-1”),4.06(d,J=9.0Hz,1H),3.76-3. 57(m,2H),3.50-3.34(m,3H),3.30-3.21(m,2H),3.19-3.08(m,2H),3.09-2.92(m,5H),2.12(d, J=11.4Hz,1H),1.06(s,3H),0.97-0.89(m,6H),0.82(d,J=5.8Hz,3H),0.76(s,3H),0.57(s,3H). 13 C NMR(126MHz,DMSO-d6)δ178.30,138.25,124.46,103.62,103.10,81.47,76.39,7 5.98,74.77,73.78,73.51,73.23,70.53,69.58,68.81,65.64,63.26,52.43,46. 88,45.18,44.88,42.67,41.80,38.50,38.43,37.11,36.31,32.12,30.20,27.50 ,23.82,23.29,23.07,21.09,17.20,16.95,16.91,16.66,14.16.HRMS(ESI):m / z calcd.for[M+Na] + C 41 H 66 O 14 Na 805.43448, found 805.43359.

[0118] Example 23

[0119] 2-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl-3β,23-dihydroxy-ursane-12(13)-en-28-acid (compound 23)

[0120] Following the methods of Examples 1 and 21, L-rhamnose in Example 1 was replaced with D-glucose, and then, following the method of Example 21, compound 23 (white solid, 115 mg, yield: 57%) was obtained: 1H NMR (300MHz, DMSO-d6) δ11.93(s,1H),5.22-5.12(m,3H),5.08(d,J=4.7Hz,1H),5.04(d,J=4.5Hz,1H) ,4.97-4.90(m,2H),4.76(d,J=2.1Hz,1H),4.53(t,J=5.8Hz,1H),4.46(t,J=4.9Hz,1H),4.32(d,J=7. 7Hz,1H,H-1'),4.15(d,J=7.7Hz,2H),3.76-3.57(m,2H),3.49-3.35(m,4H),3.23-2.89(m,9H),2.12( d,J=11.1Hz,1H),1.06(s,3H),0.94(d,J=7.0Hz,6H),0.82(d,J=5.8Hz,3H),0.76(s,3H),0.57(s,3H). 13 C NMR(126MHz,DMSO-d6)δ178.29,138.25,124.45,103.24,102.87,81.76,76.82,76 .44,76.04,74.69,73.93,73.57,73.26,70.57,70.02,68.90,63.21,60.95,52.43, 46.86,45.14,44.97,42.67,41.80,38.49,38.43,37.09,36.30,32.11,30.20,27. 49,23.82,23.29,23.06,21.08,17.18,16.94,16.90,16.66,14.16.HRMS(ESI):m / z calcd.for[M+Na] + C 42 H 68 O 15 Na 835.44504, found 835.44540.

[0121] Example 24

[0122] 2-O-α-L-Pyranoarabinosyl-(1→6)-β-D-Pyranoglucopyranosyl-3β,23-dihydroxy-ursane-12(13)-en-28-acid (compound 24)

[0123] Following the methods of Examples 1 and 21, L-rhamnose in Example 1 was replaced with L-arabinose, and then, following the method of Example 21, compound 24 (white solid, 165 mg, yield: 88%) was obtained: 1H NMR (300MHz, DMSO-d6) δ11.93 (s, 1H), 5.42-4.84 (m, 4H), 4.72 (d, J = 3.6Hz, 1H), 4.66-4.58(m,2H),4.54-4.40(m,3H),4.30(d,J=7.7Hz,1H,H-1'),4.19(d,J=5. 7Hz,1H),4.00(s,1H),3.80-3.35(m,9H),3.25-2.85(m,4H),2.11(d,J=11.2Hz, 1H),1.04(s,3H),0.92(s,6H),0.81(d,J=5.7Hz,3H),0.74(s,3H),0.55(s,3H). 13 C NMR (126MHz, DMSO-d6) δ178.30,138.27,124.46,103.53,102.16,79.58,78.99,74.78,74 .42,73.27,73.08,72.71,70.61,67.83,66.29,63.50,60.10,52.42,46.95,46.86,45.45, 44.40,42.66,41.80,40.11,39.94,39.77,39.09,38.50,38.43,37.19,36.32,32.15,30. 21,27.50,23.83,23.32,23.07,21.09,17.25,16.99,16.94,16.74,14.04.HRMS(ESI):m / z calcd.for[M+Na] + C 41 H 66 O 14 Na 805.43448, found 805.43443.

