Compound produced by mangrove deposit mud-derived fungi, and preparation method and application thereof

Abstract

The invention relates to the technical field of medicines, and particularly discloses a benzaldehyde compound produced by mangrove forest deposited mud-derived fungi as well as a preparation method and application of the benzaldehyde compound. The structure of the compound is shown as a formula (I). The compound (I) provided by the invention can significantly inhibit LPS-induced NO release, and has low cytotoxicity and good cytocompatibility. In addition, the compound provided by the invention can be prepared by adopting a fungus Pseudalleschelia angusta SYSU-M4 from mangrove forest sedimentary mud, and the extraction and separation method of the compound is simple and low in cost.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to a benzaldehyde compound produced by sediment mud-derived fungi, and a preparation method and application thereof. Background Art

[0002] Alzheimer's disease (AD) is a chronic, progressive neurodegenerative disorder and one of the most common forms of dementia in the elderly. Its primary clinical manifestations are memory loss, cognitive decline, and abnormal changes in behavior and mood. Although the exact etiology of AD remains unclear, research suggests that the disease may be closely related to abnormal neuronal metabolism and abnormal protein aggregation in the brain. Neuronal loss and abnormal deposition of β-amyloid protein (Aβ), forming so-called "senile plaques," are observed in the brain tissue of AD patients. Furthermore, hyperphosphorylation of the tau protein leads to the formation of neurofibrillary tangles, accompanied by the proliferation of glial cells. These pathological features are considered to be important mechanisms of disease progression.

[0003] BV-2 cells are a mouse microglial cell line commonly used to study neurodegenerative diseases such as Parkinson's disease, Alzheimer's disease, and multiple sclerosis. In AD, the deposition of amyloid-β (Aβ) can activate BV-2 cells, triggering an inflammatory response, including the release of inflammatory cytokines (such as TNF-α, IL-1β, and IL-6) and the production of reactive oxygen species (ROS). These cytokines and substances can further exacerbate neuronal damage and death. Several studies have used BV-2 cells as a model to investigate the neuroinflammatory mechanisms of AD. For example, one study found that Aβ can activate BV-2 cells, leading to the release of inflammatory cytokines and activation of the NF-κB signaling pathway. Furthermore, studies have explored the possibility of alleviating AD symptoms by inhibiting the inflammatory response in BV-2 cells. Currently, the effects of drugs on cell viability are assessed using methods such as the MTT assay or Annexin V / PI staining. Next, the effects of the drugs on LPS-induced inflammatory cytokines (such as TNF-α and IL-1β) were measured using ELISA or qRT-PCR, while the effects on LPS-induced NO release were measured using the Griess assay. Furthermore, the effects of the drugs on LPS-induced morphological changes in BV-2 cells, such as cell flattening and swelling, were observed. Finally, the regulatory effects of the drugs on key signaling pathways (such as NF-κB and MAPK) were examined by Western blot or qRT-PCR. These methods comprehensively assess the anti-inflammatory activity of the drugs in BV-2 cells and their underlying mechanisms, providing strong support for in-depth investigation of the drugs' anti-inflammatory effects.

[0004] On the other hand, among similar azole compounds, studies on similar compound (ⅠⅠ) have shown that this analog has cholesterol-inhibiting activity. However, there are no reports showing whether it has anti-inflammatory effects.

[0005]

[0006] (ⅠⅠ) Summary of the Invention The technical problem to be solved by the present invention is to provide a compound produced by fungi derived from mangrove sediment mud with medicinal value, as well as a preparation method and application thereof.

[0007] Specifically, the present invention provides a compound represented by formula (I):

[0008] (I) In addition, according to some common methods in the technical field of the present invention, the compound of formula (I) can be reacted with a base to form a stable pharmaceutically acceptable salt. The base can be an organic base or an inorganic base, exemplified by, but not limited to, sodium hydroxide, potassium hydroxide, sodium methoxide, sodium ethoxide, and the like.

[0009] Specifically, conventional excipients are added to the compounds of the present invention and prepared according to conventional processes into various pharmaceutically acceptable dosage forms, such as tablets, capsules, oral solutions, injections, ointments, granules, suspensions, or sustained-release formulations, but not limited thereto. Excipients may include, but not limited to, adhesives, lubricants, disintegrants, diluents, solubilizers, stabilizers, and suspending agents commonly used in the pharmaceutical field.

