Sterol compound, preparation method and medicine for treating pulmonary fibrosis

Abstract

The invention relates to a sterol compound, a preparation method and a medicine for treating pulmonary fibrosis, and belongs to the technical field of medicines. The demethylated cryptosterol separated from the fungus coniothyrium minitans has good anti-pulmonary fibrosis activity, and can be used as a lead compound for developing drugs for treating idiopathic pulmonary fibrosis. The invention provides an alternative compound for developing a new medicine for treating idiopathic pulmonary fibrosis, and has very important significance for development and utilization of natural product resources.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, and more specifically, relates to sterol compounds, the isolation and preparation of drugs for the treatment of idiopathic pulmonary fibrosis, and especially to the process of isolation and purification of desmethylcryptostol, structural confirmation and anti-pulmonary fibrosis activity. Background Technology

[0002] Epidemiological meta-analyses show that the global incidence of idiopathic pulmonary fibrosis (IPF) is 0.09–1.30 cases per 10,000 person-years, with a sharp increase in incidence with age. Current drug treatments, including glucocorticoids, broad-spectrum immunosuppressants, and the antifibrotic drug combination of pirfenidone / nintedanib, can only moderately slow the decline in forced vital capacity and are limited by dose-limiting toxicities. Therefore, the median survival after diagnosis remains only 3–5 years, making IPF one of the most deadly chronic respiratory diseases and placing an increasingly heavy burden on global public health systems.

[0003] Histopathologically, IPF is characterized by persistent fibroblast proliferation and continuous accumulation of pathological extracellular matrix (ECM), ultimately leading to thickening and obstruction of the alveolar septa and irreversible distortion of the lung structure. Transforming growth factor- β 1 (TGF- β 1) It plays a central role in this process, triggering a series of self-amplifying cascade reactions that promote the transdifferentiation of fibroblasts into myofibroblasts (FMT), clonal proliferation, and excessive synthesis and deposition of extracellular matrix (ECM). The resulting positive feedback loop continuously increases tissue stiffness, leading to progressive respiratory failure. The significant treatment gap underscores the urgent need to discover and develop next-generation antifibrotic drugs from natural products and synthetic chemistry.

[0004] Fungi are an important source of sterols. These compounds possess a unique tetracyclic steroid core structure and are widely found in fungal metabolites. Thanks to advances in natural product chemistry, numerous fungal sterols have been discovered and elucidated in recent years, highlighting their increasing medicinal value. Studies have shown that fungal sterols possess diverse pharmacological activities, exhibiting significant therapeutic potential in anti-inflammatory, anticancer, immunomodulatory, and cardiovascular protective effects. Based on their unique mechanism of action and excellent biological activity, fungal sterols are considered potential drug candidates for the treatment of idiopathic pulmonary fibrosis. Summary of the Invention

[0005] The objective of this invention is to provide a fungus called *Pseudomonas scutellatus*. paraconiothyriumThis invention relates to a method for isolating and purifying anti-pulmonary fibrosis compounds and their applications. The compounds in this invention are potential dual inhibitors of fibroblast activation and extracellular matrix deposition, exhibiting excellent anti-pulmonary fibrosis activity and can serve as lead compounds for the development of anti-pulmonary fibrosis drugs. According to a first aspect of this invention, a highly degraded sterol compound is provided; furthermore, this compound has been found to possess excellent anti-pulmonary fibrosis activity and shows promising development potential.

[0006] A sterol compound, the structural formula of which is shown in formula (1): Equation (1).

[0007] The method for preparing the sterol compound includes the following steps: (1) After fermenting the fungi of the genus Pseudomonas with rice, the alcohol was extracted, the alcohol was recovered by vacuum concentration, and then extracted with ethyl acetate to obtain the ethyl acetate extract. (2) The ethyl acetate fraction obtained in step (1) was subjected to column chromatography and then eluted with petroleum ether-ethyl acetate gradient to obtain 7 components with increasing polarity. The 6th component was then separated by gel chromatography, silica gel column chromatography and high performance liquid chromatography to obtain the sterol compound.

[0008] The pseudoscuta mold paraconiothyrium The strain was purchased from the China Marine Microbial Culture Collection Center, and its strain number is MCCC 3A00275.

[0009] The fungus *Pseudomonas scutellatus* was inoculated into a solid rice fermentation medium and cultured at 25-28 ℃ for 28 days.

[0010] The alcohol mentioned in step (1) is ethanol, and the mass concentration of the alcohol is 95% or higher.