[0124] Example 25

[0125] 2-O-β-D-galactopyranose-(1→6)-β-D-glucopyranose-3β,23-dihydroxy-ursane-12(13)-en-28-acid (compound 25)

[0126] Following the methods of Examples 1 and 21, L-rhamnose in Example 1 was replaced with D-galactose, and then, following the method of Example 21, compound 25 (white solid, 351 mg, yield: 91%) was obtained: 1H NMR (300MHz, DMSO-d6) δ11.94(s,1H),5.23-5.11(m,3H),5.08(d,J=4.8Hz,1H),5.03(d,J=4.7 Hz,1H),4.70(d,J=4.2Hz,1H),4.64(s,1H),4.62-4.56(m,1H),4.46(d,J=5.7Hz,1H,H-1'),4. 32(t,J=6.1Hz,2H),4.12(t,J=9.0Hz,2H),3.71-3.37(m,9H),3.28-2.87(m,6H),2.12(d,J=11 .1Hz,1H),1.06(s,3H),0.93(d,J=7.8Hz,6H),0.82(d,J=6.1Hz,3H),0.76(s,3H),0.57(s,3H). 13 C NMR (126MHz, DMSO-d6) δ178.30,138.26,124.46,103.47,103.33,81.86,76.46,75.24,74 .77,73.57,73.26,72.78,71.14,70.60,68.76,68.24,63.23,60.48,52.44,46.90,46.86, 45.15,45.00,42.69,41.81,40.10,39.94,39.78,38.50,38.44,37.10,36.31,32.12,30. 21,27.50,23.83,23.30,23.08,21.09,17.19,16.95,16.91,16.67,14.15.HRMS(ESI):m / z calcd.for[M+Na] + C 42 H 68 O 15 Na 835.44504, found 835.44383.

[0127] Example 26

[0128] 2α,3β,23-Trihydroxy-ursane-12(13)-ene-28-O-α-L-rhamnopyranosyl-(1→6)-β-D-glucopyranoside (Compound 26)

[0129] Following the method of Example 1, intermediates 1-4 were replaced with 21-9 to obtain compound 26 (white solid, 190 mg, yield: 76%). 1H NMR(300MHz,Methanol-d4)δ5.32(d,J=7.8Hz,1H,H-1'),5.25(s,1H),4.70(s, 1H,H-1”),3.92(d,J=11.0Hz,1H),3.82(s,1H),3.78-3.58(m,4H),3.55-3.34(m ,6H),3.29-3.19(m,2H),2.24(d,J=10.8Hz,1H),1.24(d,J=5.9Hz,3H),1.13(s, 3H),1.05(s,3H),0.97(s,3H),0.90(d,J=5.3Hz,3H),0.83(s,3H),0.69(s,3H). 13 C NMR (126MHz, DMSO-d6) δ175.06,137.75,124.87,100.41,93.87,76.56,76.18,75.59, 72.24,71.90,70.66,70.41,69.62,68.27,67.47,66.15,63.94,52.24,47.37,47.05, 46.06,42.48,41.71,40.09,39.95,39.78,38.33,38.29,37.25,35.80,32.19,30.06, 27.58,23.63,23.27,23.06,21.00,17.92,17.39,16.98,16.82,13.80.HRMS(ESI):m / z calcd.for[M+Na] + C 42 H 68 O 14 Na 819.45013, found 819.45059.

