Application of benzofuran active ingredient in eupatorium chinense in reducing uric acid

Abstract

The invention discloses application of benzofuran components in eupatorium chinense to reduction of uric acid, compounds 1-3 are obtained by being separated from an eupatorium chinense extract, and the compound 1 is a new compound. The eupatorium chinense root extract is separated by using an extraction method, macroporous adsorption resin impurity removal, HW-40F gel column chromatography, high performance liquid chromatography and other methods to obtain the benzofuran compound. According to the present invention, the inhibition activity of the gout disease related target xanthine oxidase of the separated compound 1-3 is tested, the experimental result shows that the compound 1-3 has good inhibition activity on the xanthine oxidase, and the molecular docking experiment shows that the compound 1-3 has good binding energy with the target protein. Therefore, the compounds 1-3 can be used for preparing a natural lead compound for delaying or treating diseases caused by xanthine oxidase and providing a natural lead compound in the field of treatment of hyperuricemia and gout.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology and relates to the isolation, purification and application of benzofuran compounds from the roots of Zephyranthes bidentata. Specifically, it relates to the isolation, purification and preparation methods of a new benzofuran compound 1 from the roots of Zephyranthes bidentata, as well as the inhibitory activity of compounds 1-3 on xanthine oxidase. Background Technology

[0002] Xanthine oxidase is the rate-limiting enzyme in uric acid synthesis; therefore, xanthine oxidase inhibitors (XOIs) are the first-line drugs for intermittent gout attacks after acute attacks are controlled. Common xanthine oxidase inhibitors include allopurinol and febutatin. Allopurinol, as a first-line drug for gout treatment, is a xanthine oxidase inhibitor that reduces uric acid production and is used for gout and hyperuricemia. It has the advantages of high efficacy, low price, and good drug resistance. However, allopurinol often causes adverse reactions in patients, among which the most common adverse reaction is skin rash, with an incidence rate as high as 10%. Its incidence is not significantly related to the dosage, and it can range from mild to severe, with severe rashes being more common. SJS / TEN syndrome is particularly dangerous, with a mortality rate of up to 40%, and survivors often suffer permanent complications such as eye damage. HLA-B*5801 screening is recommended before medication (especially for Asians). Adverse reactions include bone marrow suppression (granulocytopenia, thrombocytopenia), hepatotoxicity (elevated transaminases), and hypersensitivity syndrome (fever, kidney damage, eosinophilia). Therefore, developing more effective and safer drugs for treating hyperuricemia is urgently needed in clinical practice. In recent years, scholars at home and abroad have turned their attention to extracts from traditional Chinese medicine, discovering many new natural drugs with uric acid-lowering effects, such as Compound Smilax Glabra Granules. Therefore, finding natural drugs from traditional Chinese medicine to treat hyperuricemia and gout or gout complications holds great promise.

[0003] Huazelan ( Eupatorium chinense*Eupatorium fortunei*, a plant belonging to the genus *Eupatorium* in the Asteraceae family, is named "Duoxugong" (meaning "many-rooted man") because of its fibrous roots. In Guangdong, it is called "Tu Niuxi" (meaning "earth ox knee"). *Eupatorium fortunei* is widely distributed in many parts of my country and is a traditional medicinal plant used by ethnic minorities in Hunan and Hubei provinces. Its chemical composition includes benzofurans, flavonoids, terpenes, and phenols. Books such as *Fujian Materia Medica* record that *Eupatorium fortunei* has the effects of "soothing the liver and relieving depression, opening the chest and benefiting the diaphragm." In the Jiangsu and Zhejiang regions, people often use slices of *Eupatorium fortunei* to make tea, which can effectively control blood sugar levels. Modern pharmacological research on *Eupatorium fortunei* focuses on its anti-inflammatory, antiviral, antitumor, and antibacterial effects, but research on its uric acid-lowering and anti-gout activities is relatively limited. The inventor has long been committed to the research of the chemical composition and pharmacological activities of *Eupatorium fortunei*. Previously, through isolation and purification, compounds 2 and 3 described in this invention were obtained (Chinese Invention Patent Application No.: 202510598431.0), and the antidiabetic activities of compounds 2 and 3 were reported. However, a literature review revealed no prior reports on the uric acid-lowering activity of compounds 2 and 3. Furthermore, the inventors previously reported that extracts of *Eupatorium fortunei* and its various extract fractions exhibited good uric acid-lowering activity (Chinese Invention Patent Application No.: 202510893853.0). This invention isolated three compounds (1-3) from the extract of *Eupatorium fortunei*, with compound 1 being a novel compound. In vitro xanthine oxidase inhibitory activity experiments showed that compounds 1-3 all exhibited good inhibitory activity, superior to the positive control drug allopurinol. Molecular docking experiments also demonstrated that compounds 1-3 have good binding energies to xanthine oxidase. The excellent xanthine oxidase inhibitory activity of these three compounds provides a safe and effective natural lead molecule for the treatment of hyperuricemia and gout or gout complications. Summary of the Invention