[0011] In step (2), petroleum ether-ethyl acetate-methanol gradient elution is performed under the following conditions: 50-1:1:0-1.

[0012] Separation was achieved through gel chromatography, normal-phase and reverse-phase silica gel column chromatography, and high-performance liquid chromatography. Gel chromatography was performed using a 1:1 (v / v) dichloromethane-methanol elution solution. The separation conditions for silica gel column chromatography were elution with petroleum ether-ethyl acetate-methanol at a volume ratio of 20-1:1:0-1. The high-performance liquid chromatography (HPLC) separation conditions are methanol-water elution with a volume ratio of 50-75:25-50.

[0013] A drug for treating pulmonary fibrosis, comprising the aforementioned sterol compounds.

[0014] The medications for treating pulmonary fibrosis also include pharmaceutically acceptable excipients.

[0015] The use of the sterol compounds in the preparation of drugs with anti-pulmonary fibrosis activity and / or for the treatment of pulmonary fibrosis.

[0016] In summary, compared with the prior art, the above-described technical solutions conceived by this invention mainly possess the following technical advantages: 1. From fungi paraconiothyrium Sterol compounds were found in sp.; 2. Further bioactivity evaluation results showed that the new compound has a strong inhibitory effect on fibroblast activation and can effectively combat idiopathic pulmonary fibrosis. Attached Figure Description

[0017] Figure 1 This is the single-crystal diffraction pattern of the compound demethylcryptostol A2. Figure 2 This is the hydrogen spectrum of compound demethylcryptostol A2. Figure 3 This is the carbon spectrum of compound demethylcryptostol A2. Figure 4 The CCK-8 assay was used to verify the anti-fibrotic pharmacological effects of compound desmethylcryptostol A2; where: A represents the cytotoxicity of compound desmethylcryptostol A2 on NIH / 3T3 cells; B represents the TGF-β-Cytotoxicity of compound desmethylcryptostol A2 on NIH / 3T3 cells. β 1. Cell proliferation of activated NIH / 3T3 cells was detected by CCK-8 assay after 48 hours of treatment with different concentrations of the compound demethylcryptostol A2 or the solvent. Figure 5 The cell proliferation of activated and quiescent cells was assessed by the EdU assay after 48 hours of treatment with different doses of the compound desmethylcryptostol A2 or the solvent. Figure 6 Cell migration ability was assessed using a wound healing assay under the same treatment conditions. Figure 7 To analyze and detect [the virus] using Western blot under the same experimental protocol. α -SMA and COL1A1 protein levels. Figure 8 The study used methods such as weight monitoring, lung coefficient calculation, and hydroxyproline content determination to verify the effect of the compound desmethylcryptostrol A2 in improving the pathological changes of bleomycin (BLM)-induced pulmonary fibrosis in vivo. Among them: A is the change in mouse body weight throughout the experiment; B is the lung coefficient of mice calculated at the end of the experiment; C is the quantitative detection of hydroxyproline content in lung tissue to assess collagen deposition. Figure 9This study uses tissue staining and pathological scoring methods to verify the effect of compound desmethylcryptostol A2 in improving the pathological changes of bleomycin (BLM)-induced pulmonary fibrosis in vivo. Among them, A is a representative micrograph of lung tissue sections stained with hematoxylin-eosin (H&E) and Masson trichrome; BC is a semi-quantitative analysis of lung tissue pathological changes using fibrosis scoring and Ashcroft scoring. Detailed Implementation

[0018] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention. Furthermore, the technical features involved in the various embodiments of this invention described below can be combined with each other as long as they do not conflict with each other.

[0019] The compound isolated and purified from the fungus *Pseudomonas* in this invention belongs to the sterol class of compounds, and its chemical name is (3a). S ,5a R 6 R ,8a R )-6-((2 R 5 S , E )-5,6-dimethylhept-3-en-2-yl)-3a-hydroxy-5a-methyl-3a,4,5,5a,6,7,8,8a-octahydro-2H-indeno[5,4-b]furan-2-one was named demethylcryptostrol A2, and its structural formula is shown in Formula 1;

[0020] Formula 1 The method for preparing sterol compounds in this invention includes the following steps: extracting rice fermentation product of the fungus *Pseudomonas* genus with industrial alcohol (95% ethanol), concentrating under reduced pressure to recover the industrial alcohol, and then extracting with ethyl acetate to obtain an ethyl acetate fraction extract; subjecting the ethyl acetate fraction extract to column chromatography: specifically, mixing with silica gel and dry packing the column; then eluting with a gradient of petroleum ether-ethyl acetate, combining the same components to obtain a total of 7 components with polarity from smallest to largest; wherein component 6 is further separated by repeated gel chromatography, reversed-phase silica gel column chromatography, and high-performance liquid chromatography to obtain the compound demethylcryptostrol A2.