[0010] The inventors' studies on the compound represented by formula (I) have demonstrated that it significantly inhibits lipopolysaccharide-induced inflammatory responses in BV-2 cells, including the production of inflammatory mediators such as nitric oxide. Furthermore, the compound further mitigates inflammatory responses by inhibiting signaling pathways in BV-2 cells. These properties suggest that compound I has potential clinical application in the treatment of neuroinflammatory diseases, such as Alzheimer's disease and Parkinson's disease.

[0011] Based on this, the present invention provides several uses of the compound of formula (I): (1) Use of the compound represented by formula (I) and its pharmaceutically acceptable salt as an active ingredient in the preparation of anti-inflammatory drugs.

[0012] (2) Use of the compound represented by formula (I) and its pharmaceutically acceptable salt as an active ingredient in the preparation of a drug for inhibiting the production of NO in cells.

[0013] (3) Use of the compound represented by formula (I) and its pharmaceutically acceptable salt as an active ingredient in the preparation of drugs for neuroinflammatory diseases.

[0014] (4) Use of the compound represented by formula (I) and its pharmaceutically acceptable salt as an active ingredient in the preparation of a drug for treating Parkinson's disease.

[0015] (5) Use of the compound represented by formula (I) and its pharmaceutically acceptable salt as an active ingredient in the preparation of a drug for treating Alzheimer's disease.

[0016] Accordingly, the present invention also discloses a method for preparing the above compound, which comprises: Pseudallescheria angusta SYSU-M4 is inoculated into a culture medium and obtained through fermentation, extraction, and separation. The present invention prepares the compound represented by formula (I) through fungal fermentation, and the raw materials are widely available, allowing for large-scale production. The preparation method is simple and the production cost is low.

[0017] Wherein, the mangrove sediment mud origin fungus Pseudallescheria angusta SYSU-M4 was deposited in the Guangdong Provincial Microbiological Culture Collection on May 7, 2025, with the deposit number GDMCC No: 66263. The address of the Guangdong Provincial Microbiological Culture Collection is: 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou.

[0018] The culture medium may be beef peptone culture medium, rice culture medium, potato dextrose culture medium, etc., but is not limited thereto.

[0019] Preferably, in some embodiments, the method for preparing the compound comprises: (1) Fungi derived from mangrove sediments Pseudallescheria angusta SYSU-M4 is inoculated into a culture medium for fermentation to obtain a fermentation product; (2) extracting the fermentation product with an extraction solvent, extracting with an extraction solvent, separating with a separation solvent, and separating with semi-preparative HPLC to obtain; The extraction solvent is methanol and / or dichloromethane; the extraction solvent is ethyl acetate; the separation solvent is one or more of petroleum ether, ethyl acetate, dichloromethane, methanol, chloroform, and n-hexane; the mobile phase of the semi-preparative HPLC is acetonitrile and water, and the volume ratio of acetonitrile to water is (20~30): (70~80).

[0020] More preferably, in some embodiments, step (1) comprises: (1.1) Mangrove sediment-derived fungi Pseudallescheria angusta SYSU-M4 activation; Specifically, the mangrove sediment mud-derived fungus Pseudallescheria angusta SYSU-M4 was activated in an incubator with a temperature of 20-30°C and a humidity of 60-90% (RH).

[0021] (1.2) The activated mangrove sediment fungi Pseudallescheria angusta SYSU-M4 was inoculated into liquid culture medium and cultured to obtain seed solution; The liquid culture medium may be PDB medium, SBM medium, Martin medium, etc., but is not limited thereto. Preferably, in some embodiments, the liquid culture medium is PDB medium, which is composed of: 200-300 g PDB / L, 20-40 g salt / L. PDB is composed of glucose and potato in a weight ratio of 1: (5-10). The culture medium is heated at 0.5×10 5 ~3×10 5 Sterilize at 40°C for 10 to 40 minutes.

[0022] Specifically, the inoculum size is 10-20 g of colony plate per 500 mL of culture medium. After inoculation, culture the culture medium at 28°C on a shaker (100-300 rpm) for 3-4 days to obtain a seed solution.

[0023] (1.3) Inoculate the seed liquid into a solid culture medium and ferment at 20-30°C for 30 days to obtain the fermentation product.