[0004] The first objective of this invention is to provide a novel benzofuran compound 1 from *Eupatorium fortunei* extract and a method for its preparation. The second objective is to provide the use of compounds 1-3 in the preparation of medicaments for the prevention or treatment of xanthine oxidase-mediated diseases.

[0005] The compound having the function of inhibiting xanthine oxidase activity has any one of the following structural formulas: .

[0006] The technical solution of this invention is the application of the benzofuran compounds in *Eupatorium fortunei* in drugs for delaying or treating xanthine oxidase-mediated diseases.

[0007] The preparation method of the aforementioned benzofuran compound 1 includes the following steps: A1. Solvent extraction: The roots of Zephyranthes cuspidatum were crushed and then successively extracted with ethanol, petroleum ether, ethyl acetate and n-butanol. The n-butanol extract was then concentrated to obtain extract a. B1. Removal of impurities from macroporous adsorption resin for extract a: Dissolve extract a in deionized water by ultrasonication. Soak macroporous adsorption resin (D101 type) in deionized water and load it onto the column. Rinse, load the sample by wet method, let it stand overnight for adsorption, and elute with ethanol / deionized water system. Concentrate the eluent under reduced pressure to obtain extract b. C1. Dissolve the above extract b in methanol, perform crude separation of the extract using reversed-phase silica gel column chromatography, use wet packing and dry loading, use methanol / deionized water system for gradient elution, and collect the eluent. D1. Take the eluent and separate it by HW-40F gel column chromatography to obtain the separated product; E1. The separated products were initially separated by reverse silica gel column chromatography, and the obtained fragments were prepared by semi-preparative HPLC to obtain compounds 1-3.

[0008] In step C1, gradient elution is performed using 10% methanol / 90% water, 15% methanol / 85% water, 20% methanol / 80% water, 30% methanol / 70% water, 40% methanol / 60% water, 60% methanol / 40% water, 80% methanol / 20% water, and 100% methanol. The eluent from the 20% methanol / 80% water fraction, or the 40% methanol / 60% water fraction, or the 60% methanol / 40% water fraction is collected.

[0009] The eluent from step D1 was separated by gel column chromatography (HW-40F) using methanol-water at a volume fraction of 10-25: 70-85 to obtain the separated product.

[0010] In step E1, the separated products are separated by reverse silica gel column chromatography with methanol-water at a volume fraction of 55-70: 55-70. In the semi-preparative HPLC, the mobile phase was acetonitrile-water 22:78, the flow rate was 2 mL / min, and compound 1 was prepared.

[0011] A medicine for treating gout or gouty arthritis or gout complications, or hyperuricemia, said medicine comprising compounds 1-3 prepared therefrom.

[0012] A compound that inhibits xanthine oxidase activity, said compound comprising compounds 1-3 prepared above.

[0013] A medicament for treating hyperuricemia and gout or gouty arthritis or gout complications by inhibiting the activity of xanthine oxidase, the medicament comprising compounds 1-3 prepared therefrom.

[0014] Compounds 1-3 form sites containing hydrophobic interactions and hydrogen bond interactions by binding to amino acids at the active site of xanthine oxidase protein; The compounds 1-3 or the drug of the present invention also include pharmaceutically acceptable excipients.

[0015] Compared with the prior art, this application has at least one of the following beneficial effects: 1. Compounds 1-3 of this application have significant xanthine oxidase inhibitory activity, which is higher than that of the positive control drug allopurinol. There are no publicly available reports on the xanthine oxidase inhibitory activity of the compounds and their application in related diseases.

[0016] 2. A novel compound with xanthine oxidase inhibitory activity was first discovered and isolated from the n-butanol extract of *Eupatorium fortunei* roots. This compound has broad application prospects for development into a drug for the prevention or treatment of xanthine oxidase-mediated diseases such as gout, gouty arthritis, gout complications, and hyperuricemia. Attached Figure Description

[0017] Figure 1 The chemical structure diagram and two-dimensional key correlation diagram of compound 1 are shown.