[0021] The application of sterol compounds in the anti-pulmonary fibrosis of this invention, wherein the compounds can inhibit TGF-β through targeted inhibition. βIt drives fibroblast activation and extracellular matrix production, thereby inhibiting fibrosis-related inflammatory responses and suppressing fibroblast proliferation and migration, reducing collagen deposition, thus alleviating symptoms of lung tissue structural damage and functional impairment caused by pulmonary fibrosis.

[0022] In summary, this invention, through the application of... paraconiothyrium sp . The fermentation broth was extracted and purified by column chromatography to obtain one sterol compound. paraconiothyrium sp . Secondary metabolites. Their structures were determined using various spectroscopic analysis techniques and other methods, as shown in Formula 1. Evaluation of the anti-pulmonary fibrosis activity of the compound of Formula 1 revealed that it significantly inhibited TGF-β. β The 1-driven fibroblast activation and extracellular matrix production have therapeutic activity against pulmonary fibrosis and can serve as a lead compound for the development of drugs to treat pulmonary fibrosis.

[0023] The following are specific embodiments. Example 1 1. Isolation and preparation of compound demethylcryptostrol A2 as shown in Formula 1 The fungi of the genus *Pseudomonas* were inoculated into 100 kg of rice solid fermentation medium and cultured at 28 ℃ for 28 days. The rice fermentation product was extracted 8 times with industrial alcohol. The industrial alcohol was concentrated and recovered under reduced pressure at below 50 ℃ and then extracted with ethyl acetate to obtain 1.12 kg of ethyl acetate extract.

[0024] The ethyl acetate fraction extract was subjected to column chromatography: specifically, the sample was mixed with 200-300 mesh silica gel and dry-packed into a column, eluted with a gradient of petroleum ether-ethyl acetate-methanol (50:1:1), detected by TLC, and the same components were combined to obtain 7 fractions with increasing polarity. Fraction 6 was then separated by repeated gel chromatography, normal and reverse phase silica gel column chromatography, and high performance liquid chromatography (gel chromatography separation conditions were elution with dichloromethane-methanol at a volume ratio of 1:1; normal phase silica gel column chromatography separation conditions were elution with petroleum ether-ethyl acetate-methanol at a volume ratio of 20:1:1; high performance liquid chromatography separation conditions were elution with methanol-water at a volume ratio of 75:25) to obtain compound desmethylcryptostrol A2 (113.2 mg), which is the compound of formula 1 of this application.

[0025] 2. Structural identification of compound demethylcryptostrol A2 as shown in Formula 1 Nuclear magnetic resonance (NMR) and mass spectrometry (MS) analyses of compound demethylcryptostol A2 revealed that its proton and carbon spectra were similar to those of compounds reported in the literature. 1The high-field methyl region of its ¹³C NMR spectrum, except for the H3-19 methyl singlet, shows the characteristic methyl signal of steroids. However, its ¹³C NMR data shows only 21 carbon atoms, indicating that it is chemically a degradation sterol. 13 C NMR in δ Signals were observed at 173.6, 172.9, 112.6, and 104.1, all indicating the presence of a signal within the molecule. γ -hydroxy- α , β -Unsaturated γ - Lactone system, 1 H NMR spectrum δ The triple and double peaks at 5.36 and δ The doublet doublet at 5.24 was identified as the olefin protons H-15 and H-16 in the side chain. Further X-ray single-crystal diffraction data analysis of demethylcryptostol A2 confirmed the molecular formula of the compound as C2. 21 H 32 Based on O3, combined with the Flake parameters and chiral center data, the absolute stereoconfiguration of this molecule can be confirmed as 4. S 7 R 8 R ,11 R ,13 R 15 E 17 S This confirmed that the structure of the compound was consistent with literature reports. The crystal structure is as follows: Figure 2 As shown, the crystal structure data of compound demethylcryptostol A2 is as follows. 13 C NMR and 1 The 1H NMR data are shown in Table 1. The NMR 1H spectrum is shown in Table 1. Figure 2 As shown, the carbon spectrum is as follows Figure 3 As shown.