[0024] Specifically, the solid culture medium can be beef peptone culture medium, rice culture medium, potato dextrose culture medium, etc., but is not limited thereto. Preferably, in some embodiments, the solid culture medium is rice culture medium, and its specific composition is: Specifically, the composition of the rice culture medium is: 30-100 g rice / 50-70 mL seawater, where the sea salt content in the seawater is 10-30 g / L. Preferably, it is 50 g rice / 50 mL seawater, where the sea salt content in the seawater is 30 g / L. The culture medium is sterilized before inoculation. Specifically, at 121°C, 1×10 5 Pa sterilization for 20~40min.

[0025] Specifically, the inoculation amount of the seed liquid is 1-10 vol%, preferably 5 vol%. The fermentation temperature is 20-35° C., and the fermentation time is 10-50 days; preferably, the fermentation temperature is 25-30° C., and the fermentation time is 30 days.

[0026] More preferably, in some embodiments, step (2) comprises: (2.1) Extracting the fermentation product 1 to 3 times with an extraction solvent, combining the extracts and drying to obtain a first crude extract; The extraction solvent is methanol and / or dichloromethane, but is not limited thereto, preferably methanol.

[0027] (2.2) dissolving the first crude extract in water, then extracting with ethyl acetate 5 to 6 times, combining the extracts and drying to obtain a second crude extract; (2.3) eluting the second crude extract using a normal phase silica gel column to obtain a first intermediate product; Among them, the silica gel packing used in the forward silica gel column is 100~200 mesh, and the eluents used are petroleum ether-ethyl acetate with volume ratios of 10:0, 8:2, 7:3, 6:4, 5:5, and 4:6, and ethyl acetate-methanol with volume ratios of 10:0, 1:1, and 0:10.

[0028] Specifically, 36 subfractions (Fr.1-36) were obtained by normal silica gel column chromatography. The 36 subfractions were combined and analyzed by thin layer chromatography (TLC, developing solvent: petroleum ether: ethyl acetate = 7:3, thin layer silica gel plate, color developer: vanillin), and the relative migration value (R f values) are as follows: R of Fr.A (subcomponents 1-10) f The value is 0.9, the R of Fr.B (subcomponents 11-16) f The value is 0.8, R of Fr.C (subcomponents 17-20) f The value is 0.65, the R of Fr.D (subcomponents 21-24) f The value is 0.4, the R of Fr.E (subcomponents 25-29) f The value is 0.2. f The main component Fr.C with a value of 0.65 serves as the first intermediate product.

[0029] (2.4) eluting the first intermediate product using a gel column to obtain a second intermediate product; The matrix of the gel column is agarose gel with a pore size of 25 to 100 μ More specifically, the gel column is Sephadex LH-20 and the eluent used is dichloromethane-methanol in a volume ratio of 1:1.

[0030] Specifically, five components (Fr.CA to Fr.CE) were obtained by gel column chromatography. Thin layer chromatography (TLC, developing solvent: petroleum ether: ethyl acetate = 7:3, thin layer silica gel plate, color developer: vanillin) was used to analyze the 50 subcomponents obtained, and the relative migration value (R f ) values are as follows: R of Fr.CA (subfractions 1-10) f The value is 0.9, the R of Fr.CB (subcomponents 11-20)f The value is 0.7, the R of Fr.CC (subfractions 21-30) f The value is 0.7, R of Fr.CD (subcomponents 31-40) f The value is 0.65, and the R f The value is 0.65. f The component Fr.CB with a value of 0.7 serves as the second intermediate product.

[0031] (2.5) Separating the second intermediate product by semi-preparative HPLC to obtain a compound; The mobile phase of the semi-preparative HPLC was acetonitrile and water in a volume ratio of 30:70, with a flow rate of 3 mL / min. The chromatographic column used was Aglient-XB-C18, with a column length of 250 mm and a diameter of 10 mm. The particle size of the stationary phase in the chromatographic column was 5 μ m; the residence time of the compound is 11 min.