[0130] Example 27

[0131] 2α,3β,23-Trihydroxy-ursane-12(13)-ene-28-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranoside (Compound 27)

[0132] Following the methods of Examples 1 and 21, intermediate 27-1 was obtained. Then, following the method of Example 1, intermediate 1-4 was replaced with 27-1 to obtain compound 27 (white solid, 140 mg, yield: 82%). 1H NMR(300MHz, Methanol-d4)δ5.30(d,J=7.8Hz,1H,H-1'),5.24(s,1H),4.33(d,J=7.8 Hz,1H,H-1”),4.11(d,J=10.8Hz,1H),3.85(d,J=11.0Hz,1H),3.79-3.62(m,3H),3.5 4-3.33(m,7H),3.29-3.22(m,3H),3.19(d,J=8.5Hz,1H),2.24(d,J=11.3Hz,1H),1.1 3(s,3H),1.06(s,3H),0.97(s,3H),0.90(d,J=6.1Hz,3H),0.84(s,3H),0.70(s,3H). 13 C NMR(101MHz,DMSO-d6)δ175.51(C-28),138.24(C-13),125.27(C-12),103.53, 94.50,77.29,77.18,77.04,76.99,76.05,73.97,72.62,70.44,67.94,64.40, 61.49,52.73,47.80,47.52,46.50,42.95,42.21,38.79,37.73,36.35,30.52, 28.10,24.18,23.67,23.51,21.49,17.86,17.48,17.36,14.27.HRMS(ESI):m / z calcd.for[M+Na] + C 42 H 68 O 15 Na 835.44504, found 835.44464.

[0133] Example 28

[0134] 2α,3β,23-Trihydroxy-ursane-12(13)-ene-28-O-β-D-galactopyranosyl-(1→6)-β-D-glucopyranoside (compound 28)

[0135] Following the methods of Examples 1 and 21, intermediate 28-1 was obtained. Then, following the method of Example 1, intermediate 1-4 was replaced with 28-1 to obtain compound 28 (white solid, 191 mg, yield: 78%). 1H NMR(300MHz, Methanol-d4)δ5.31(d,J=7.9Hz,1H,H-1'),5.25(s,1H),4.29(d,J=7.5Hz,1 H,H-1”),4.11(d,J=11.2Hz,1H),3.82(d,J=2.5Hz,1H),3.79-3.65(m,4H),3.58-3.45(m,5 H),3.44-3.38(m,2H),3.35(t,J=4.8Hz,2H),3.26(d,J=11.3Hz,1H),2.23(d,J=11.2Hz,1H ),1.13(s,3H),1.05(s,3H),0.97(s,3H),0.90(d,J=6.1Hz,3H),0.84(s,3H),0.70(s,3H). 13 C NMR(101MHz,DMSO-d6)δ175.06(C-28),137.82(C-13),124.82(C-12),103.74,94.06, 76.53,76.47,75.62,75.21,73.50,72.21,70.66,69.35,68.12,67.92,67.50,63.97, 60.33,52.33,47.35,47.08,46.08,42.52,41.78,38.36,37.30,35.87,32.19,30.09, 27.67,23.75,23.24,23.08,21.05,17.42,17.06,17.01,16.91,13.83.HRMS(ESI):m / z calcd.for[M+Na] + C 42 H 68 O 15 Na 835.44504, found 835.44558.

[0136] Example 29

[0137] 2α,3β,23-Trihydroxy-ursane-12(13)-ene-28-O-α-L-arabinopyranosyl-(1→6)-β-D-glucopyranoside (Compound 29)

