Diterpenoid compound, extraction method thereof and application of diterpenoid compound in preparation of anti-inflammatory drugs
Four novel diterpenoid compounds were isolated and purified from the whole plant of Euphorbia pekinensis using a multi-step chromatography method. This solved the problem of underutilization of the medicinal components of Euphorbia pekinensis, achieved significant anti-inflammatory effects, and provided a basis for new drug development.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- SHENYANG PHARMA UNIV
- Filing Date
- 2026-03-03
- Publication Date
- 2026-05-08
AI Technical Summary
In the existing technology, the medicinal value of Euphorbia pekinensis has not been fully utilized, and there is a lack of systematic research on its components, especially the isolation and application of diterpenoids, which has resulted in the ineffective utilization of its anti-inflammatory activity.
Four novel diterpenoid compounds were isolated from the whole herb of Euphorbia pekinensis using a multi-step chromatographic method. The compounds were purified by ethanol-water extraction, polyamide column chromatography, silica gel column chromatography, MCI column chromatography, ODS column chromatography, and Sephadex LH-20 gel chromatography. Combined with HPLC, compounds 1, 2, 3, and 4, as well as their pharmaceutically acceptable salts, were obtained for the preparation of anti-inflammatory drugs.
This study achieved efficient separation and purification of diterpenoids from Euphorbia pekinensis in southern Europe, providing compounds with significant anti-inflammatory activity that can inhibit LPS-induced NO production in RAW264.7 cells, thus providing active lead compounds for new drug development.
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Abstract
Description
Technical Field
[0001] This invention belongs to the field of traditional Chinese medicine extraction, specifically relating to a diterpenoid compound isolated from the whole herb of Euphorbia pekinensis in Southern Europe, its extraction method, and its application in the preparation of anti-inflammatory drugs. Background Technology
[0002] Southern European Euphorbia ( Euphorbia peplus *Euphorbia tirucalli*, commonly known as milkweed or sapgrass, is a small annual weed characterized by its white latex. Native to Europe and North Africa, particularly the Mediterranean region, this plant has become widely distributed globally due to its rapid growth and invasive nature. In China, it has been introduced to Yunnan and Guangdong provinces (Hua J, Liu Y, Xiao CJ, ...). and others .Chemical profile and defensive function of the latexof Euphorbia peplus [J]. Phytochemistry, 2017, 136: 56-64; Pu XX, Ran XQ, YanY, and others . Three new jatrophane diterpenoids from Euphorbia peplus Linn. withactivity towards autophagic flux [J]. Phytochemistry Letter, 2022, 50: 141-146. In traditional folk medicine, Euphorbia pekinensis is widely used to treat various diseases, including skin diseases, diabetes, asthma, inflammation, and tumors (Anastasiou E, Lorentz KO, Stein GJ, ...). et al. Prehistoricschistosomiasis parasite found in the Middle East [J]. The Lancet InfectiousDiseases, 2014, 14: 553-554; Chen H, Wang H, Yang B, and others .Diterpenesinhibiting NO production from Euphorbia helioscopia[J]. Fitoterapia, 2014, 95: 133-138.). Phytochemical studies on the whole herb of Euphorbia pekinensis have shown that this species is rich in diterpenoids, including compounds with skeletons of the pepluane, paraliane, and ent-rosine types, and exhibit various biological activities, such as anticancer, anti-inflammatory, antibacterial, and multidrug resistance reversal activities (Ali AA, Sayed HM, Ibrahim SRM, et al. Chemical constituents, antimicrobial,analgesic, antipyretic, and anti-inflammatory activities of Euphorbia peplus L[J]. Phytopharmacology, 2013, 4: 69-80; Chen L, Liu L, Li Y, et al. Macrocyclic Diterpenoids from Euphorbia peplus Possessing Activity Towards Autophagic Flux[J]. International Journal of Molecular Sciences, 2025, 26: 299; Gao Y, ZhouJS, Liu HC, et al. Phonerilins AK, cytotoxic ingenane and ingol diterpenoidsfrom Euphorbia neriifolia [J]. Tetrahedron, 2022, 123: 132955; Li Y, Yu ZP, LiYP, et al. Diterpenoids from Euphorbia peplus possessing cytotoxic and anti-inflammatory activities [J]. Bioorganic Chemistry, 145: 107194; Rizk AM,Hammouda FM, El-Missiry MM, et