Squalene type triterpenoids, preparation method thereof and application of squalene type triterpenoids in anti-inflammatory drugs

Abstract

The invention discloses a squalene type triterpenoid compound, a preparation method thereof and application of the squalene type triterpenoid compound to anti-inflammatory drugs. According to the invention, the irpolacrin C with remarkable anti-inflammatory activity is separated from a rice solid fermentation culture of a casuarina equisetifolia endophytic fungus Irpus lactus SC4004, and the irpolacrin C is as shown in a formula (I) which is described in the specification. The irplacrin C is a novel squalene type triterpenoid compound, the half inhibitory concentration (IC50) of the irplacrin C is 32.53 mu M, and the IC50 value of the positive control indometacin is 29.44 mu M. The irplacrin C is a novel squalene type triterpenoid compound. The invention provides a candidate drug for research and development of novel anti-inflammatory drugs, and provides a scientific basis for development and utilization of casuarina equisetifolia endophytic fungus resources.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical biotechnology, and particularly relates to a compound irpelacerin C, a preparation method thereof, and an application thereof in the preparation of anti-inflammatory drugs. Background Art

[0002] Casuarina equisetifolia L., also known as horsetail tree or hairy beard, is a deep-rooted tree of the Casuarina family. It was introduced to China over 80 years ago. Native to Australia and the Pacific Islands, it is cultivated in Guangdong, Guangxi, and Fujian provinces of China. The traditional medicinal properties of Casuarina equisetifolia were first documented in the Xinhua Compendium of Materia Medica. Its bark, branches, leaves, and seeds are all used medicinally, primarily to treat colds and coughs, hernias, diarrhea caused by cold and dampness, urinary incontinence, and athlete's foot.

[0003] Microorganisms are a rich resource for natural product research. Advanced genome sequencing technologies reveal that microorganisms possess greater biosynthetic capabilities than previously anticipated. Plant endophytes, a highly diverse group of organisms, are considered an excellent option for sustainable agricultural development. They are gaining increasing attention due to their widespread distribution, variability, and diverse and novel chemical compositions. Among them, endophytic fungi, present in nearly every plant species, are widely considered a significant source of active natural compounds. They offer new avenues for sustainable development in medicine, agriculture, and industry, representing a valuable treasure trove worthy of exploration.

[0004] Inflammation is the immune system's first response to external stress and injury and serves as a protective mechanism. It is often accompanied by localized symptoms of redness, swelling, fever, and pain, and in extreme cases, can be fatal. Inflammation is generally characterized by the production and secretion of proinflammatory cytokines, such as interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), nitric oxide (NO), and prostaglandin E2 (PGE2). The vast majority of currently developed anti-inflammatory drugs, primarily glucocorticoids and nonsteroidal anti-inflammatory drugs (NSAIDs), are designed to inhibit the production and secretion of these proinflammatory cytokines. Due to their widespread clinical use, these drugs have been found to have adverse effects that can be detrimental to the body. Glucocorticoids can inhibit the expression of proinflammatory genes, but they carry the risk of initiating and exacerbating infections. Furthermore, long-term, high-dose use of glucocorticoids in clinical practice can lead to Cushing's syndrome, sodium and water retention, and an increased risk of hypertension. NSAIDs achieve their antipyretic and analgesic effects by inhibiting cyclooxygenase (COX) activity, thereby blocking the synthesis of prostaglandins. Because prostaglandins protect the gastric mucosa, the use of NSAIDs can cause a variety of gastrointestinal reactions, including loss of appetite, abdominal pain, and even severe myocardial infarction and stroke. Under the principle of rational medication, safer drugs should be used whenever possible. Therefore, identifying natural compounds with no toxic side effects and good efficacy to replace traditional anti-inflammatory drugs is a major challenge that needs to be addressed in the clinical treatment of inflammatory diseases.