[0138] Following the methods of Examples 1 and 21, intermediate 29-1 was obtained. Then, following the method of Example 1, intermediate 1-4 was replaced with 29-1 to obtain compound 29 (white solid, 141 mg, yield: 80%). 1H NMR(300MHz, Methanol-d4)δ5.31(d,J=7.9Hz,1H,H-1'),5.25(s,1H),4.29(d,J=7.5Hz,1 H,H-1”),4.11(d,J=11.2Hz,1H),3.82(d,J=2.5Hz,1H),3.79-3.65(m,4H),3.58-3.45(m,5 H),3.44-3.38(m,2H),3.35(t,J=4.8Hz,2H),3.26(d,J=11.3Hz,1H),2.23(d,J=11.2Hz,1H ),1.13(s,3H),1.05(s,3H),0.97(s,3H),0.90(d,J=6.1Hz,3H),0.84(s,3H),0.70(s,3H). 13 C NMR(101MHz,DMSO-d6)δ175.06(C-28),137.82(C-13),124.82(C-12),103.74,94.06, 76.53,76.47,75.62,75.21,73.50,72.21,70.66,69.35,68.12,67.92,67.50,63.97, 60.33,52.33,47.35,47.08,46.08,42.52,41.78,38.36,37.30,35.87,32.19,30.09, 27.67,23.75,23.24,23.08,21.05,17.42,17.06,17.01,16.91,13.83.HRMS(ESI):m / z calcd.for[M+Na] + C 42 H 68 O 15 Na 835.44504, found 835.44558.

[0139] Example 30

[0140] 2α,3β,23-Trihydroxy-ursane-12(13)-ene-28-O-β-D-xylanopyranosyl-(1→6)-β-D-glucopyranoside (Compound 30)

[0141] Following the methods of Examples 1 and 21, intermediate 30-1 was obtained. Then, following the method of Example 1, intermediate 1-4 was replaced with 30-1 to obtain compound 30 (white solid, 196.5 mg, yield: 84%). 1H NMR (300MHz, Methanol-d4) δ5.32 (d, J = 8.0 Hz, 1H, H-1'), 5.25 (s, 1H), 4.29 (d, J = 6. 1Hz,1H,H-1”),4.03(d,J=11.1Hz,1H),3.93-3.77(m,2H),3.75-3.65(m,2H),3.63- 3.44(m,6H),3.43-3.33(m,3H),3.26(d,J=11.3Hz,1H),2.23(d,J=11.4Hz,1H),1.1 3(s,3H),1.05(s,3H),0.97(s,3H),0.90(d,J=5.9Hz,3H),0.83(s,3H),0.69(s,3H). 13 C NMR(126MHz,DMSO-d6)δ175.01,137.75,124.85,103.00,93.95,76.47,76.23,75.5 8,72.45,72.18,70.44,69.57,67.61,67.46,67.12,64.59,63.94,52.30,47.32,47. 06,46.04,42.49,41.73,40.11,39.94,39.78,38.34,37.27,35.83,32.17,30.05,27 .60,23.71,23.23,23.04,21.01,17.39,17.02,16.96,16.87,13.80.HRMS(ESI):m / z calcd.for[M+Na] + C 41 H 66 O 14 Na805.43448, found 805.43393.

[0142] Example 31

[0143] In vitro anti-inflammatory effects of the compound

[0144] The inhibitory effect of the compound on LPS-induced inflammatory response in THP1 differentiated macrophages was detected using an ELISA assay.

[0145] 1. Cell Culture

[0146] THP1 cells: RPMI 1640 complete medium (10% inactivated fetal bovine serum and 1% streptomycin / penicillin) were cultured in an incubator at 37°C with 5% CO2.

[0147] THP1 cell plating and drug administration: Human THP1 cells with a viable cell ratio of over 90% were used for the experiment. Cell suspension was placed in 15ml centrifuge tubes, centrifuged at 900rpm for 5min, the supernatant was discarded, and the pellet was resuspended for cell counting. Cell suspensions were then prepared according to the standard of 400,000 cells per well in a 12-well plate. PMA (50ng / ml) was added, and the cell suspension was evenly spread to each well (1ml per well). After differentiation and adhesion for 48h, the culture medium was replaced with fresh medium, and cultured for another 24h before drug administration. LPS modeling was initiated, and the corresponding compound was administered simultaneously. The final concentration of the test compound was set at 10μM. 200μM AICAR was used as a positive control compound. Drug administration was carried out for 12 hours under complete culture conditions.

[0148] 2. Collect cell supernatant

[0149] After simultaneous modeling and drug administration under complete culture conditions for 12 hours, the cell supernatant was collected into a 1.5 ml EP tube, centrifuged at 800 rpm at room temperature for 5 min, the supernatant was aliquoted into different EP tubes, and stored at -80℃ for later use.