al. Biologically active diterpene estersfrom euphorbia peplos [J]. Phytochemistry, 24: 1605-1606; Wan LS, Chu R, PengXR, et al. Pepluane and Paraliane Diterpenoids from Euphorbia peplus withPotential Anti-inflammatory Activity [J]. Journal of Natural Products, 2016,79: 1628-1634; Wang W, Xiong L, Wu Y, and others . New lathyrane diterpenoid hybridshave anti-inflammatory activity through the NF-κB signaling pathway andautophagy [J]. Acta Materia Medica, 2022, 1: 224-243; Yan Y, Peng MY, Yang Y, et al. Highly oxygenated ent-abietane diterpenoid lactones from Euphorbia more and their anti-inflammatory activity [J]. Bioorganic Chemistry, 2025,154: 107989; Yan Y, Zhou Q, Ran X, et al. Jatrophane Diterpenoids from Euphorbia more Linn. as Activators of Autophagy and Inhibitors of Tau Pathology [J].International Journal of Molecular Sciences, 2023, 24: 1088; Yang Y, Zhou M,Wang D, et al. Jatrophane Diterpenoids from Euphorbia peplusAs MultidrugResistance Modulators with Inhibitory Effects on the ATR-Chk-1 Pathway [J]. Journal of Natural Products, 2021, 84: 339-351). To maximize the medicinal value of Euphorbia salina, a systematic component study was conducted, and new diterpenoid compounds were isolated. The structures of the compounds were confirmed using NMR and mass spectrometry, and the inhibitory effect of the extracted compounds on LPS-induced NO production in RAW264.7 cells was detected. Summary of the Invention
[0003] The primary objective of this invention is to provide four diterpenoid compounds.
[0004] A second objective of this invention is to provide a method for extracting the diterpenoid compounds.
[0005] A third object of the present invention is to provide a pharmaceutical composition containing the diterpenoid compound.
[0006] A fourth object of the present invention is to provide the use of the diterpenoid compound, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the compound, in the preparation of an anti-inflammatory medicament.
[0007] To achieve the above objectives, the technical solution of the present invention is summarized as follows: A diterpenoid compound of formula 1, 2, 3 or 4 isolated from Euphorbia pekinensis in southern Europe:
[0008] The pharmaceutically acceptable salts include sodium, potassium, ammonium, hydrochloride, and sulfate salts of the compound.
[0009] The extraction method for diterpenoid compounds described in this invention includes the following steps: (1) Using the whole herb of Euphorbia pekinensis as raw material, add 8-10 times the mass of the raw material in an aqueous ethanol solution, reflux extract, recover the solvent under reduced pressure from the extract, and concentrate to obtain the total extract. (2) Disperse the total extract into 2-6 times its mass of water, and extract it sequentially with petroleum ether and ethyl acetate. Concentrate the extract to recover the solvent, and obtain petroleum ether extract concentrate, ethyl acetate extract concentrate and water, respectively. (3) The petroleum ether extract concentrate was separated by polyamide column chromatography, and gradient elution was performed using ethanol-water with a volume ratio of 100:1-0:1. The crude fraction with an ethanol volume fraction of 80%-100% was collected. (4) The crude fraction was separated by silica gel column chromatography, and gradient elution was performed using petroleum ether-ethyl acetate with a volume ratio of 100:1-0:1. The fraction E4 with a volume ratio of 10:1-5:1 was collected. (5) Concentrated fraction E4: The concentrate was separated by MCI column chromatography and gradient elution was performed using methanol-water with a volume ratio of 100:1-0:1. Fraction E41 with a methanol volume fraction of 90%-100% was collected. (6) Concentrated fraction E41: The concentrate was separated by silica gel column chromatography and gradient elution was performed using petroleum ether-acetone with a volume ratio of 100:1-0:1. Fraction E412 with a volume ratio of 20:1-8:1 was collected. (7) Concentrated fraction E412: The concentrate was separated by silica gel column chromatography, and gradient elution was performed using petroleum ether-acetone with a volume ratio of 100:1-0:1. The fraction E4122 with a volume ratio of 20:1-5:1 was collected. (8) The concentrated fraction E4122 was separated by ODS column chromatography and further purified to obtain diterpenoid compound 1, compound 2, compound 3 and compound 4.