[0018] Figure 2 ECD calculation spectrum of compound 1 Figure 3 For compound 1 1 H-NMR spectrum.

[0019] Figure 4 For compound 1 13 C-NMR spectrum.

[0020] Figure 5 The image shows the DEPT135 spectrum of compound 1.

[0021] Figure 6 The image shows the HSQC spectrum of compound 1.

[0022] Figure 7 For compound 1 1 H- 1 H COSY spectrum.

[0023] Figure 8 The image shows the HMBC spectrum of compound 1.

[0024] Figure 9 The NOESY spectrum of compound 1 is shown.

[0025] Figure 10 The diagram shows the interaction analysis between compound 1 and xanthine oxidase protein (where A: 3D binding mode of compound 1 with xanthine oxidase protein; B: amino acid residue binding mode of compound 1 with xanthine oxidase protein; C: 2D binding mode of compound 1 with xanthine oxidase protein).

[0026] Figure 11 The diagram shows the interaction between compound 2 and xanthine oxidase protein (where A: 3D binding mode of compound 2 with xanthine oxidase protein; B: amino acid residue binding mode of compound 2 with xanthine oxidase protein; C: 2D binding mode of compound 2 with xanthine oxidase protein).

[0027] Figure 12 The diagram shows the interaction between compound 3 and xanthine oxidase protein (where A: 3D binding mode of compound 3 with xanthine oxidase protein; B: amino acid residue binding mode of compound 3 with xanthine oxidase protein; C: 2D binding mode of compound 3 with xanthine oxidase protein). Detailed Implementation

[0028] The following embodiments are used to further explain and illustrate the present invention. Obviously, the described embodiments are only some embodiments of this application, and not all embodiments. Based on the embodiments in this application, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this application.

[0029] Example 1: Isolation, preparation, and structural analysis of a novel benzofuran compound 1 from the roots of Zephyranthes cusia. (1) The separation and preparation method includes the following steps: Step A, Solvent Extraction: The roots of *Eupatorium fortunei* were dried and pulverized. They were then extracted three times under reflux with 95% ethanol at room temperature. The extract was concentrated using a rotary evaporator and dispersed in water. Extraction was then performed sequentially with petroleum ether, ethyl acetate, and n-butanol, each repeated three times. The n-butanol extract was concentrated to obtain extract a.

[0030] Step B: Removing impurities from macroporous adsorption resin of extract a: Take extract a, add 300g of deionized water and dissolve by ultrasonication. Soak 1000g of macroporous adsorption resin (D101 type) in deionized water for 6 hours and then load it onto the column. Rinse with deionized water for at least three column volumes. Lower the solvent to about 2 cm above the packing surface, load the sample by wet method, let it stand overnight for adsorption, and elute with 50% ethanol / 50% deionized water until the eluent is colorless. Concentrate the eluent under reduced pressure to obtain extract b.

[0031] Step C, Column Chromatography Crude Separation: Take the above-mentioned extract b, dissolve it in a small amount of methanol, add it to 100g of reversed-phase silica gel (200~300 mesh), and mix the sample; weigh 1200g of reversed-phase silica gel, soak it in 10% methanol / 90% deionized water, pack the column using the wet method, the column volume is 1 L, wash with 10% methanol / 90% deionized water for at least three column volumes, until the solvent level in the column is about 3cm higher than the packing surface, then load the sample using the dry method, after loading the sample, insert an appropriate amount of degreased cotton as a protective layer, and combine the high performance liquid chromatography analysis results, use the methanol-water system to elute, and perform preliminary separation of extract b. Gradient elution was performed using 10% methanol / 90% water, 15% methanol / 85% water, 20% methanol / 80% water, 30% methanol / 70% water, 40% methanol / 60% water, 60% methanol / 40% water, 80% methanol / 20% water, and 100% methanol, with each gradient elution lasting two column volumes, yielding eight fractions (Fr.AH).

[0032] Thin-layer chromatography, high-performance liquid chromatography, and HPLC-DAD scanning analysis showed that the 20% methanol / 80% water fraction Fr.C was the component containing target compound 1.

[0033] Step D: Take the 20% methanol / 80% water fraction Fr.C and elute it with an isogradient gel column chromatography HW-40F (methanol / water = 2:8) to obtain fractions Fr.C (1~22).