[0150] 3. Reagent preparation

[0151] (1) 1×Coating Buffer A solution: Dilute 5×Coating Buffer A with ddH2O at a ratio of 1:5 to the required volume;

[0152] (2) 1×Capture Antibody solution: Dilute 200×Capture Antibody with 1×Coating Buffer A at a ratio of 1:200 to the required volume;

[0153] (3) 1×Assay Diluent A solution: Dilute 5×Assay Diluent A with 1×PBS at a ratio of 1:5 to the required volume;

[0154] (4) 1×PBST: 1×PBS+0.05% Tween-20;

[0155] (5) 1×Detection Antibody solution: 200×Detection Antibody and 1×Assay Diluent A are diluted to the required volume at a ratio of 1:200;

[0156] (6) 1×Avidin-HRP: 1000×Avidin-HRP and 1×Assay Diluent A were diluted to the required volume at a ratio of 1:1000;

[0157] (7) TMB Substrate Solution: Mix Substrate Solution A and Substrate Solution B in a 1:1 ratio and prepare fresh before use;

[0158] (8) Stop Solution: 2N H2SO4 solution.

[0159] 4. ELISA experimental procedure

[0160] (1) Coating the plate: Add 100 μl of 1×Capture Antibody to each well of the 96 plate and let it stand at 2-8℃ for 16-18 hours to coat the antibody in preparation for the ELISA experiment the next day.

[0161] (2) Discard the coating solution, wash 4 times with 1×PBST, add 200μl of 1×Assay Diluent A to each well for blocking, and incubate with gentle shaking at room temperature for 1h;

[0162] (3) Discard the blocking solution, wash 4 times with 1×PBST, add 100μl of diluted standard and test sample to each well, and incubate gently at room temperature for 2 hours.

[0163] (4) Discard the sample, wash 4 times with 1×PBST, add 100μl of 1×Detection Antibody solution to each well to detect the antibody, and incubate gently at room temperature for 1h.

[0164] (5) Discard the detection antibody, wash 4 times with 1×PBST, add 100μl of 1×Avidin-HRP solution to each well, and incubate gently at room temperature for 30 min;

[0165] (6) Discard the Avidin-HRP solution, wash 5 times with 1×PBST, each time for 30s-1min, add 100μl of freshly prepared TMB Substrate Solution to each well, and incubate at room temperature in the dark for 15min.

[0166] (7) Add 100 μl of Stop Solution to each well. On the microplate reader, select wavelengths of 450 nm and 570 nm and complete the reading within 15 min.

[0167] 5. The experimental results are shown in Figure 1. Asiatic acid and its saponins significantly inhibited the secretion of the pro-inflammatory factor IL-6 in THP1-differentiated macrophages induced by lipopolysaccharide. Furthermore, most saponins exhibited superior in vitro anti-inflammatory activity compared to asiatic acid (AA) and asiaticoside (As), particularly compounds 3, 14, 17, 18, 20, 24, and 26, which showed very strong in vitro anti-inflammatory activity.

[0168] Example 32

[0169] Protective effect of the compound on an anhydrous ethanol-induced rat gastric ulcer model

[0170] 1. Laboratory animals

[0171] Male SD rats, 6-7 weeks old and weighing 220-250g, were purchased from Vital River Laboratory Animal Technology Co., Ltd. The SD rats were housed under SPF conditions at a temperature of 25±2℃ and humidity of 50%-70%, with alternating 12-hour light and dark cycles. They had free access to food and water, and their corn cob bedding was changed twice a week. Experiments began after the rats had acclimatized to their environment for 7 days.

[0172] 2. Experimental apparatus

[0173] Animal weighing scale, balance, vortex mixer, magnetic stirrer, CNC rocker bed

[0174] 3. Preparation of experimental reagents and drugs

[0175] Sodium carboxymethyl cellulose (Sinopharm), the test compound, anhydrous ethanol (GENERAL-REAGENT, ≥99.8%), and omeprazole (positive control, purchased from Shanghai Yuanye Biotechnology). Weigh 5g of sodium carboxymethyl cellulose powder into a 1000mL wide-mouth bottle, add 1000mL of double-distilled water, and stir evenly on a magnetic stirrer to prepare a 0.5% sodium carboxymethyl cellulose solution. Grind the positive control or test compound thoroughly in a mortar, add an appropriate amount of the 0.5% sodium carboxymethyl cellulose solution, and grind to suspend the compound, preparing a 2mg / mL test solution.

