Preparation method and application of benzofuran component in eupatorium chinense

Abstract

The invention discloses a preparation method and application of a benzofuran component in eupatorium chinense. The benzofuran component is separated from an eupatorium chinense extract. 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 separated compound on the diabetes-related target alpha-glucosidase is tested, the experimental result shows that the compound has good inhibition activity on the alpha-glucosidase, and the molecular docking experiment shows that the compound has good binding energy with the target protein alpha-glucosidase. Therefore, the compound can be used for preparing drugs for preventing, delaying or treating alpha-glucosidase diseases, especially type II diabetes and obesity, or can be used as a lead compound of the drugs.

Description

Technical Field

[0001] The present invention belongs to the field of biology and medicine technology, and relates to the separation, purification and application of benzofuran compounds from the roots of Huazelan, and specifically to a separation, purification and preparation method of new benzofuran compounds from the roots of Huazelan, and their inhibitory activity on α-glucosidase. Background Art

[0002] Eupatorium chinense is a plant of the genus Eupatorium in the Asteraceae family. It is named "Duoxugong" because of its beard-like roots. It is called Tuniuxi in Guangdong. Eupatorium chinense is widely distributed in many parts of my country and is a traditional ethnic medicinal plant in Hunan, Hubei and other places. The chemical components of Eupatorium chinense cover a variety of categories such as benzofurans, flavonoids, terpenes and phenols. Books such as "Fujian Materia Medica" record that Eupatorium chinense has the effects of "soothing the liver and relieving depression, opening the chest and relieving diaphragm". In the areas of Jiangsu and Zhejiang, people often use Eupatorium chinense roots to cut into slices and make tea, which can effectively control elevated blood sugar. Modern pharmacological research on Eupatorium chinense focuses on anti-inflammatory, antiviral, anti-tumor, and antibacterial effects, but there are fewer studies on its hypoglycemic and antidepressant activities.

[0003] α-glucosidase inhibitors can facilitate the passage of some undigested carbohydrates into the small intestine, stimulating the secretion of glucagon-like peptide 1 (GLP-1) there. This alteration in GLP-1 secretion contributes to further blood sugar control. Furthermore, α-glucosidase inhibitors can effectively ameliorate and prevent the development of some T2DM complications without significant hepatotoxicity or renal side effects. Therefore, α-glucosidase inhibitors have a promising therapeutic effect in Asian patients with T2DM who primarily consume carbohydrates, demonstrating unique advantages in managing T2DM and its complications. (Wang Meiyan. Design, synthesis, activity, and mechanism of action of dual-target inhibitors of α-glucosidase and PTP1B [D]. Tianjin Medical University, 2017.) Summary of the Invention

[0004] The first purpose of the present invention is to provide benzofuran compounds in the root of Huazelan and a preparation method thereof. The second purpose is to provide the use of the benzofuran compounds in the preparation of drugs for preventing or treating α-glucosidase-mediated diseases.

[0005] The compound having the ability to inhibit α-glucosidase activity has any one of the following structural formulas:

[0006] The technical solution of the present invention is to use the benzofuran compounds in the roots of Huazelan in the preparation of medicines for preventing, delaying or treating α-glucosidase-mediated diseases.

[0007] The preparation method of the Huazelan benzofuran compound comprises the following steps:

[0008] A1. Solvent extraction: The root of Herba Cynanchifoliae was crushed and extracted with ethanol, petroleum ether, ethyl acetate, and n-butanol in sequence. The n-butanol extract was concentrated to obtain extract a.

[0009] B1. Extract a impurity removal by macroporous adsorption resin: Extract a was dissolved in deionized water by ultrasonication. Macroporous adsorption resin (D101 type) was soaked in deionized water and loaded onto the column. The column was rinsed, wet-loaded, and allowed to stand overnight for adsorption. The column was eluted with ethanol / deionized water system, and the eluate was concentrated under reduced pressure to obtain extract b.

[0010] C1. Dissolve the extract b in methanol and perform a crude fractionation of the extract by reverse-phase silica gel column chromatography. Use wet column packing and dry loading, perform gradient elution using a methanol / deionized water system, and collect the eluate.

[0011] D1, taking the eluate and subjecting it to HW-40F gel column chromatography to obtain the separated product;

[0012] E1. The separated product was subjected to preliminary separation by reverse-phase silica gel column chromatography, and the obtained fragments were prepared by semi-preparative HPLC to obtain compounds 1-3.

[0013] In step C1, 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 were used for gradient elution, and the eluate from the 20% methanol / 80% water portion, the eluate from the 40% methanol / 60% water portion, or the eluate from the 60% methanol / 40% water portion was collected.

[0014] The eluate in step D1 was separated by gel column chromatography HW-40F with methanol-water at a volume fraction of 10-25:70-85 to obtain the separated product.

[0015] In step E1, the separated product is separated by reverse silica gel column chromatography using methanol-water with a volume fraction of 55-70:55-70.

[0016] In the semi-preparative HPLC, the mobile phase was acetonitrile-water 20:80, and the flow rate was 1-2 mL / min.