[0005] Natural products have stronger pharmacological activity and lower toxicity, and can be used as potential resources for the development of natural anti-inflammatory drugs. Although natural products have limited therapeutic effects on the severe symptoms of certain chronic diseases, they are more effective in relieving mild pain in the early stages of these diseases. Current studies have found that natural products with anti-inflammatory activity include polysaccharides, flavonoids, polyphenols, alkaloids, terpenes, natural pigments, plant volatile oils, quinones and other compounds. In order to discover natural products with significant activity, this application isolated an endophytic fungus Irpex laceratus SC4004 from a sample of the medicinal plant Casuarina equisetifolia. By isolating and purifying the solid fermentation product of rice of this strain, we obtained a new squalene-type triterpenoid compound, which can inhibit the production of nitric oxide (NO) and has the potential to be developed into an anti-inflammatory drug. Summary of the Invention

[0006] The present invention separates and purifies a squalene-type triterpenoid compound from the fermentation product of the endophytic fungus Irpex laceratus SC4004 of the medicinal plant Casuarina equisetifolia. The present invention uses high-resolution mass spectrometry, spectroscopy (hydrogen spectrum, carbon spectrum, HSQC spectrum, HMBC spectrum and 1 H- 1Its chemical structure was confirmed by H COSY spectroscopy and spectroscopy (infrared and ultraviolet). Activity testing revealed that the compound has significant anti-inflammatory activity and has good development prospects.

[0007] The first object of the present invention is to provide a compound represented by formula (I): irpelacerin C.

[0008]

[0009] The second object of the present invention is to provide a method for preparing the compound irpelacerin C, which is isolated and prepared from the fermentation culture of the endophytic fungus Irpex laceratus SC4004 of Casuarina equisetifolia.

[0010] Preferably, the preparation method comprises the following steps:

[0011] a. Preparation of solid fermentation culture of Casuarina endophyte Irpex laceratus SC4004, the solid fermentation product was extracted with ethyl acetate, and the extract was concentrated to obtain an extract;

[0012] b. The extract was separated by macroporous adsorption resin chromatography and eluted with water, 20% ethanol by volume, 40% ethanol by volume, 60% ethanol by volume and 80% to 100% ethanol by volume;

[0013] The 80% to 100% ethanol elution fraction was collected and eluted with a normal phase petroleum ether / ethyl acetate / methanol system v:v:v, 30:1:0→0:3:1, and the petroleum ether / ethyl acetate / methanol v / v / v=2:1:0→1:1:0 gradient elution was collected and TLC thin layer chromatography with dichloromethane: methanol=10:1v / v developed to obtain a component Fr.4.5 with Rf=0.5-0.9. Fr.4.5 was eluted with a normal phase silica gel column chromatography with dichloromethane / methanol v / v=100:1→0:1 system, and the dichloromethane / methanol v / v=100:1→50:1 gradient elution was collected to obtain The obtained product was developed by TLC thin layer chromatography with dichloromethane: methanol = 50:1 v / v to obtain component Fr.4.5.3 with Rf = 0.1-0.5. After Fr.4.5.3 was eluted with petroleum ether / ethyl acetate v / v = 10:1→0:1 system on a normal phase silica gel column, the product was collected by petroleum ether / ethyl acetate v / v = 5:1→2:1 gradient elution and developed by TLC thin layer chromatography with dichloromethane: methanol = 10:1 v / v to obtain component Fr.4.5.3.2 with Rf < 0.3. Fr.4.5.3.2 was purified by Sephadex LH-20 under dichloromethane / methanol v / v, 1:3 conditions and collected by Sephadex gel column The product eluted from LH-20 and developed by TLC thin layer chromatography with dichloromethane:methanol=15:1 v / v gave component Fr.4.5.3.2.1 with Rf=0.3-0.7. Fr.4.5.3.2.1 was eluted by silica gel column chromatography with petroleum ether / ethyl acetate v / v=10:1→1:1 system, and the product obtained by petroleum ether / ethyl acetate v / v=5:1→2:1 gradient elution was collected and developed by TLC thin layer chromatography with petroleum ether / ethyl acetate=2:1 v / v system to give component F with Rf=0.5-0.8. R.4.5.3.2.1.1 and Fr.4.5.3.2.1.1 were eluted on a silica gel column using a gradient of petroleum ether / ethyl acetate (v / v) = 10:1 → 1:1. The fraction obtained by the gradient of petroleum ether / ethyl acetate (v / v) = 2:1 was collected and developed by thin-layer chromatography (TLC) with petroleum ether / ethyl acetate = 1:1 (v / v) to obtain Fr.4.5.3.2.1.1.1 with an Rf = 0.7-0.9. Fr.4.5.3.2.1.1.1 was purified by high-performance liquid chromatography (HPLC) using a mobile phase of acetonitrile / water (78:22, v / v) at a flow rate of 2 mL / min to obtain irpelacerin C.