[0032] The implementation of the present invention has the following beneficial effects: The compound (I) provided by the present invention has low cytotoxicity at different concentrations, good cell compatibility, and μ g / mL significantly inhibited LPS-induced NO release (p < 0.05), and has broad application prospects in the preparation of drugs for the treatment of Alzheimer's disease. In addition, the invented compound is derived from fungi derived from mangrove sediment mud. Pseudallescheria angusta SYSU-M4, its extraction and separation method is simple and low-cost. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1 is the ethyl group in Example 1 of the present invention ( S )-2-[( R H NMR spectrum of 3-methyl-2,5-dioxopyrrolidin-1-yl]propanoate; Figure 2 is the ethyl group in Example 1 of the present invention ( S )-2-[( R )-3-methyl-2,5-dioxopyrrolidin-1-yl] propionate; Figure 3 is the ethyl group in Example 1 of the present invention ( S )-2-[( R )-3-methyl-2,5-dioxopyrrolidin-1-yl]propanoate 1 H- 1 Schematic diagram of H COSY and HMBC; Figure 4 is the ethyl group in Example 1 of the present invention ( S )-2-[( R Two possible stereoisomers of 3-methyl-2,5-dioxopyrrolidin-1-yl]propanoate 1 H / 13 C NMR calculation results (revTPSS / pcSseg-1 theoretical level); Figure 5 is the ethyl group in Example 1 of the present invention ( S )-2-[( R Experimental CD and calculated ECD curves of )-3-methyl-2,5-dioxopyrrolidin-1-yl] propanoate; Figure 6 is the ethyl group in Example 2 of the present invention ( S )-2-[( R )-3-methyl-2,5-dioxopyrrolidin-1-yl] propionate cytotoxicity test results; Figure 7 is the ethyl group in Example 3 of the present invention ( S )-2-[( R Figure 3 shows the inhibitory effect of [3-methyl-2,5-dioxopyrrolidin-1-yl] propionate on LPS-induced NO in BV2 cells. DETAILED DESCRIPTION

[0034] In order to make the objectives, technical solutions and advantages of the present invention more clear, the present invention will be further described in detail below with reference to the accompanying drawings and specific implementation methods.

[0035] Example 1 Ethyl ( S )-2-[( R )-3-methyl-2,5-dioxopyrrolidin-1-yl] propionate and its preparation The fungi derived from the mangrove sediment in this example Pseudallescheria angusta SYSU-M4 was deposited in the Guangdong Provincial Microbiological Culture Collection on May 7, 2025, with the deposit number GDMCC No: 66263. The address of the Guangdong Provincial Microbiological Culture Collection is: 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou.

[0036] The specific preparation method is: (1) Fungi derived from mangrove sediments Pseudallescheria angusta SYSU-M4 cryopreserved tubes were placed in a mold incubator at 28°C and 80% humidity for activation overnight; (2) The activated strain was inoculated into a pre-sterilized seawater potato liquid culture medium (200 g potato, 20 g glucose, 20 g sea salt, add pure water to 1 L, and5 The mixture was cultured in a shaking incubator (100 rpm) for 4 days (sterilized with Pa for 30 min) to obtain seed solution.

[0037] (3) The seed solution was inoculated into rice culture medium (50 g of rice / bottle, 50 mL of seawater with a sea salt content of 30 g / L / bottle, 121°C, 1×10 5 The culture was placed in a conical flask (1 L) sterilized with Pa for 30 min and cultured at room temperature for 30 days. The fermentation was completed when the mycelium completely filled the culture medium.

[0038] (4) The fermented bacterial cake was extracted with methanol three times, and the extracts were combined and concentrated under reduced pressure to obtain a first crude extract; (5) The first crude extract was redissolved in deionized water, extracted 5 to 6 times with ethyl acetate as the extractant, and the combined extracts were vacuum dried to obtain the second crude extract; (6) The second crude extract was subjected to column chromatography using normal phase silica gel chromatography; specifically, the second crude extract was fully mixed with blank silica gel (100-200 mesh) in a weight ratio of 1:2 to obtain sample silica gel. The column was wet packed by first adding 100-200 mesh blank silica gel, and then spreading the sample silica gel on the upper layer (the volume ratio of sample silica gel: blank silica gel was 1:7). A layer of cotton was spread on the top of the column to prevent the surface from being washed away by the solvent. Then, petroleum ether-ethyl acetate (volume ratio of 10:0, 8:2, 7:3, 6:4, 5:5, 4:6) and ethyl acetate-methanol (10:0, 1:1, 10:0) were used for gradient elution, and a total of 36 subcomponents (Fr.1-36) were obtained. After thin layer chromatography (TLC) analysis of the characteristics of each component, they were merged into five main components, namely Fr.A-Fr.E, according to the similarity of polarity. The relative transfer value (R f ) with a main component Fr.C of 0.65 as the first intermediate product.