[0010] In step (1), the whole herb of Euphorbia pekinensis is used as raw material. An ethanol aqueous solution with a volume concentration of 60%-95% is added at 8-10 times the mass of the raw material. The mixture is refluxed and extracted 2-4 times, with each extraction lasting 2-4 hours. The extracts are combined, the solvent is recovered under reduced pressure, and the total extract is obtained after concentration.
[0011] The specific separation and purification process of fraction E4122 in step (8) is as follows: Fraction E4122 was concentrated and separated by ODS column chromatography with gradient elution using methanol-water (v / v) at a ratio of 50:50-100:0. The fraction with a methanol-water ratio of 70:30 was collected and designated as E41222. Fraction E41222 was then concentrated and separated by silica gel column chromatography with gradient elution using petroleum ether-acetone (v / v) at a ratio of 100:1-0:1. The fraction with a petroleum ether-acetone ratio of 20:1-8:1 was collected and designated as E412222. Fraction E412222 was purified by Sephadex LH-20 gel chromatography with methanol as the elution solvent, followed by preparative HPLC purification using acetonitrile-water (v / v) at a ratio of 60:40 to obtain compounds 1, 3, and 4.
[0012] Fraction E4122 was concentrated and separated by ODS column chromatography with gradient elution using methanol-water at a volume ratio of 50:50-100:0. The fraction with a methanol-water volume ratio of 50:50 was collected and designated as E41221. Fraction E41221 was concentrated and purified by Sephadex LH-20 gel chromatography with methanol as the elution solvent, followed by preparative HPLC purification with methanol-water at a volume ratio of 65:35 as the mobile phase to obtain compound 2.
[0013] The present invention also provides the use of the four diterpenoid compounds or pharmaceutically acceptable salts of the compounds in the preparation of anti-inflammatory drugs.
[0014] A pharmaceutical composition comprising one or more of the diterpenoid compound or a pharmaceutically acceptable salt thereof.
[0015] A pharmaceutical formulation comprising the diterpenoid compound or a pharmaceutically acceptable salt thereof as an active ingredient, and further comprising pharmaceutically acceptable excipients.
[0016] The drug preparation is administered orally or by injection, and the dosage form is: tablet, capsule, powder, syrup or injection.
[0017] The present invention also provides the use of the pharmaceutical composition or pharmaceutical preparation in the preparation of anti-inflammatory drugs.
[0018] The diterpenoid compound, or its isomer, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition thereof, has an inhibitory effect on NO production induced by LPS in RAW264.7 cells and can be used to prepare anti-inflammatory drugs.
[0019] Advantages of this invention: The diterpenoid compounds of the present invention, or pharmaceutically acceptable salts thereof, or pharmaceutical compositions containing such compounds, inhibit LPS-induced NO production in RAW264.7 cells, and therefore can be used to prepare anti-inflammatory drugs. The method of the present invention further enriches the structural diversity of Euphorbia species, and on this basis, lays the foundation for subsequent bioactivity testing of the obtained monomeric compounds, provides active lead compounds for new drug development, and also provides a theoretical basis for in-depth research and development of Euphorbia species. Attached Figure Description
[0020] Figure 1 Compounds 1-4 exhibit inhibitory activity against LPS-induced NO production in RAW264.7. Detailed Implementation
[0021] The technical solution of the present invention will be further described below with reference to specific embodiments.