[0014] The potato glucose liquid culture medium is prepared by the following method per liter: 200g of potatoes are boiled in 1000mL of pure water for 20 minutes, 20g of glucose, 3g of KH2PO4, 1.5g of MgSO4, and 10mg of vitamin B1 are added after filtering, and water is added to make up to 1000mL, and the culture medium is sterilized.

[0015] Preferably, the HPLC purification is performed using a YMC pack ODS-A / AQ column with a mobile phase of acetonitrile / water in a volume ratio of 78:22 at a flow rate of 2 mL / min to obtain compound irpelacerin C, t R =62min.

[0016] Preferably, the step a of preparing a solid fermentation culture of the Casuarina endophytic fungus Irpex laceratus SC4004 comprises the following steps: on a clean bench under sterile conditions, the Irpex laceratus SC4004 strain is transferred to a sterilized potato glucose liquid culture medium, and cultured on a shaking table at 28°C and 150 r / min for 7 days to obtain a fungal seed liquid, 90 mL of water is added to every 100 g of rice, and the rice culture medium is obtained by sterilization, the cultured seed liquid is poured into the sterilized rice culture medium, and incubated at 28°C for 30 days. After fermentation is completed, a solid fermentation culture is obtained.

[0017] The third object of the present invention is to provide the use of the compound irpelacerin C or a pharmaceutically acceptable salt thereof in the preparation of anti-inflammatory drugs.

[0018] Preferably, the anti-inflammatory drug is a drug that inhibits the production of NO in mouse mononuclear macrophage leukemia cells (RAW264.7) induced by lipopolysaccharide (LPS). A fourth object of the present invention is to provide an anti-inflammatory drug comprising an effective amount of the compound irpelacerin C, or a pharmaceutically acceptable salt thereof, as an active ingredient, and a pharmaceutically acceptable carrier.

[0019] Preferably, the anti-inflammatory drug is a drug that has an inhibitory effect on the production of NO in mouse mononuclear macrophage leukemia cells (RAW264.7) induced by lipopolysaccharide (LPS).

[0020] The present invention also provides use of the Casuarina equisetifolia endophytic fungus Irpex laceratus SC4004 in preparing the compound irpelacerin C.

[0021] The present invention found through experiments that the compound irpelacerin C has significant anti-inflammatory activity, with a half-maximal inhibitory concentration (IC 50) was 32.53 μM, and the IC of the positive control indomethacin was 50 The value was 29.44 μM. This result indicates that the compound irpelacerin C of the present invention has relatively significant anti-inflammatory activity.

[0022] The present invention prepares and isolates the compound irpelacerin C from the endophytic fungus Irpex laceratus SC4004 of Casuarina equisetifolia. The compound irpelacerin C has anti-inflammatory activity and can be used to prepare anti-inflammatory drugs. It provides a candidate compound for the research and development of new anti-inflammatory drugs and provides a scientific basis for the development and utilization of natural active substances of endophytic fungi in medicinal plants.