[0017] A medicine for treating diabetes and obesity, comprising the prepared compounds 1 to 3.

[0018] A compound for inhibiting alpha-glucosidase activity, comprising the prepared compounds 1 to 3.

[0019] A drug for preventing, delaying or treating diabetes and obesity is prepared by inhibiting the activity of α-glucosidase, and the drug comprises the prepared compounds 1 to 3.

[0020] The compounds 1 to 3 are combined with amino acids in the active site of the α-glucosidase protein to form sites containing hydrophobic interaction and hydrogen bond interaction.

[0021] The compound or drug of the present invention further includes a pharmaceutically acceptable excipient.

[0022] Compared with the prior art, this application has at least one of the following beneficial effects:

[0023] 1. Compounds 1 to 3 of the present application have significant α-glucosidase inhibitory activity, but there are no public reports on the α-glucosidase inhibitory activity of the compounds and their use in related diseases.

[0024] 2. A new compound 1, isolated from the n-butanol extract of the roots of Huazelan, was discovered for the first time to inhibit α-glucosidase activity. This compound has broad application prospects for developing drugs to prevent or treat α-glucosidase-mediated diseases, such as diabetes and obesity. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 The chemical structure diagram and two-dimensional correlation diagram of compound 1.

[0026] Figure 2 For compound 1 1 H-NMR.

[0027] Figure 3 For compound 1 13 C-NMR.

[0028] Figure 4 This is the DEPT135 spectrum of compound 1.

[0029] Figure 5 is the HSQC spectrum of compound 1.

[0030] Figure 6 For compound 1 1 H- 1 H COSY spectrum.

[0031] Figure 7 is the HMBC spectrum of compound 1.

[0032] Figure 8 is the NOESY spectrum of compound 1.

[0033] Figure 9 This is an analysis diagram of the interaction between compound 1 and α-glucosidase protein (wherein, A: 3D binding mode of compound 1 and α-glucosidase protein; B: amino acid residue binding mode of compound 1 and α-glucosidase protein; C: 2D binding mode of compound 1 and α-glucosidase protein). DETAILED DESCRIPTION

[0034] The following examples are used to further illustrate the present invention. Obviously, the examples described are only part of the embodiments of this application, not all of them. Based on the examples in this application, all other examples obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of this application.

[0035] Example 1: Isolation, preparation and structural analysis of benzofuran compounds 1 to 3 from the root of Huazelan (1) The separation and preparation method comprises the following steps:

[0036] Step A, Solvent Extraction: The roots of Huazelan were dried and crushed. Refluxing extraction was performed three times with 95% ethanol at room temperature. After concentration on a rotary evaporator, the extract was dispersed in water and extracted three times with petroleum ether, ethyl acetate, and n-butanol. The n-butanol extract was concentrated to obtain extract A.

[0037] Step B, extract a macroporous adsorption resin impurity removal: extract a was taken, 300 g of deionized water was added and ultrasonically dissolved, 1000 g of macroporous adsorption resin (D101 type) was soaked in deionized water for 6 h and then loaded onto the column, rinsed with deionized water for at least three column volumes, the solvent was lowered to about 2 cm above the filler surface, wet sample was loaded, allowed to stand overnight for adsorption, eluted with 50% ethanol / 50% deionized water, rinsed until the eluent was colorless, and the eluent was concentrated under reduced pressure to obtain extract b.

[0038] Step C, column chromatography crude fractionation: Take the above extract b, dissolve it with a small amount of methanol, add it to 100g of reverse-phase silica gel (200-300 mesh), and mix the sample; weigh 1200g of reverse-phase silica gel, soak it with 10% methanol / 90% deionized water, use wet column packing, the column volume is 1L, and rinse with 10% methanol / 90% deionized water for at least three column volumes until the solvent level in the column is approximately 3cm higher than the packing surface. Then, use dry method to load the sample, plug an appropriate amount of absorbent cotton into it as a protective layer after loading, and use methanol-water system elution to perform preliminary separation of extract b based on the results of high performance liquid chromatography analysis. 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, respectively, with each gradient washing 2 column volumes to obtain 8 fractions (named sequentially including Fr.AH).

[0039] Thin layer chromatography, high performance liquid chromatography and HPLC-DAD scanning analysis showed that the 20% methanol / 80% water part Fr.C was the component where the target compounds 1 and 3 were located, and the 60% methanol / 40% water part Fr.F was the component where the target compound 2 was located.

[0040] Step D: The 20% methanol / 80% water fraction Fr.C was subjected to isocratic elution on a gel column chromatography HW-40F (methanol / water = 2:8) to obtain fractions Fr.C (1-22). The 60% methanol / 40% water fraction Fr.F was subjected to reverse-phase silica gel column chromatography (10% MeOH / 90% dd H2O-100% MeOH) to obtain eight fractions Fr.F.1-8.