[0022] Example 1 The extraction method for diterpenoid compounds from Euphorbia pekinensis in Southern Europe includes the following steps: (1) Using the whole herb of Euphorbia pekinensis as raw material (15 kg), add 10 times the mass of the raw material (150 L) of 95% ethanol aqueous solution, reflux extract three times, each extraction for 3 hours, combine the extracts, recover the solvent under reduced pressure, concentrate and obtain the total extract. (2) Disperse the total extract into 4 times its mass of water, and extract it sequentially with petroleum ether and ethyl acetate. Concentrate the extract to recover the solvent, and obtain petroleum ether extract concentrate, ethyl acetate extract concentrate and water, respectively. (3) The petroleum ether extract concentrate was separated by polyamide column chromatography, and gradient elution was performed using ethanol-water with a volume ratio of 100:1-0:1. The crude fraction with an ethanol volume fraction of 80%-100% was collected. (4) The crude fraction was separated by silica gel column chromatography, and gradient elution was performed using petroleum ether-ethyl acetate with a volume ratio of 100:1-0:1. The fraction E4 with a volume ratio of 10:1-5:1 was collected. (5) Concentrated fraction E4 (60.0 g): The concentrate was separated by MCI column chromatography and gradient elution was performed using methanol-water with a volume ratio of 100:1-0:1. The fraction E41 with a methanol volume fraction of 90%-100% was collected. (6) Concentrated fraction E41 (42.2 g): The concentrate was separated by silica gel column chromatography and gradient elution was performed using petroleum ether-acetone with a volume ratio of 100:1-0:1. Fraction E412 with a volume ratio of 20:1-8:1 was collected. (7) Concentrated fraction E412 (31.6 g): The concentrate was separated by silica gel column chromatography and gradient elution was performed using petroleum ether-acetone with a volume ratio of 100:1-0:1. The fraction E4122 with a volume ratio of 20:1-5:1 was collected. (8) The concentrated fraction E4122 (19.2 g) was separated by ODS column chromatography and further purified to obtain diterpenoid compound 1, compound 2, compound 3 and compound 4.
[0023] The specific separation and purification process of fraction E4122 in step (8) is as follows: Fraction E4122 was concentrated and separated by ODS column chromatography with gradient elution using methanol-water (v / v) at a ratio of 50:50-100:0. The fraction with a methanol-water ratio of 70:30 was collected and designated as E41222. Fraction E41222 (7.6 g) was concentrated and separated by silica gel column chromatography with gradient elution using petroleum ether-acetone (v / v) at a ratio of 100:1-0:1. The fraction with a petroleum ether-acetone ratio of 20:1-8:1 was collected and designated as E412222. Fraction E412222 (6.0 g) was concentrated and purified by Sephadex LH-20 gel chromatography with methanol as the elution solvent, followed by preparative HPLC purification using acetonitrile-water (v / v) at a ratio of 60:40 to obtain compounds 1 (34.0 mg), 3 (16.9 mg), and 4 (13.9 mg).
[0024] Fraction E4122 was concentrated and separated by ODS column chromatography with gradient elution using methanol-water at a volume ratio of 50:50-100:0. The fraction with a methanol-water volume ratio of 50:50 was collected and designated as E41221. Fraction E41221 (2.8 g) was concentrated and purified by Sephadex LH-20 gel chromatography with methanol as the elution solvent, followed by preparative HPLC purification using methanol-water at a volume ratio of 65:35 as the mobile phase to obtain compound 2 (4.1 mg).