[0026] Compound demethylcryptostol A2: Molecular formula: C 21 H 32 O3, molecular weight: 332.46 g / mol, unit cell parameters: a = 7.98040(10) Å, b = 6.67820(10) Å, c = 18.6960(2) Å, V = 993.15(2) Å 3 , α = 90°, β = 94.6260(10)°, γ = 90°, cell volume (V): 993.15(2) ų, temperature (T): 99.99(10) K, space group: P21, number of molecules in the cell (Z): 2, absorption coefficient ( μ Cu Kα): 0.568 mm -1 Number of diffraction points measured: 22315; Number of independent diffraction points: 3860; Diffraction point repeatability (… R int ): 0.0486, structural refinement result: R 1 (I>2) σ ( I ): 0.0337, wR ( F 2 )( I >2 σ ( I ): 0.0881, R 1 (Full Data): 0.0339, wR ( F 2 (Full data): 0.0883, Goodness of fit (GOFon) F 2): 1.049, Flack parameter: 0.10(10).

[0027] Table 1. Compounds of Formula (1) 1 H-NMR (400MHz, CD3OD) and 13 C-NMR (100MHz, CD3OD) data

[0028] Example 2 1. In vitro pharmacodynamics and mechanism studies (1) For TGF- β 1. Evaluation of the inhibitory activity of induced fibroblast proliferation The basic safety of compound demethylcryptostrol A2 in fibroblasts and its specific antagonistic effect on TGF-β were verified using methods such as CCK-8 assay and EdU detection. β 1. Activity of the proliferative effect.

[0029] CCK-8 method: NIH / 3T3 cells were seeded at a density of 8 × 10³ cells / well in 96-well plates, treated with a series of compound concentrations for 48 hours, and then analyzed according to the kit instructions. In constructing TGF-... β In the induced pulmonary fibrosis cell model, NIH / 3T3 cells (density 5×10⁻⁶) were used. 4 Cells (100 μL / mL) were seeded into each well of a 96-well plate. After cell adhesion, the plates were divided into a control group and a model group (containing 5 ng / mL TGF-β). β 1) and compound treatment group (in TGF- βDifferent concentrations of the compound were added in the presence of 1. After treatment, CCK-8 reagent was added to each well and incubated for 1 hour, and the absorbance was measured at 450 nm. Cell viability was calculated using the formula: Cell viability (%) = [(As - Ab) / (Ac - Ab)] × 100%, where As, Ac, and Ab are the absorbance values ​​of the experimental group, normal control group, and blank well (containing only culture medium), respectively. The effect of the compound on TGF-β was calculated based on the absorbance values. β 1. The inhibition rate of induced proliferation was determined, and its dose-response relationship was analyzed using GraphPad Prism software. For example... Figure 4 As shown in Figure A, the toxicity test results indicated that at concentrations below 2.5 μM, the cell viability of the demethylcryptostol A2 treatment group was not statistically different from that of the normal control group, indicating that the compound had no significant cytotoxicity within this concentration range. Figure 4 As shown in Figure B, in the inhibition activity experiment, TGF- β Cell viability in the stimulation group was significantly higher than that in the normal control group, confirming that it successfully induced cell proliferation; while demethylcryptosidol A2 significantly reversed TGF-β in a concentration-dependent manner. β This proliferative effect of 1. At a concentration of 1.25 μM, it has an effect on TGF-β. β The inhibition rate of induced proliferation reached a significant level, and the inhibitory effect was further enhanced with increasing concentration, which was better than the positive control SB431542.

[0030] 5-Ethynyl-2'-deoxyuridine (EdU) labeling detection method: NIH / 3T3 cells were labeled with 1.5 × 10⁻⁶ cells. 5 Cells were seeded at a density per well in 6-well plates. After cell adhesion and 80% confluence, the cells were divided into control and experimental groups, and both groups were treated with 10 ng / mL LTGF-1. β 1. The experimental group was treated with gradient concentrations of the compound for 48 hours. Subsequently, it was incubated with EdU for 2 hours, fixed at room temperature, stained using an EdU cell proliferation assay kit, and quantitatively analyzed using ImageJ software. (Example: ...) Figure 5 As shown in the fluorescence microscopy images, compared with the normal control group, TGF-β... β The number of EdU-positive (red fluorescent) cells was significantly increased in the stimulation group, which directly confirmed the presence of TGF-β. β The proliferative effect of 1. And demethylcryptostol A2 and TGF-β. β In the co-treatment group, the proportion of EdU-positive cells decreased in a dose-dependent manner. Quantitative analysis data further confirmed that at a concentration of 1.25 μM, desmethylcryptosidol A2 significantly reduced TGF-β. β The induced EdU incorporation rate was inhibited in a concentration-dependent manner.