[0039] (7) The first intermediate product was eluted using a gel column to obtain a second intermediate product; specifically, the gel column was Sephadex LH-20 and the eluent was dichloromethane-methanol with a volume ratio of 1:1. Five components (Fr.CA to Fr.CE) were obtained by gel column chromatography. After thin layer chromatography analysis of the characteristics of each component, the relative transfer value (R f ) is 0.7 component Fr.CB as the second intermediate product.

[0040] (8) Separating the second intermediate product by semi-preparative HPLC to obtain a compound; The mobile phase of the semi-preparative HPLC was acetonitrile and water in a volume ratio of 30:70, with a flow rate of 3 mL / min. The chromatographic column used was Aglient-XB-C18, with a column length of 250 mm and a diameter of 10 mm. The particle size of the stationary phase in the chromatographic column was 5 μ m; the residence time of compound (I) is 11 min.

[0041] The compound in Example 1 was subjected to structural analysis and testing, and the following physicochemical properties were obtained: brown powder; the HP-ESI-MS positive spectrum showed a molecular ion peak m / z of 213.7146 ([MH] - , theoretical value: 213.1000) (specific reference Figure 2 ). 1 H and 13 C NMR (see Table 1 for details) shows that the molecular formula is C 10 H 15 NO4, unsaturation Ω is 4.

[0042] According to (Table 1) 1 Analysis of the H NMR spectrum data shows that there are three methyl proton signals in the molecule δ H 1.56 (d, J =7.3Hz, H3-10), 1.35 (d, J = 6.9 Hz, H3-5) and 1.25 (t, J =7.1Hz, H3-9); 1 oxymethylene proton signal δ H 4.77 (q, J =7.3Hz, H2-8) and 2.89 (dd, J =6.9, 3.5Hz, H2-8); 1 heteroatom methine proton signal δ H 4.18 (m, 2H, H-6); 1 methylene proton signal not connected to a heteroatom δ H 2.95 (dd, J = 16.9, 8.6 Hz, H-6) and 2.36 (m, H-6); and a methine proton signal δ H 2.89 (dd, J =6.9, 3.5Hz, H2-2). 13 The C NMR spectrum shows 10 carbon signals, including 3 carbonyl carbon signals. δ C179.8 (C-1), 175.6 (C-4) and 169.3 (C-7), combined with the degree of unsaturation, it can be inferred that the compound contains one ring. 1 H- 1 H COSY spectrum ( Figure 3 ), H-2 is correlated with H-3 and H-5, H-6 is correlated with H-10, and H-8 is correlated with H-9, indicating the existence of structural fragments CH3-CH2- and CH3-CH-CH2-. Combined with the HMBC spectrum ( Figure 3 ), H-5 is related to C-2, H-3 is related to C-1, C-2, and C-4; H-10 is related to C-6 and C-7; H-6 is related to C-1, C-4, C-7, and C-10; and H-8 is related to C-7 and C-9. In summary, the structural characteristics of this compound are as follows: a methyl group is attached to positions 2 and 6, an ethoxy group is attached to position 7, and the molecule contains three carbonyl carbons and one oxymethylene carbon.

[0043] Furthermore, no cross peaks between the protons were detected in the NOESY spectrum, indicating that the spatial distance between the two may exceed 5Å under experimental conditions, which is outside the effective observation range of the NOE effect. The chemical shifts of the two possible isomers (2R*,6S*)-1 and (2R*,6R*)-1 were calculated using the sub-chemical calculation method (e.g. Figure 4 The results showed that the DP4+ probability of compound 1 was 100%. Figure 5 ) and determined its absolute configuration to be (2R*,6S*)-1.

[0044] Based on the above determination and calculation, the absolute configuration of the compound obtained in this example is determined to be as shown in formula (I), and its IUPAC standard name is ethyl ( S )-2-[( R )-3-methyl-2,5-dioxopyrrolidin-1-yl] propionate [ethyl ( S )-2-(( R )-3-methyl-2,5-dioxopyrrolidin-1-yl)propanoate].