[0025] The structures of the extracted diterpenoid compounds 1-4 were identified, and the specific physicochemical data are as follows: Compound 1: White amorphous powder (CH3OH); +21.6 ( c = 0.50, CH3OH); UV (CH3OH) λ max (log ε ) 200 (0.85), 225 (0.71) nm; 1 H-NMR (600 MHz, CDCl3) and 13 C-NMR (150MHz, CDCl3) data are shown in Table 1; HRESIMS m / z 557.2520 [M + Na] + (calcd. for C 32 H 38 O7Na + ,557.2618), with the molecular formula C 32 H 38 O7.
[0026] Compound 2: White amorphous powder (CH3OH); +29.4 ( c = 0.50, CH3OH); UV (CH3OH) λ max (log ε 202 (1.02) nm; 1 H-NMR (600 MHz, CDCl3) and 13 C-NMR (150 MHz, CDCl3) data are shown in Table 2; HRESIMS m / z : 453.2253 [M + Na] + (calcd. for C 25 H 34 O6Na + (453.2355), molecular formula is C 25 H 34 O6.
[0027] Compound 3: White powder (CH3OH); +30.6 ( c = 0.50, CH3OH), UV (CH3OH) λ max (log ε ) 201 (1.95), 230 (3.46) nm; 1 H-NMR (600 MHz, CDCl3) and 13 C-NMR (150 MHz, CDCl3) data are shown in Table 3. HR-ESI-MS m / z 595.2309 [M + Na] + (calcd. for C 34 H 36 O8Na + ,595.2410), molecular formula C 34 H 36 O8.
[0028] Compound 4: White powder (CH3OH); +25.4 c = 0.50, CH3OH), UV (CH3OH) λ max (log ε ) 201 (1.95), 246 (3.52) nm; 1 H-NMR (600 MHz, CDCl3) and 13C-NMR (150 MHz, CDCl3) data are shown in Table 4. HR-ESI-MS m / z 573.2468 [M + Na] + (calcd. for C 32 H 38 O8Na + ,573.2567), molecular formula C 32 H 38 O8.
[0029] Table 1. Carbon and proton spectrum data of compound 1
[0030] Table 2. Carbon and proton spectrum data of compound 2
[0031] Table 3. Carbon and hydrogen spectral data of compound 3
[0032] Table 4. Carbon and proton spectrum data of compound 4
[0033] Using physicochemical data and modern spectroscopic techniques (HR-ESI-MS and NMR), combined with relevant data from published literature, the structures of the above compounds were identified, and compounds 1-4 were determined to be novel compounds not previously reported in the literature, as shown below:
[0034] Example 2 The extraction method for diterpenoid compounds from Euphorbia pekinensis in Southern Europe includes the following steps: (1) Using the whole herb of Euphorbia pekinensis as raw material (10 kg), add 8 times the mass of the raw material (80 L) of 90% ethanol aqueous solution, reflux extract 3 times, each extraction for 3 hours, combine the extracts, recover the solvent under reduced pressure, concentrate and obtain the total extract. (2) Disperse the total extract into water at twice its mass, and extract it sequentially with petroleum ether and ethyl acetate. Concentrate the extract to recover the solvent, and obtain petroleum ether extract concentrate, ethyl acetate extract concentrate and water, respectively. (3) The petroleum ether extract concentrate was separated by polyamide column chromatography, and gradient elution was performed using ethanol-water with a volume ratio of 100:1-0:1. The crude fraction with an ethanol volume fraction of 80%-100% was collected. (4) The crude fraction was separated by silica gel column chromatography, and gradient elution was performed using petroleum ether-ethyl acetate with a volume ratio of 100:1-0:1. The fraction E4 with a volume ratio of 10:1-5:1 was collected. (5) Concentrated fraction E4: The concentrate was separated by MCI column chromatography and gradient elution was performed using methanol-water with a volume ratio of 100:1-0:1. Fraction E41 with a methanol volume fraction of 90%-100% was collected. (6) Concentrated fraction E41: The concentrate was separated by silica gel column chromatography and gradient elution was performed using petroleum ether-acetone with a volume ratio of 100:1-0:1. Fraction E412 with a volume ratio of 20:1-8:1 was collected. (7) Concentrated fraction E412: The concentrate was separated by silica gel column chromatography and gradient elution was performed using petroleum ether-acetone with a volume ratio of 100:1-0:1. The fraction E4122 with a volume ratio of 20:1-5:1 was collected. (8) The concentrated fraction E4122 was further purified by ODS column chromatography to obtain diterpenoid compound 1 (22.5 mg), compound 2 (2.7 mg), compound 3 (11.3 mg) and compound 4 (9.2 mg). The specific separation and purification process was the same as in Example 1.