[0031] (2) Regarding TGF-β Evaluation of the inhibitory activity of induced fibroblast migration ability The scratch assay was used to simulate key steps in the migration of fibroblasts to the injury site during the fibrosis process, and to examine the intervention effect of compounds on cell motility.

[0032] Scratch healing assay: NIH / 3T3 cells in logarithmic growth phase were cultured at 5 × 10⁶ cells per well. 5 Cells were seeded at a density of [number] cells per well in pre-marked 12-well plates and cultured overnight at 37°C and 5% CO2 until a dense monolayer formed. Uniform scratches were made using the tip of a 200 μL sterile pipette. After washing with PBS to remove detached cells, the medium was replaced with low-serum medium containing 1% FBS (to reduce the impact of basal proliferation), and different treatments were added: control group, TGF-β-[treatment name missing]. β 1 (5 ng / mL) stimulation group, and TGF- β Group 1 was co-treated with different concentrations of compounds. Images were taken at the same location at 0 and 24 hours after scratching, and the scratch area was quantified using ImageJ software. The scratch healing rate (%) was calculated using the formula: [(A0- A 24 [) / A0] × 100%, where A0 is the initial scratch area at hour 0, A 24 This represents the remaining area of ​​the scratch after 24 hours. For example... Figure 6 As shown, after 24 hours of culture, the control group exhibited slower cell migration and a lower scratch closure rate due to the low serum environment. TGF- β In the stimulation group, cells at the edge of the scratches extended significantly into the blank area, and the scratch closure rate was significantly higher than that in the control group, confirming that TGF-β- β 1. It strongly promoted cell migration. In contrast, the extension of cells toward the scratch center was significantly inhibited in groups treated with different concentrations of desmethylcryptosidol A2. Even at a concentration of 1.25 μM, the compound significantly inhibited scratch healing, showing a dose-dependent trend.

[0033] (3) Regulatory effects on key signaling pathways and effector protein expression in fibrosis Based on the confirmed effects of the compound on cell function (proliferation, migration), the effect of demethylcryptostol A2 on TGF-β was directly detected and quantified at the protein molecular level. β 1. To investigate the regulatory role of key effector molecules in the downstream core fibrosis signaling pathway, thereby elucidating the specific molecular mechanism by which they exert antifibrotic activity.

[0034] Extracted from different treatments (control group, TGF-) β 1. Stimulation group, TGF- βTotal protein from NIH / 3T3 cells (treated with different concentrations of compounds) was collected by electrophoresis, transfer, and blocking. The cells were then sequentially incubated with primary and secondary antibodies targeting α-smooth muscle actin, type I collagen α1 chains, and the internal reference protein GAPDH. Finally, the cells were visualized using chemiluminescence. Figure 7 As shown, compared with the control group, TGF- β Stimulation significantly upregulated the protein expression levels of α-SMA and COL1A1. Meanwhile, demethylcryptosidol A2 and TGF-β... β Co-treatment with COL1A1 significantly reduced the expression levels of the aforementioned fibrosis marker proteins in a dose-dependent manner, reversing this upregulation effect. Quantitative analysis showed that at a concentration of 1.25 μM, the expression level of COL1A1 protein recovered to near or even below the basal state. This result confirms that demethylcryptostrol A2 inhibits TGF-β-reactive protein expression. β The signal can directly block key processes that lead to cell activation into myofibroblasts (decreased α-SMA expression) and excessive extracellular matrix deposition (decreased COL1A1 expression), thus elucidating the molecular basis of its antifibrotic effect.

[0035] 1. In vivo pharmacodynamic validation Based on the established in vitro mechanism, the anti-pulmonary fibrosis activity of desmethylcryptosidol A2 was further evaluated at the whole animal level. A pulmonary fibrosis model of C57BL / 6 mice induced by bleomycin intratracheal infusion was established. A model group, a desmethylcryptosidol A2 treatment group, and a normal control group were set up. Body weight was monitored regularly. After euthanasia, the body weight and wet weight of the separated and washed lung tissue were measured. The lung coefficient was calculated as (lung wet weight / body weight) × 100%. Lung tissue was fixed in 4% paraformaldehyde, embedded in paraffin, and sectioned. HE staining and Masson staining were performed according to the kit instructions. After mounting, the sections were used for later use. The degree of inflammatory infiltration, collagen deposition, and structural damage in the stained sections was scored according to the Ashcroft criteria.