[0045] Table 1 Compounds represented by formula (I) 13 C and 1 H NMR (400 MHz, CDCl3) data

[0046] Example 2 Ethyl ( S )-2-[( RCytotoxicity Experiment of 3-Methyl-2,5-dioxopyrrolidin-1-yl] Propionate 1. Experimental methods: BV2 cell lines were obtained from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China). Cells were cultured in 90 mL of DMEM (Gibco, Catalog No. 11995065) supplemented with 10 mL of fetal bovine serum (FBS) (Gibco) at 37°C in a 5% CO2 incubator. The potential cytotoxicity of the compound represented by Formula (I) was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay.

[0047] The complete culture medium for BV-2 cells was prepared by adding 10 mL of fetal bovine serum (FBS) and 1 mL of 100× penicillin-streptomycin solution to every 100 mL of high-glucose DMEM medium, mixing thoroughly, and then filtration with 0.22 μ Sterilize by filtration through a m filter membrane and store at 4°C until ready for use. This medium formula provides the nutrients required for cell growth while effectively preventing bacterial contamination.

[0048] Cell seeding: per well The cells were seeded into 96-well plates at a density of 10 cells per well to ensure that the cells were evenly distributed and grew adherently.

[0049] Drug treatment: After cell inoculation, add the final concentration of 1~20 μ g / mL of the test sample. Set up multiple replicate wells for each concentration to improve the reliability of the experimental results. For colorimetric detection: 24 hours after drug treatment, add MTT reagent to each well and incubate for an additional 4 hours. Then, remove the culture medium and dissolve the resulting formazan crystals in DMSO. Finally, measure the absorbance (OD value) of each well at a wavelength of 490 nm.

[0050] Result calculation: The cell viability percentage was calculated according to the following formula: Cell viability (%) = (OD value of experimental group / OD value of control group) × 100%. By comparing the OD values of the experimental group and the control group, the effects of samples at different concentrations on cell viability were evaluated to determine their potential cytotoxicity.

[0051] 2. Experimental results: The compound (I) obtained in Example 1 was added at 0, 1, 10 and 20 μ g / mL dose effect on BV2 cells. Cell viability of each dose group was significantly different from that of 0 μ g / mL were similar, with no significant difference ( Figure 6 ).

[0052] Example 3 Ethyl ( S )-2-[( REffects of [(3-methyl-2,5-dioxopyrrolidin-1-yl)-3-propionate] on NO release in BV-2 cells 1. Experimental methods: The BV2 cell line and culture medium information are as in Example 2.

[0053] Preparation of standard curve: Prepare the standard curve according to the instructions of the kit (Beyotime S0021M). The regression equation of the standard curve established is y=0.0036x+0.0542, and its correlation coefficient is It reached 0.999, indicating a good linear relationship.

[0054] Experimental grouping: The experimental design includes three main groups: normal control group, LPS stimulation group (concentration of 1 μ g / mL), and sample treatment groups (concentrations ranging from 1 to 20 μ g / mL). Five replicate wells were set in each group to ensure the reliability of the data.

[0055] Specific detection steps: First, collect the cell supernatant from each experimental group. Then, mix equal volumes of the cell supernatant with Griess reagent and react for 10 minutes in the dark. Finally, measure the absorbance at 540 nm using a microplate reader.

[0056] Statistical Analysis: One-way ANOVA was used to analyze the data. The results were compared with those in the LPS-stimulated group. Significance levels are indicated as *p<0.05, **p<0.01, and ***p<0.001, respectively, to indicate the statistical significance of the differences in NO content between the different treatment groups.

[0057] 2. Experimental results: Specific experimental results such as Figure 7 As shown in the figure, it can be seen that compound (I) μ After treatment with 20 μg / mL concentration, the concentration of NO was significantly lower than that of LPS group. This indicates that compound (I) μ g / mL concentration can significantly inhibit the production of NO.

[0058] The above is a preferred embodiment of the invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the invention. These improvements and modifications are also considered to be within the scope of protection of the present invention.

Claims

1. Compounds of formula (I) and pharmaceutically acceptable salts thereof; （Ⅰ）。 2. A pharmaceutical composition, characterized in that The invention comprises the compound as claimed in claim 1 and a pharmaceutically acceptable salt thereof.