[0035] Example 3 The Griess method was used to detect the effect of compounds 1-4 on inhibiting lipopolysaccharide-induced nitric oxide (NO) release from mouse RAW264.7 macrophages: 1. Principle: Excessive LPS induces the activation and expression of nitric oxide synthase (NOS) in macrophages, leading to the production of NO. NO then reacts with oxygen free radicals to rapidly form NO2. - NO2 formed - It can be quantitatively detected by Griess Reagent under acidic conditions. First, NO2 - It undergoes a diazotization reaction with sulfanilamide, and then with... N -(1-Naphthyl)ethylenediamine dihydrochloride ( N (1-naphthyl)ethylenediamine dihydrochloride undergoes a coupling reaction to generate a purple-red azo compound. Finally, the absorbance value is measured at 540 nm using an enzyme-linked immunosorbent assay (ELISA) reader. Combined with the standard curve, the NO content in the sample can be determined.
[0036] 2. Methods: RAW264.7 mouse mononuclear macrophages in the logarithmic growth phase were collected and their concentration adjusted to 3.5 × 10⁻⁶ cells. 4Cells / wells were seeded in 96-well plates, with 100 μL of cell suspension added to each well. The experiment included a control group (RAW264.7 cells, DMSO), a model group (RAW264.7 cells, DMSO, 0.5 μg / mL LPS), a positive control group (RAW264.7 cells, dexamethasone (20 μM), 0.5 μg / mL LPS), and a test drug group (RAW264.7 cells, each test compound (20 μM), 0.5 μg / mL LPS). Cells were incubated in a 5% CO2, 37℃ incubator for 24 h. Then, 40 μL of cell supernatant was transferred to an ELISA plate, and an equal volume of Griess reagent was added. The accumulation of nitrite in the culture medium was measured at 540 nm using an ELISA reader. Results are as follows: Figure 1 As shown, compounds 1 and 3-4 exhibit extremely strong activity in inhibiting NO release.
[0037] The above description of the embodiments is only for the purpose of helping to understand the method and central idea of the present invention. It should be noted that those skilled in the art can make several improvements and modifications to the present invention without departing from the principle of the present invention, and these improvements and modifications also fall within the protection scope of the present invention.