[0036] like Figure 8 As shown, desmethylcryptosidol A2 has a comprehensive therapeutic effect on bleomycin-induced pulmonary fibrosis models. The progressive weight loss trend in the treated mice was significantly alleviated, effectively improving disease-related systemic wasting; the abnormal increase in the lung coefficient was also significantly suppressed, directly indicating a reduction in the pathological burden on lung tissue. Crucially, the fibrosis pathological score of the lung tissue in the treated group was significantly reduced, quantitatively confirming its ability to effectively reverse structural remodeling and abnormal repair of lung tissue.

[0037] like Figure 9As shown, HE and Masson staining results of lung tissue provide direct histopathological evidence. HE staining in the model group revealed severe alveolar structural damage, extensive inflammatory cell infiltration, and interstitial thickening; Masson staining showed widespread and dense collagen fiber deposition (large and deep blue-stained areas). In contrast, the desmethylcryptostol A2 treatment group showed better alveolar structural integrity, reduced inflammatory infiltration, significantly smaller and lighter blue-stained areas in Masson staining, and significantly reduced collagen deposition. These results, corroborated by macroscopic indicators, confirm that this compound intervenes in the core pathological processes of pulmonary fibrosis through multiple targets: protecting alveolar structure, inhibiting inflammatory responses, and reducing abnormal collagen deposition.

[0038] 2. Results and Analysis: In summary, the compound desmethylcryptostol A2 provided by this invention can intervene in the pathological process of pulmonary fibrosis through multiple targets and multiple links: in vitro, it can effectively inhibit the proliferation and migration of fibroblasts and the transdifferentiation of myofibroblasts; in vivo, it can significantly improve inflammatory infiltration and fibrotic pathological damage in lung tissue. Therefore, desmethylcryptostol A2 has significant pharmaceutical development value and can be used to prepare drugs for the treatment of pulmonary fibrosis and related fibrotic diseases.

[0039] Those skilled in the art will readily understand that the above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A sterol compound, characterized by, The structural formula of the sterol compound is shown as formula (1): Formula (1).

2. The method for preparing sterol compounds according to claim 1, characterized in that, The method comprises the following steps: (1) After the Aschersonia sp. is fermented with rice, alcohol extraction is performed, alcohol is recovered after concentration under reduced pressure, and ethyl acetate extraction is performed to obtain an ethyl acetate extract; (2) The ethyl acetate extract obtained in step (1) is subjected to column chromatography, and then gradient elution is performed with petroleum ether-ethyl acetate-methanol to obtain seven components with decreasing polarity, and the sixth component is sequentially subjected to gel chromatography separation, silica gel column chromatography separation and high performance liquid chromatography separation, thereby obtaining the sterol compound.

3. The production method according to claim 2, wherein the proposed sphaerulina paraconiothyrium sp. was purchased from China General Microbiological Culture Collection Center with the accession number MCCC 3A00275.

4. The production method according to claim 3, wherein The Aschersonia sp. is inoculated into a rice solid fermentation medium and cultured at a temperature of 25-28°C for 28 days.

5. The production method according to claim 2, wherein In step (1), the alcohol is ethanol, and the mass concentration of the alcohol is greater than 95%; in step (2), the gradient elution is performed with petroleum ether-ethyl acetate-methanol, and the gradient elution conditions are 50-1:1:0-1.

6. The production method according to claim 2, wherein The gel chromatography separation, normal and reversed-phase silica gel column chromatography separation and high performance liquid chromatography separation; the gel chromatography separation is performed with dichloromethane-methanol with a volume ratio of 1:1 as the eluent; The silica gel column chromatography separation is performed with petroleum ether-ethyl acetate-methanol with a volume ratio of 20-1:1:0-1 as the eluent; The high performance liquid chromatography separation is performed with methanol-water with a volume ratio of 50-75:25-50 as the eluent.

7. A medicament for treating pulmonary fibrosis, characterized by, The sterol compound of claim 1.

8. The medicament according to claim 7, characterized in that, The drug for treating pulmonary fibrosis further comprises a pharmaceutically acceptable excipient.

9. The sterol compound of claim 1 for use in the preparation of a drug with anti-pulmonary fibrosis activity.

10. Use according to claim 9, wherein The application of the sterol compound in the preparation of a drug for treating pulmonary fibrosis.

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