[0023] The Casuarina equisetifolia endophytic fungus Irpex laceratus SC4004 of the present invention has been disclosed in Chen C, Wei LJ, Yuan YF, Chen YC, Li SR, Lai JY, Duan FF, Qin SY, Jiang B, Wei XY, Zou ZX, Tan HB. New Squalene Triterpenes from the Fungus Irpex laceratus. Chemistry & Biodiversity, 2025 [i.e., the strain Irpex laceratus in the document]. The strain is deposited in the South China Agricultural Plant Molecular Analysis and Genetic Improvement Laboratory of the South China Botanical Garden in Guangzhou, and is guaranteed to be provided to the public within 20 years from the date of application. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 It is the compound irpelacerin C 1 H-NMR spectrum;

[0025] Figure 2 It is the compound irpelacerin C 13 C-NMR spectrum;

[0026] Figure 3 It is the compound irpelacerin C 1 H- 1 H COSY spectrum;

[0027] Figure 4 is the HSQC spectrum of compound irpelacerin C;

[0028] Figure 5 is the HMBC spectrum of compound irpelacerin C;

[0029] Figure 6This is the HRESIMS data of compound irpelacerin C;

[0030] Figure 7 The structure and 1 H- 1 H COSY and HMBC related signals. DETAILED DESCRIPTION

[0031] The following examples are provided to further illustrate the present invention, but are not intended to limit the present invention.

[0032] Example 1:

[0033] 1. Isolation, purification and identification of the endophytic fungus Irpex laceratus SC4004 from Casuarina equisetifolia

[0034] The endophytic fungus Irpex laceratus SC4004 of the present invention was isolated from Casuarina equisetifolia plants collected from Yongxing Island, Hainan Province. The strain was identified as Irpex laceratus by ITS sequencing analysis and blast comparison and named Irpex laceratus SC4004 (hereinafter referred to as strain SC4004).

[0035] 2. Solid-state fermentation of strain SC4004

[0036] Under sterile conditions on a clean bench, the Irpex laceratus SC4004 strain was transferred to sterilized potato glucose liquid medium (prepared per liter by boiling 200 g of potatoes in 1000 mL of pure water for 20 minutes, filtering, adding 20 g of glucose, 3 g of KH2PO4, 1.5 g of MgSO4, and 10 mg of vitamin B1, and then sterilizing). The culture was incubated on a shaker at 28°C and 150 rpm for 7 days to obtain a fungal seed solution. 28 kg of rice was distributed into 268 jars, and approximately 90 mL of water was added to 100 g of rice. The culture was then sterilized (autoclaved at 121°C for 20 minutes). The seed solution was then poured into sterilized jars and incubated at 28°C for 30 days. After the fermentation was completed, the rice culture medium was crushed and extracted three times with ethyl acetate and 90% ethanol aqueous solution, respectively, to obtain about 3 kg (not completely dried) of the crude extract of the fungus.

[0037] 3. Preparation of compound irpelacerin C

[0038] The crude extract (3 kg) was separated by macroporous adsorption resin chromatography (XAD-7HP) and gradient eluted with water, 20% ethanol by volume, 40% ethanol by volume, 60% ethanol by volume and 80% to 100% ethanol by volume to obtain five fractions: aqueous layer, 20% ethanol fraction, 40% ethanol fraction, 60% ethanol fraction and 80% to 100% ethanol fraction (Fr.0 to Fr.4).