3. Use of the compound according to claim 1 and its pharmaceutically acceptable salt as an active ingredient in the preparation of anti-inflammatory drugs.

4. Use of the compound according to claim 1 and its pharmaceutically acceptable salt as an active ingredient in the preparation of a drug for inhibiting intracellular NO production.

5. Use of the compound according to claim 1 and its pharmaceutically acceptable salt as an active ingredient in the preparation of a medicament for neuroinflammatory diseases.

6. Use of the compound according to claim 5 and its pharmaceutically acceptable salt as an active ingredient in the preparation of a drug for treating Parkinson's disease. The neuroinflammatory diseases include Alzheimer's disease and Parkinson's disease.

7. A method for preparing the compound according to claim 1, characterized in that: include: Mangrove sediment-derived fungi Pseudallescheria angusta SYSU-M4 is inoculated into the culture medium and obtained through fermentation, extraction, and separation; Wherein, the mangrove sediment mud originates from fungi Pseudallescheria angusta SYSU-M4 was deposited in the Guangdong Provincial Microbiological Culture Collection on May 7, 2025, with the deposit number GDMCC No: 66263. The address of the Guangdong Provincial Microbiological Culture Collection is: 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou.

8. The method for preparing the compound according to claim 7, wherein include: (1) Fungi derived from mangrove sediments Pseudallescheria angusta SYSU-M4 is inoculated into a culture medium for fermentation to obtain a fermentation product; (2) extracting the fermentation product with an extraction solvent, extracting with an extraction solvent, separating with a separation solvent, and separating with semi-preparative HPLC to obtain; Wherein, the extraction solvent is selected from methanol and / or dichloromethane; The extraction solvent is ethyl acetate; The separation solvent is selected from one or more of petroleum ether, ethyl acetate, dichloromethane, methanol, chloroform, and n-hexane; The mobile phase of the semi-preparative HPLC was acetonitrile and water, and the volume ratio of acetonitrile to water was (20-30): (70-80).

9. The method for preparing the compound according to claim 8, wherein The extraction solvent is methanol; The separation solvents are petroleum ether-ethyl acetate with a volume ratio of 10:0, 8:2, 7:3, 6:4, 5:5, and 4:6, ethyl acetate-methanol with a volume ratio of 10:0, 1:1, and 0:10, and dichloromethane-methanol with a volume ratio of 1:1; The mobile phase of the semi-preparative HPLC was acetonitrile and water in a volume ratio of 30:70, with a flow rate of 3 mL / min. The chromatographic column used was Aglient-XB-C18, with a column length of 250 mm and a diameter of 10 mm. The particle size of the stationary phase in the chromatographic column was 5 μ m; the residence time of the compound is 11 min.

10. The method for preparing the compound according to claim 8, wherein Step (2) includes: (2.1) extracting the fermentation product 1 to 3 times with an extraction solvent, combining the extracts and drying to obtain a first crude extract; wherein the extraction solvent is methanol; (2.2) dissolving the first crude extract in water, then extracting with ethyl acetate 5 to 6 times, combining the extracts and drying to obtain a second crude extract; (2.3) Eluting the second crude extract using a normal phase silica gel column to obtain a first intermediate product; wherein the silica gel packing used in the normal phase silica gel column is 100-200 mesh, and the eluents used are petroleum ether-ethyl acetate in volume ratios of 10:0, 8:2, 7:3, 6:4, 5:5, and 4:6, and ethyl acetate-methanol in volume ratios of 10:0, 1:1, and 0:10, respectively; the transferability of the first intermediate product is 0.65; (2.4) The first intermediate product is eluted using a gel column to obtain a second intermediate product; wherein the matrix of the gel column is agarose gel with a pore size of 25 to 100 μ m; the eluent used was dichloromethane-methanol with a volume ratio of 1:1; the transfer value of the second intermediate product was 0.7; (2.5) The second intermediate product was separated by semi-preparative HPLC to obtain a compound; wherein the mobile phase of the semi-preparative HPLC was acetonitrile and water in a volume ratio of 30:70, the flow rate was 3 mL / min, the chromatographic column used was Aglient-XB-C18, the column length was 250 mm, the diameter was 10 mm, and the particle size of the stationary phase in the column was 5 μ m; the residence time of the compound is 11 min.

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