Claims
1. A diterpenoid compound or a pharmaceutically acceptable salt thereof, characterized in that, The diterpenoids are selected from the following compounds: 。 2. A method for extracting the diterpenoid compound of claim 1 or a pharmaceutically acceptable salt thereof, characterized in that, Includes the following steps: (1) Using the whole herb of Euphorbia pekinensis as raw material, add 8-10 times the mass of the raw material in an aqueous ethanol solution, reflux extract, recover the solvent under reduced pressure from the extract, and concentrate to obtain the total extract. (2) Disperse the total extract into 2-6 times its mass of water, and extract it sequentially with petroleum ether and ethyl acetate. Concentrate the extract to recover the solvent, and obtain petroleum ether extract concentrate, ethyl acetate extract concentrate and water, respectively. (3) The petroleum ether extract concentrate was separated by polyamide column chromatography, and gradient elution was performed using ethanol-water with a volume ratio of 100:1-0:
1. The crude fraction with an ethanol volume fraction of 80%-100% was collected. (4) The crude fraction was separated by silica gel column chromatography, and gradient elution was performed using petroleum ether-ethyl acetate with a volume ratio of 100:1-0:
1. The fraction E4 with a volume ratio of 10:1-5:1 was collected. (5) Concentrated fraction E4: The concentrate was separated by MCI column chromatography and gradient elution was performed using methanol-water with a volume ratio of 100:1-0:
1. Fraction E41 with a methanol volume fraction of 90%-100% was collected. (6) Concentrated fraction E41: The concentrate was separated by silica gel column chromatography and gradient elution was performed using petroleum ether-acetone with a volume ratio of 100:1-0:
1. Fraction E412 with a volume ratio of 20:1-8:1 was collected. (7) Concentrated fraction E412: The concentrate was separated by silica gel column chromatography, and gradient elution was performed using petroleum ether-acetone with a volume ratio of 100:1-0:
1. The fraction E4122 with a volume ratio of 20:1-5:1 was collected. (8) The concentrated fraction E4122 was separated by ODS column chromatography and further purified to obtain diterpenoid compound 1, compound 2, compound 3 and compound 4.
3. The extraction method for the diterpenoid compound or its pharmaceutically acceptable salt according to claim 2, characterized in that, In step (1), the whole herb of Euphorbia pekinensis is used as raw material. An ethanol aqueous solution with a volume concentration of 60%-95% is added at 8-10 times the mass of the raw material. The mixture is refluxed and extracted 2-4 times, with each extraction lasting 2-4 hours. The extracts are combined, the solvent is recovered under reduced pressure, and the total extract is obtained after concentration.
4. The extraction method for the diterpenoid compound or its pharmaceutically acceptable salt according to claim 2, characterized in that, The specific separation and purification process of fraction E4122 in step (8) is as follows: Fraction E4122 was concentrated and then separated by ODS column chromatography with gradient elution using methanol-water at a volume ratio of 50:50-100:
0. The fraction with a methanol-water volume ratio of 70:30 was collected and designated as E41222. Fraction E41222 was then concentrated and separated by silica gel column chromatography with gradient elution using petroleum ether-acetone at a volume ratio of 100:1-0:
1. The fraction with a petroleum ether-acetone volume ratio of 20:1-8:1 was collected and designated as E412222. Fraction E412222 was purified by Sephadex LH-20 gel chromatography with methanol as the elution solvent, followed by preparative HPLC chromatography with acetonitrile-water at a volume ratio of 60:40 as the mobile phase to separate compounds 1, 3, and 4. Fraction E4122 was concentrated and separated by ODS column chromatography with gradient elution using methanol-water at a volume ratio of 50:50-100:
0. The fraction with a methanol-water volume ratio of 50:50 was collected and designated as E41221. Fraction E41221 was concentrated and purified by Sephadex LH-20 gel chromatography with methanol as the elution solvent, followed by preparative HPLC purification with methanol-water at a volume ratio of 65:35 as the mobile phase to obtain compound 2.
5. A pharmaceutical composition, characterized in that, The pharmaceutical composition contains one or more of the diterpenoid compounds of claim 1 or their pharmaceutically acceptable salts.
6. A pharmaceutical preparation, characterized in that, The pharmaceutical preparation uses one or more of the diterpenoid compounds of claim 1 or their pharmaceutically acceptable salts as active ingredients, and also contains pharmaceutically acceptable excipients.
7. The pharmaceutical preparation according to claim 6, characterized in that, The drug formulation can be administered orally or by injection, and the dosage forms are: tablets, capsules, powders, syrups, or injections.
8. The use of the diterpenoid compound of claim 1 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 5, or the pharmaceutical preparation of claim 6 or 7, in the preparation of an anti-inflammatory drug.