[0039] Fr.4 (220 g, 80%-100% ethanol elution fraction) was eluted with a normal phase petroleum ether / ethyl acetate / methanol system (v:v:v, 30:1:0 → 0:3:1). The fraction obtained by gradient elution with petroleum ether / ethyl acetate / methanol (v / v / v = 2:1:0 → 1:1:0) was collected and developed by thin-layer chromatography (TLC) with dichloromethane:methanol = 10:1 v / v to afford fraction Fr.4.5 with an Rf = 0.5-0.9. Fr.4.5 was eluted with a normal phase silica gel column chromatography with dichloromethane / methanol (v / v = 100:1 → 0:1). The fraction obtained by gradient elution with dichloromethane / methanol (v / v = 100:1 → 50:1) was collected and developed by TLC with dichloromethane:methanol = 50:1 v / v to afford fraction Fr.4.5.3 with an Rf = 0.1-0.5. Fr.4.5.3 was eluted on a normal phase silica gel column with petroleum ether / ethyl acetate (v / v = 10:1 → 0:1). The fraction obtained by gradient elution with petroleum ether / ethyl acetate (v / v = 5:1 → 2:1) was collected and developed by thin-layer chromatography (TLC) with dichloromethane:methanol = 10:1 v / v to afford Fr.4.5.3.2 with an Rf < 0.3. Fr.4.5.3.2 (1.5 g) was purified on Sephadex LH-20 in dichloromethane / methanol (v / v, 1:3). The fraction eluted on a Sephadex LH-20 column was collected and developed by TLC with dichloromethane:methanol = 15:1 v / v to afford Fr.4.5.3.2.1 with an Rf = 0.3-0.7. Fr.4.5.3.2.1 was eluted on a silica gel column using a petroleum ether / ethyl acetate (v / v = 10:1 → 1:1) gradient elution system. The fraction obtained by gradient elution using petroleum ether / ethyl acetate (v / v = 5:1 → 2:1) was collected and developed by thin-layer chromatography (TLC) using petroleum ether / ethyl acetate = 2:1 v / v to obtain Fr.4.5.3.2.1.1. with an Rf = 0.5-0.8. Fr.4.5.3.2.1.1 was eluted on a silica gel column using a petroleum ether / ethyl acetate (v / v = 10:1 → 1:1) gradient elution system. The fraction obtained by gradient elution using petroleum ether / ethyl acetate (v / v = 2:1) was collected and developed by thin-layer chromatography (TLC) using petroleum ether / ethyl acetate = 1:1 v / v to obtain Fr.4.5.3.2.1.1.1 with an Rf = 0.7-0.9. Fr.4.5.3.2.1.1.1 was purified by semi-prep HPLC (acetonitrile / water, 78:22, v / v, 2 mL / min) and irpelacerin C (9.5 mg, t R = 62 min). TLC thin-layer chromatography was developed with petroleum ether / ethyl acetate = 2:1 (v / v). Rf = 0.6, and vanillin was colored as irpelacerin C, a dark purple compound.

[0040] 4. Structural identification of compound irpelacerin C

[0041] 1 H NMR, 13 C NMR and HMBC NMR spectra were measured using a Bruker Advance-500 NMR spectrometer with tetramethylsilane (TMS) as the internal standard; ESI-MS data were measured using a VGAutospec-3000 mass spectrometer.

[0042] like Figure 1-7 As shown, Figure 1 It is the compound irpelacerin C 1 H-NMR spectrum; Figure 2 It is the compound irpelacerinC 13 C-NMR spectrum; Figure 3 It is the compound irpelacerin C 1 H- 1 H COSY spectrum; Figure 4 is the HSQC spectrum of compound irpelacerin C; Figure 5 is the HMBC spectrum of compound irpelacerin C; Figure 6 This is the HRESIMS data of the compound irpelacerin C. Figure 7 The structure and 1 H- 1 H COSY and HMBC related signals.

[0043] Compound 1 (irpelacerin C): colorless oil. (c 0.10, MeOH); UV (CH3CN): λmax (logε): 190 (3.33) nm; HRESIMS: m / z 563.4303[M+H] + (Calculated value C 34 H 59 O6 + ,563.4306),585.4093[M+Na] + (Calculated value C 34 H 58 O6Na + ,585.4126), suggesting that its molecular formula is C 34 H 58 O6, the degree of unsaturation is 6. 1 The H NMR data (CD3OD, 500 MHz) show that there are 10 methyl signals [δ H1.11 (6H, s, H3-27 / 28), 1.60 (6H, s, H3-25 / 30), 1.62 (6H, s, H3-26 / 29), 1.67 (6H, s, H3-1 / 24) and 2.08 (6H, s, H3-2' / 2")], two oxymethine proton signals [δ H 4.92 (2H, d, J = 10.2 Hz, H-11 / 14)], four terminal olefin proton signals [δ H 5.16 (4H, m, H-7, H-3 / 22 / 7 / 18). 13 Only 16 carbon signals were found in C NMR, indicating that compound 1 has a symmetrical structure and some carbon signals overlap. 1 H and 13 In the C NMR spectrum, two groups of double bond signals (δ C / H 125.7 / 5.10) and acetyl signal (δ C 172.8, δ C / H 21.1 / 2.08), these functional groups meet the requirements of six unsaturated degrees. The above information shows that compound 1 is a chain triterpenoid compound with an interesting symmetrical structure. 1 H- 1 The H COSY spectrum shows the existence of two spin systems: a(H-3 / H2-4 / H2-5) and b(H-7 / H2-8 / H2-9). H 1.67) and H3-25(δ H 1.60) to C-2(δ C 132.1), C-3(δ C 125.4), and H-3(δ H 5.10) to C-1(δ C 25.9), C-25(δ C 17.8) HMBC related, combined 1 H- 1 H COSY fragment a, substructure A was established. H 1.62) to C-5(δ C 40.8), C-7(δ C 125.7), and H-7(δ H 5.10) to C-5(δ C 40.8), C-26(δ C 16.1) HMBC association, and combined 1 H- 1 The substructure B was further confirmed by H COSY fragment b.