[0034] Step E: After preliminary separation by reverse silica gel column chromatography (methanol / water = 5:5), Fr.C.20 was obtained as four fragments (Fr. C20-1~4). Step F: The second fragment Fr. C20-2 from step E was separated by semi-preparative HPLC (acetonitrile-water 22:78, 2 mL / min, YMC C18 column) to obtain compound 1 (4.6 mg).

[0035] (2) Structural analysis of new compound 1: Yellow, oily liquid. Optical rotation data [α] 25 D -6.9°C (c 0.15, methanol), UV (methanol) λ max = 239, 350 nm; measured by high-resolution mass spectrometry HR-ESI-MS m / z : 253.1068 [M+H] + (C) 13 H 17 O5, calcd. 253.1071), its molecular formula is determined to be C 13 H 16 O5, with an unsaturation degree of 6. Analysis of compound 1... 1The H-NMR spectrum (Table 1) showed three olefin proton signals. δ H 7.94 (H-4), 7.87 (H-6), 6.90 (H-7), 2 methyl signals δ H 0.97 (H-14), 2.52 (H-11). Compound 1 13 The C-NMR and DEPT135 spectra (Table 1) show 14 carbon signals, including 2 methyl carbon signals. δ C 27.0 (C-11), 20.1 (C-14); 3 oxygen-carbon signals δ C 72.8 (C-12), 70.8 (C-3), and 66.6 (C-13); DEPT135 spectra revealed that among the three oxygen-bonded carbon signals, one was a methylene carbon signal with a single oxygen atom. δ C 66.6 (C-13); 1 carbonyl carbon signal δ C 196.5 (C-10); 6 olefin carbon signals δ C 164.4 (C-8), 131.67 (C-6), 109.8 (C-7), 131.7 (C-9), 126.7 (C-4), 130.6 (C-5). Compound 1 is similar to the compound megapodiol reported in the literature [Bruce, B., Jarvis, Norman, B., et al. Non-trichothecenes from baccharis megapotamica. Phytochemistry, 1986, 25(2): 533-535.]. 1 H-NMR spectrum and 13 The C-NMR spectral data are highly similar, indicating that this compound, like those reported in the literature, belongs to the benzofuran analogue family. A comparison of the two... 13 According to the C-NMR spectral data, the difference between this compound and the literature compound is the absence of one methylene signal. δ C The presence of an additional tertiary carbon signal at position 29.6 (C-3) indicates the presence of a hydroxyl group at the C-3 position of the compound. This can be observed in the HMBC spectrum. δ H 7.94 (H-4) and δ C 70.8 (C-3) is relevant. δ C72.8 (C-12) and δ H A correlation was found at 5.33 (H-3), confirming the presence of a hydroxyl group at the C-3 position. In the HMBC spectrum, correlations were observed between H-11 and C-10, C-5; between H-3 and C-8, C-9; and between H-13 and C-3. 1 H- 1 In the HCOSY spectrum, there is a correlation between H-2 and H-3. Comprehensive analysis of the two-dimensional spectrum of this compound confirms its planar structure. Figure 1 ).

[0036] In the NOESY spectrum, we observed correlated signals between H-13 and H-2, H-3, indicating that H-2, H-3, and H-13 are on the same side, based on literature summaries (…). Tetrahedron Lett 1972, 13, 2873-2876; J. Org. Chem . 1971, 36, 1805-1807.), if the coupling constants of H-2 and H-3 are relatively small ( J H-2-H-3 If the Hz value is ≤2.0 Hz, then the hydrogen atoms at positions 2 and 3 are in the trans configuration; if the coupling constants of H-2 and H-3 are relatively large... J H-2-H-3 ≥4.0 Hz indicates that the hydrogen atoms at positions 2 and 3 are in a cis configuration. The coupling constants of H-2 and H-3 in this compound are observed to be relatively large. J H-2-H-3 =4.3 Hz, indicating that the hydrogen atoms at positions 2 and 3 are in cis configuration. Finally, the relative configuration of compound 1 was determined. Its absolute configuration was determined by comparing the ECD spectrum calculated using TD-DFT theory with the experimental ECD spectrum (which showed a negative cotton effect at 220 nm and positive cotton effects at 280 and 300 nm). Figure 2 The configurations of C-2, C-3, and C-12 are all determined to be 2. S 3 R 12 R A search revealed that it is a new compound.