[0176] 4. Experimental Procedure

[0177] (1) Modeling and drug administration

[0178] SD rats were randomly divided into groups of three. After a 24-hour fast with unlimited access to water, all rats were orally administered anhydrous ethanol solution (5 μL / g) via gavage. One hour after modeling, the model group rats were administered 0.5% sodium carboxymethyl cellulose solution via gavage, while the positive control group (omeprazole) and the test drug group rats were administered the corresponding drug suspensions via gavage at a dose of 10 mg / kg. One hour after administration, the animals were sacrificed and dissected for tissue collection.

[0179] (2) Collection of materials

[0180] Gastric tissue was harvested by opening the abdominal cavity along the midline and cutting open the gastric cavity along the greater curvature. Gastric juice and contents were washed out with physiological saline. The washed gastric mucosa was fixed on a foam board and photographed. The area of ​​the ulcer region was analyzed using ImageJ software, and the proportion of the ulcer region was calculated. Then, the ratio of the proportion of the ulcer region in the treatment group to that in the model group was calculated (Table 2).

[0181] Table 2. Results of the pharmacodynamic study of the compound in treating an anhydrous ethanol-induced gastric ulcer model.

[0182] 5. Experimental Results:

[0183] The experimental results, as shown in Table 2 and Figure 2, indicate that after modeling with anhydrous ethanol, rats experienced severe gastric mucosal bleeding and significant gastric ulceration. Treatment with the drugs reduced both bleeding and ulceration. Table 2 shows that treatment with the compounds reduced the proportion of ulcerated areas, indicating that the compounds have anti-gastric ulcer efficacy. Among them, compounds 3, 4, 6, 13, 15, 17, 19, 20, 22, 26, and 29 showed significantly better efficacy than omeprazole, a commonly used clinical drug for treating gastric ulcers, at the same dosage. This suggests that the compounds of this invention can be used for the prevention and treatment of gastric ulcers and other peptic ulcer diseases.

[0184] Example 33

[0185] Therapeutic effect of compound 6 on DSS-induced mouse ulcerative colitis model

[0186] 1. Laboratory animals

[0187] Male C57 mice, 8 weeks old and weighing approximately 25g, were purchased from Vital River Laboratory Animal Technology Co., Ltd. The C57 mice were housed under SPF conditions at a temperature of 25±2℃ and humidity of 50%-70%, with alternating 12-hour light and dark cycles. They had free access to food and water, and their corn cob bedding was changed twice a week. Experiments began after the mice had acclimatized to their environment for 7 days.

[0188] 2. Experimental apparatus

[0189] Animal weighing scale, balance, vortex mixer, magnetic stirrer, CNC rocker bed

[0190] 3. Preparation of experimental reagents and drugs

[0191] Sodium carboxymethyl cellulose (purchased from Sinopharm Group), Compound 6, sodium dextran sulfate (DSS) (36000-50000 M.Wt, purchased from MPBIO), and 5-aminosalicylic acid (positive control, purchased from Aladdin Reagent Company) were used. 5g of sodium carboxymethyl cellulose powder was weighed into a 1000mL wide-mouth bottle, and 1000mL of double-distilled water was added. The mixture was stirred evenly on a magnetic stirrer to prepare a 0.5% sodium carboxymethyl cellulose solution. The positive control or Compound 6 was then thoroughly ground in a mortar, and an appropriate amount of the 0.5% sodium carboxymethyl cellulose solution was added to suspend the mixture.