[0044] Therefore, the planar structure of compound 1 was established and named irpelacerin C.

[0045] Table 1 Compound irpelacerin C 1 H(500MHz) and 13 C NMR (125 MHz) data (δ in ppm, CD3OD)

[0046]

[0047] It was determined that the chemical structure of compound 1 (irpelacerin C) is as shown in formula (I).

[0048]

[0049] Example 2:

[0050] 1. This part of the experiment uses the Griess method reported in the literature to measure the ability of the test substance to produce NO in LPS-induced mouse RAW264.7 cells, and indomethacin is used as a positive control. RAW264.7 cells are diluted to 5×10 5 cell / mL concentration, add 180 μL of cell suspension to a 96-well plate, set up 3 blank wells, and culture in a constant temperature incubator at 37°C and 5% CO2 for 1 day. After allowing it to fully adhere, remove the supernatant. Use a pipette to draw 50 μL of the supernatant from the experimental group and the control group wells, transfer them to the corresponding wells of a clean 96-well plate, add 50 μL of Griess A and Griess B reagents and mix them, and then measure the OD value at a wavelength of 540 nm. Each group of experiments was measured in parallel 3 times, and the inhibition rate of NO generation was calculated according to the following formula: NO production inhibition rate (%) = (A 对照组 -A 样品组 ) / A 对照组 ×100%.

[0051] 2. Experimental results: The prepared compound irpelacerin C has significant anti-inflammatory activity, with a half inhibitory concentration (IC 50 ) was 32.53 μM, and the IC of the positive control indomethacin was 50 The value was 29.44 μM (Table 2). These results indicate that the compound irpelacerin C has significant anti-inflammatory activity. Therefore, the present invention provides a candidate compound for the research and development of new anti-inflammatory drugs and provides a scientific basis for the development and utilization of natural active substances from plant endophytic fungi.

[0052] Table 2 Anti-inflammatory activity data of compound irpelacerin C

[0053]

Claims

1. The compound represented by formula (I): irpelacerin C, 2. A method for preparing the compound irpelacerin C according to claim 1, characterized in that: It is isolated and prepared from the fermentation culture of the endophytic fungus Irpex laceratus SC4004 of Casuarina equisetifolia.

3. The preparation method according to claim 2, characterized in that The following steps are involved: a. Preparation of solid fermentation culture of Casuarina endophyte Irpex laceratus SC4004, the solid fermentation product was extracted with ethyl acetate, and the extract was concentrated to obtain an extract; b. The extract was separated by macroporous adsorption resin chromatography and eluted with water, 20% ethanol by volume, 40% ethanol by volume, 60% ethanol by volume and 80% to 100% ethanol by volume; The 80% to 100% ethanol elution fraction was collected and eluted with a normal phase petroleum ether / ethyl acetate / methanol system v:v:v, 30:1:0→0:3:1, and the petroleum ether / ethyl acetate / methanol v / v / v=2:1:0→1:1:0 gradient elution was collected and TLC thin layer chromatography with dichloromethane: methanol=10:1v / v developed to obtain a component Fr.4.5 with Rf=0.5-0.