[0037] Table 1. Compound 1 and its literature counterpart 1 H-NMR, 13 C-NMR data comparison

[0038] Example 2: Test of the inhibitory activity of compounds 1-3 on xanthine oxidase (1) Experimental materials Xanthine, xanthine oxidase, allopurinol, potassium dihydrogen phosphate, disodium hydrogen phosphate, etc. (2) Experimental methods Xanthine oxidase can catalyze the production of uric acid and superoxide anions from xanthine and hypoxanthine. Uric acid has a characteristic absorption peak at 290 nm. Therefore, in vitro enzymatic reactions were used to evaluate the in vitro activity of Vanisanghuang extract and its various extract fractions.

[0039] Preparation of phosphate buffer (pH 7.4): Dissolve 0.2 g potassium dihydrogen phosphate, 2.89 g disodium hydrogen phosphate, 8.0 g sodium chloride, and 0.2 g potassium chloride in 1000 mL of ultrapure water. Substrate solution: Dissolve 15.2 mg xanthine in an appropriate amount of phosphate buffer, add 1 mL of 1 mol / L sodium hydroxide to aid dissolution, add 1 mol / L hydrochloric acid to adjust the pH to 7.4, sonicate, and add phosphate buffer to bring the volume to 100 mL to obtain a substrate solution with a concentration of 100 μm.

[0040] Enzyme solution: Dissolve 1000 U xanthine oxidase in 2 mL phosphate buffer to obtain a 500 U / mL enzyme stock solution. Aliquot the solution under an ice-water bath and store at -80℃. Dilute to a working concentration of 100 U / L before use and store under an ice-water bath. Sample solution: Dissolve each part of the sample in DMSO, store at 4℃, and dilute to the working concentration of 500 μg / mL before use.

[0041] Allopurinol solution: Dissolve 200 mg of allopurinol in an appropriate amount of phosphate buffer, bring the phosphate buffer to a final volume of 100 mL, store at 4°C, and dilute to different concentrations before use.

[0042] The specific experimental steps are as follows: Using a 200 μL final volume reaction system in a 96-well plate, prepare the sample, enzyme, and substrate solutions as described above and dilute to the working concentration. Prepare the enzyme solution fresh for each use. During the experiment, keep the enzyme solution in an ice-water bath to prevent enzyme inactivation. Add 50 μL of enzyme solution, sample solution, allopurinol solution, and the corresponding phosphate buffer according to the groups in Table 2. Incubate at 37°C for 10 min. Finally, add 100 μL of 100 μM substrate to start the reaction and incubate at 37°C for 30 min. Measure the OD value at 290 nm using a microplate reader and calculate the inhibition rate.

[0043]

[0044] Table 2. Experimental methods for XOD inhibition in vitro

[0045] (3) Experimental results Table 3. Results of the inhibitory activity of compounds 1-3 on xanthine oxidase (n=3, ±SEM)

[0046] As can be seen from Table 3, compounds 1-3 all exhibited good inhibitory activity against xanthine oxidase, and their inhibitory activity was superior to that of the positive control drug allopurinol.

[0047] Example 3: Docking of compounds 1-3 with protein target molecules (1) Experimental methods Compounds 1-3 used in this docking experiment were constructed using ChemDraw, then imported into Chem3D software for optimization and energy minimization using the MM2 module, and saved as SDF files as ligand molecules for molecular docking. They were then imported into Pymol and Autodock software for further optimization and exported as PDBQT files. The xanthine oxidase (PDB ID: 1FIQ) protein structure was obtained from the RCSB database (…). https: / / www.rcsb.org / The protein structure was processed using Pymol and Autodock platforms, including water molecule removal, ligand removal, and hydrogenation. Energy minimization and geometric optimization were performed on the protein, and the result was exported as a PDBQT file. Molecular docking was handled and optimized using the Grid module in Autodock software. The PDBQT file was imported into the software, the bounding box fully encapsulated the protein, and then molecular docking was performed. Additionally, the protein-small molecule complexes were visualized and analyzed using Pymol.

[0048] (2) Docking results The molecular docking results are shown in the table below: Table 4. Docking results of compounds 1-3 with target protein 1FIQ

[0049] Compound-protein interaction analysis: In this experiment, compounds 1-3 and the positive control drug allopurinol were molecularly docked with the xanthine oxidase target protein. The docking results showed that the compounds and target proteins exhibited excellent binding interactions and high matching degrees (Table 4), with binding energies all less than -5 kcal / mol. The complexes formed by the docked compounds and proteins were visualized using Pymol 2.1 software to obtain the binding patterns. Based on the binding patterns, it was clearly observed that compound 1 formed a hydrophobic interaction with amino acid THR-1077 at the active site of the xanthine oxidase protein, and formed multiple hydrogen bonds with amino acids ALA-1079, SER-1080, and THR-1083. Figure 10Compound 2 forms hydrophobic interactions with amino acids ARG-912, GLN-1040, ALA-1078, and GLU-1261 at the active site of xanthine oxidase protein, and forms multiple hydrogen bond interactions with amino acids GLN-767, GLN-1040, and GLN-1194. Figure 11 Compound 3 forms hydrophobic interactions with the LEU-257, VAL-269, ALA-301, ILE-353, and LEU-398 amino acids at the active site of xanthine oxidase protein, and forms multiple hydrogen bond interactions with the SER-347, THR-354, and LEU404 amino acids. Figure 12 ).