[0192] 4. Experimental Procedure

[0193] Thirty C57 mice were randomly divided into 6 groups of 5 mice each. The control group was fed purified water, while the model group, drug-treated group, and positive control group were fed purified water containing 2% sodium dextran sulfate (DSS) for seven days, while receiving drug intervention simultaneously. Mice in the control and model groups were orally administered a placebo of 0.5% sodium carboxymethyl cellulose solution, mice in the treatment group were orally administered compound 6 (C6, at doses of 10, 20, and 50 mg / kg, respectively), and mice in the positive control group were orally administered the clinical drug 5-aminosalicylic acid (5-ASA, at a dose of 100 mg / kg). On the eighth day, the mice were sacrificed by cervical dislocation, and colon tissue was harvested and its length measured. The differences in colon length among the groups were statistically analyzed.

[0194] 5. Experimental Results:

[0195] The experimental results, as shown in Figures 3 and 4, indicate that DSS significantly shortened the colon length in mice, and treatment with compound 6 significantly improved this phenomenon. Furthermore, compound 6 showed a better effect on improving colon length than the positive control drug 5-ASA. This suggests that compound 6 can be used for the prevention and treatment of ulcerative colitis. Other compounds of this invention also possess similar functions.

[0196] Example 34

[0197] Single-dose toxicity studies of compounds 4 and 6 in mice

[0198] Eight-week-old C57 mice (purchased from Vital River Laboratory Animal Technology Co., Ltd.) were used in groups of 10 (5 males and 5 females) and randomly assigned to three groups: a control group, a compound 4 group, and a compound 6 group. After acclimatization, the mice were randomly assigned to groups according to weight and sex. Before administration, the mice were fasted for 12 hours but allowed free access to water. The administration groups were administered either compound 4 or compound 6 by gavage at a dose of 2 g / kg, while the control group was administered an equal volume of 0.5% CMC-Na as a control solvent. The mice were allowed to eat again 2 hours after administration. The mice were then observed for 14 consecutive days. During this period, no mice in the compound 4 or compound 6 groups died, and their weight, movement, respiration, excretion, and mental status remained normal, suggesting that compounds 4 and 6 have good safety profiles. Other compounds of this invention also exhibit good safety profiles.

[0199] Example 35

[0200] tablet

[0201] The compound 6 (50g) obtained in Example 6, hydroxypropyl methylcellulose E (150g), starch (200g), appropriate amount of povidone K30 and magnesium stearate (1g) were mixed, granulated and compressed into tablets.

[0202] In addition, according to the conventional formulation method of the 2015 edition of the Pharmacopoeia, the compounds obtained in Examples 1 to 30 can be given different pharmaceutical excipients to make capsules, powders, tablets, granules, pills, injections, syrups, oral liquids, inhalers, ointments, solutions, creams, gels, powders, lotions, tinctures, suppositories or patches.

Claims

1. A asiatic acid saponin compound or its pharmaceutically acceptable salt, ester or solvate as shown in formula (I): in, R 1 Selected from H, β-D-glucopyranosyl, β-D-galactopyranosyl, β-D-xylanosyl, α-L-arabinopyranosyl, α-L-rhamnosyl, α-L-rhamnosyl-(1→2)-α-L-arabinopyranosyl, α-L-rhamnosyl-(1→3)-α-L-arabinopyranosyl, β-D-xylanosyl-(1→2)-α-L-arabinopyranosyl, β-D-xylanosyl-(1→3)-α-L-arabinopyranosyl, β-D-glucopyranosyl-(1→2)-α-L-arabinopyranosyl, or β-D-glucopyranosyl-(1→3)-α-L-arabinopyranosyl; R 2 Selected from H, β-D-glucopyranosyl, β-D-galactopyranosyl, β-D-xylanosyl, α-L-arabinopyranosyl, α-L-rhamnosyl, α-L-rhamnosyl-(1→2)-α-L-arabinopyranosyl, α-L-rhamnosyl-(1→3)-α-L-arabinopyranosyl, β-D-xylanosyl-(1→2)-α-L-arabinopyranosyl, β-D-xylanosyl-(1→3)-α-L-arabinopyranosyl, β-D-glucopyranosyl-(1→2)-α-L-arabinopyranosyl, or β-D-glucopyranosyl-(1→3)-α-L-arabinopyranosyl; R 1 and R 2 They are not both H.