9. Fr.4.5 was eluted with a normal phase silica gel column chromatography with dichloromethane / methanol v / v=100:1→0:1 system, and the dichloromethane / methanol v / v=100:1→50:1 gradient elution was collected to obtain The obtained product was developed by TLC thin layer chromatography with dichloromethane: methanol = 50:1 v / v to obtain component Fr.4.5.3 with Rf = 0.1-0.

5. After Fr.4.5.3 was eluted with petroleum ether / ethyl acetate v / v = 10:1→0:1 system on a normal phase silica gel column, the product was collected by petroleum ether / ethyl acetate v / v = 5:1→2:1 gradient elution and developed by TLC thin layer chromatography with dichloromethane: methanol = 10:1 v / v to obtain component Fr.4.5.3.2 with Rf < 0.

3. Fr.4.5.3.2 was purified by Sephadex LH-20 under dichloromethane / methanol v / v, 1:3 conditions and collected by Sephadex gel column The product eluted from LH-20 and developed by TLC thin layer chromatography with dichloromethane:methanol=15:1 v / v gave component Fr.4.5.3.2.1 with Rf=0.3-0.

7. Fr.4.5.3.2.1 was eluted by silica gel column chromatography with petroleum ether / ethyl acetate v / v=10:1→1:1 system, and the product obtained by petroleum ether / ethyl acetate v / v=5:1→2:1 gradient elution was collected and developed by TLC thin layer chromatography with petroleum ether / ethyl acetate=2:1 v / v system to give component F with Rf=0.5-0.

8. R.4.5.3.2.1.1 and Fr.4.5.3.2.1.1 were eluted on a silica gel column using a gradient of petroleum ether / ethyl acetate (v / v) = 10:1 → 1:

1. The fraction obtained by the gradient of petroleum ether / ethyl acetate (v / v) = 2:1 was collected and developed by thin-layer chromatography (TLC) with petroleum ether / ethyl acetate = 1:1 (v / v) to obtain Fr.4.5.3.2.1.1.1 with an Rf = 0.7-0.

9. Fr.4.5.3.2.1.1.1 was purified by high-performance liquid chromatography (HPLC) using a mobile phase of acetonitrile / water (78:22, v / v) at a flow rate of 2 mL / min to obtain irpelacerin C. The potato glucose liquid culture medium is prepared by the following method per liter: 200g of potatoes are boiled in 1000mL of pure water for 20 minutes, 20g of glucose, 3g of KH2PO4, 1.5g of MgSO4, and 10mg of vitamin B1 are added after filtering, and water is added to make up to 1000mL, and the culture medium is sterilized.

4. The preparation method according to claim 3, characterized in that The high performance liquid phase purification was performed using a YMCpack ODS-A / AQ column with a mobile phase of acetonitrile / water in a volume ratio of 78:22 at a flow rate of 2 mL / min to obtain the compound irpelacerin C. R =62min.

5. The preparation method according to claim 3, characterized in that The step a of preparing a solid fermentation culture of the Casuarina endophytic fungus Irpex laceratus SC4004 comprises the following steps: on a clean bench under sterile conditions, the Irpex laceratus SC4004 strain is transferred to a sterilized potato glucose liquid culture medium, and cultured on a shaking table at 28°C and 150 r / min for 7 days to obtain a fungal seed liquid, 90 mL of water is added to every 100 g of rice, and the rice culture medium is obtained by sterilization, the cultured seed liquid is poured into the sterilized rice culture medium, and the culture is incubated at 28°C for 30 days. After fermentation is completed, a solid fermentation culture is obtained.

6. Use of the compound irpelacerin C according to claim 1, or a pharmaceutically acceptable salt thereof, in the preparation of an anti-inflammatory drug.

7. An anti-inflammatory drug, characterized in that The invention comprises the compound irpelacerin C or a pharmaceutically acceptable salt thereof according to claim 1 as an active ingredient.

8. Use of the endophytic fungus Irpex lacerates SC4004 from Casuarina equisetifolia in the preparation of the compound irpelacerin C according to claim 1.

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