[0050] The above detailed embodiments provide a specific description of the analytical methods involved in this invention. It should be noted that the above description is only intended to help those skilled in the art better understand the methods and ideas of this invention, and is not intended to limit the scope of the invention. Without departing from the principles of this invention, those skilled in the art can make appropriate adjustments or modifications to this invention, and such adjustments and modifications should also fall within the protection scope of this invention.

Claims

1. Benzofuran compounds from Zephyranthes var. huazellariae, characterized in that, The structural formula is as follows: 。 2. The method for separating and extracting benzofuran compound 1 from Zephyranthes bidentata according to claim 1, characterized in that, Includes the following steps: A1. Solvent extraction: The roots of Zephyranthes cuspidatum were crushed and then successively extracted with ethanol, petroleum ether, ethyl acetate and n-butanol. The n-butanol extract was then concentrated to obtain extract a. B1. Removal of impurities from macroporous adsorption resin of extract a: Take extract a and add it to deionized water for ultrasonic dissolution. Take macroporous adsorption resin, soak it in deionized water and load it onto the column, rinse, wet load the sample, let it stand overnight for adsorption, elute with ethanol / deionized water system, and concentrate the eluent under reduced pressure to obtain extract b. C1. Dissolve the above extract b in methanol, perform crude separation of the extract using reversed-phase silica gel column chromatography, use wet packing and dry loading, use methanol / deionized water system for gradient elution, and collect the eluent. D1. Take the eluent and separate it by HW-40F gel column chromatography to obtain the separated product; E1. The separated product was initially separated by reverse silica gel column chromatography, and the obtained fragment was prepared by semi-preparative HPLC to obtain compound 1.

3. The method for separating and extracting benzofuran compounds from the roots of *Eupatorium fortunei* according to claim 2, characterized in that, In step C1, gradient elution is performed using 10% methanol / 90% water, 15% methanol / 85% water, 20% methanol / 80% water, 30% methanol / 70% water, 40% methanol / 60% water, 60% methanol / 40% water, 80% methanol / 20% water, and 100% methanol. The eluent from the 20% methanol / 80% water fraction, or the 40% methanol / 60% water fraction, or the 60% methanol / 40% water fraction is collected.

4. The method for separating and extracting benzofuran compounds from the roots of *Eupatorium fortunei* according to claim 2, characterized in that, The eluent from step D1 was separated by gel column chromatography (HW-40F) using methanol-water at a volume fraction of 10-25: 70-85 to obtain the separated product.

5. The method for separating and extracting benzofuran compounds from the roots of *Eupatorium fortunei* according to claim 2, characterized in that, In step E1, the separated products are separated by reverse silica gel column chromatography with methanol-water at a volume fraction of 55-70: 55-70. In the semi-preparative HPLC, the mobile phase was acetonitrile-water 22:78, the flow rate was 2 mL / min, and compound 1 was prepared.

6. The method for separating and extracting benzofuran compound 1 from Zephyranthes bidentata according to claim 5, characterized in that, Compound 1 was isolated from the eluent of the 20% methanol / 80% water fraction.

7. A compound or composition that inhibits xanthine oxidase activity, characterized in that, The compounds include compounds 1 to 3 as described in claim 1 or a combination thereof.

8. A drug for treating hyperuricemia, gout, gouty arthritis, or gout complications by inhibiting the activity of xanthine oxidase, characterized in that, The drug comprises compounds 1-3 or a combination thereof as described in claim 1.

9. The compound or drug according to claim 7 or 8, characterized in that, Compounds 1-3 of claims 1 form sites containing hydrophobic interactions and hydrogen bond interactions by binding to amino acids at the active site of xanthine oxidase protein.

10. The compound or drug for treating hyperuricemia, gout, gouty arthritis, or gout complications according to claim 7 or 8, characterized in that, The compound or drug also includes pharmaceutically acceptable excipients.

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