2. The asiatic acid saponin compound, its pharmaceutically acceptable salt, ester, or solvate according to claim 1, characterized in that, The compound or its pharmaceutically acceptable salt, ester, or solvent compound is selected from any one of the following compounds:

3. Use of the compound according to any one of claims 1-2 or a pharmaceutically acceptable salt, ester or solvate thereof in the preparation of a medicament for the prevention or treatment of inflammatory diseases, ulcerative diseases or tumor diseases.

4. The use according to claim 3, characterized in that, The inflammatory diseases mentioned are metabolic diseases, including: insulin resistance, metabolic syndrome, type 1 or type 2 diabetes, hyperlipidemia, obesity, atherosclerosis, myocardial ischemia, myocardial infarction, arrhythmia, coronary heart disease, hypertension, heart failure, myocardial hypertrophy, myocarditis, diabetic complications, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis, alcoholic fatty liver disease, cirrhosis, hyperuricemia, gout, osteoporosis, stroke or cerebral infarction.

5. The use according to claim 3, characterized in that, The inflammatory diseases mentioned are secondary diseases caused by autoimmune diseases, organ fibrosis, neurodegenerative diseases, or pathogen infections, including: pneumonia, tuberculosis, inflammatory bowel disease, Behcet's disease, asthma, chronic obstructive pulmonary disease, chronic bronchitis, emphysema, bronchiolitis obliterans, systemic lupus erythematosus, rheumatoid arthritis, spondyloarthritis, osteoarthritis, synovitis, tendinitis, thromboangiitis obliterans, phlebitis, intermittent claudication, keloids, and psoriasis. Ichthyosis, bullous pemphigoid, dermatitis, contact dermatitis, pancreatitis, chronic nephritis, cystitis, meningitis, gastritis, sepsis, pyoderma gangrenosa, uveitis, idiopathic pulmonary fibrosis, cystic fibrosis, Parkinson's disease, Alzheimer's disease, alpha-common protein disease, depression, multiple sclerosis, systemic sclerosis, amyotrophic lateral sclerosis, fibromyalgia syndrome, neuralgia, Down syndrome, Hallewarden-Scholes disease, Huntington's disease, or Wilson's disease.

6. The use according to claim 3, characterized in that, The ulcerative diseases mentioned are peptic ulcers, including: gastric ulcers, duodenal ulcers, reflux esophagitis, or erosive esophagitis.

7. The use according to claim 3, characterized in that, The tumor diseases mentioned include: bone cancer, acute myeloid leukemia, chronic myeloid leukemia, acute lymphoblastic leukemia, chronic lymphoblastic leukemia, myeloproliferative disorders, multiple myeloma, myelodysplastic syndrome, Hodgkin's lymphoma, non-Hodgkin's lymphoma, hemangioma, granuloma, xanthoma, meningeal sarcoma, glioma, astrocytoma, medulloblastoma, ependymoma, germ cell tumor (pineal tumor), glioblastoma multiforme, oligodendroglioma, schwannoma, retinoblastoma, neurofibroma, sarcoma, esophageal cancer, gastric cancer, pancreatic cancer, colorectal cancer, colon cancer, rectal cancer, kidney cancer, prostate cancer, lymphoma, testicular cancer, interstitial cell carcinoma, lung cancer, liver cancer, skin cancer, malignant melanoma, or basal cell carcinoma.

8. A pharmaceutical composition for the prevention or treatment of inflammatory or ulcerative diseases, characterized in that, The composition comprises the compound of any one of claims 1 to 2 or a pharmaceutically acceptable salt, ester, or solvent compound thereof as an active ingredient and a pharmaceutically acceptable excipient.

9. The pharmaceutical composition according to claim 8, characterized in that, The pharmaceutical composition is preferably in the form of capsules, powders, tablets, granules, pills, injections, syrups, oral liquids, inhalers, ointments, suppositories, or patches.

Images