[0041] Step E: Fr.C.19 was subjected to preliminary separation by reverse silica gel column chromatography (methanol / water = 5:5) to obtain four fragments of compound (Fr.C19-15 to 18). After semi-preparative HPLC separation (acetonitrile-water 20:80, 2 mL / min, YMC C18 column), compound 1 (2.0 mg) was obtained from Fr.C-19-17. Compound 3 (2.1 mg) was obtained from Fr.C.16. Fr.F.8 was separated by gel column chromatography HW-40 to obtain four fragments (Fr.F.8.1-4). After further purification by semi-preparative HPLC, compound 2 (28.0 mg) was obtained from Fr.F.8.4. (2) Structural analysis of compounds 1 to 3:

[0042] Structural elucidation of compound 1

[0043] The NMR spectroscopy data and mass spectrometry data are as follows:

[0044] Compound 1: Yellow oily liquid. Optical rotation data: [α] 25 D -5.6 (c 0.12, methanol), UV (methanol) λmax = 220, 270, 305 nm; high resolution mass spectrometry HR-ESI-MS m / z: 267.1229 [M+H] + (C 14 H 19 O5, calcd.267.1227), and its molecular formula is determined to be C 14 H 18 O5, the degree of unsaturation is 6. According to compound 1 1 From the H-NMR spectrum, two olefin proton signals δ H 7.43 (1H, s, H-4), 7.61 (1H, d, H-6), 3 methyl signals δ H1.12 (1H, s, H-14), 1.17 (1H, s, H-15), 2.53 (1H, s, H-13). Combined with the spectral analysis, the 13 There are 14 carbon signals in the C-NMR spectrum and DEPT135 spectrum, including 3 methyl carbon signals δ C 27.0 (C-13), 26.2 (C-14), 25.7 (C-15); 3 oxygen-linked carbon signals δ C 71.8 (C-3), 70.1 (C-11) and 98.5 (C-2), 1 methoxy signal δ C 56.2 (C-12), 1 carbonyl carbon signal δ C 196.5 (C-10), 6 olefin carbon signals δ C 112.5 (C-4), 153.3 (C-5), 119.8 (C-6), 144.2 (C-7), 132.0 (C-8), 131.2 (C-9). The DEPT135 spectrum showed that there was no methylene carbon signal. Compound 1 is similar to the megapodiol compound reported in the literature (Bruce, B., Jarvis, Norman, B., et al. Non-trichothecenes from baccharismegapotamica [J]. Phytochemistry, 1986, 25 (2): 533-535.). 1 H-NMR spectrum and 13 The C-NMR spectral data were highly similar, indicating that compound 1 was a structural analogue of the compound reported in the literature.

[0045] By comparing the two 13 C-NMR spectrum data, the difference between compound 1 and this compound is that compound 1 has one more methoxy carbon signal δ C 56.2 (C-12), and while the methyl carbon signal remained unchanged, an additional oxygen-linked carbon signal appeared, and the DEPT135 spectrum showed no methylene signal. This suggests that the C-3 position of compound 1 has changed from the original methylene to a hydroxyl group, and the C-3 position has become a tertiary carbon atom linked to oxygen. The C-10 position has a methyl group attached to the side chain, and C-10 has changed from the original aldehyde carbon to a carbonyl carbon. In the HMBC spectrum, correlations can be observed between H-14 and C-2, between H-4 and C-3, C-5, C-9, and C-10, and between H-11 and C-6 and C-8. 1 H- 1 In the H COSY spectrum, there was a correlation between H-3 and H-2. The planar structure of compound 1 was determined by comprehensive analysis of the two-dimensional spectrum.

[0046] In the NOESY spectrum, we observed that H-2 and H-3 had related signals, indicating that H-2, H-3 and are on the same side. According to the literature summary (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, the hydrogen atoms at positions 2 and 3 are in trans configuration; if the coupling constants of H-2 and H-3 are relatively large, then the hydrogen atoms at positions 2 and 3 are in trans configuration. H-2-H-3 ≥4.0Hz, the hydrogen atoms at positions 2 and 3 are in a cis configuration. The coupling constants for H-2 and H-3 of this compound were observed to be relatively large, at J = 6.6Hz, indicating that the hydrogen atoms at positions 2 and 3 are in a cis configuration. Ultimately, the relative configuration of compound 1 was determined. Its absolute configuration was determined by comparing the TD-DFT theoretically calculated ECD spectrum with the experimental ECD spectrum (negative cotton effect at 218 and 276nm, positive cotton effect at 236 and 313nm), confirming that the configuration of C-2 and C-3 is 2R, 3S. After searching, it was found to be a new compound and named Eupbenzofuran H.

[0047] Table 1. Comparison of compound 1 with reference compounds 1 H-NMR and 13 C-NMR data comparison (solvent DMSO-d6)

[0048]

[0049] Structural analysis of compound 2:

[0050] The NMR spectroscopy data and mass spectrometry data are as follows:

[0051] Compound 2: C 15 H 18 O5,ESI-MS:m / z 279.1[M+H] + , 1 H-NMR (400 MHz, DMSO-d6) δ H :1.08(3H,t,J=7.3Hz,H-16),6.63(1H,s,H-3),7.28(1H,s,H-4),7.15(1H,s,H-7),3.54(1H,d,J=8.6Hz,H-11),3.43(1H,d,J=8.6Hz, H-11),3.30(1H,dd,J=8.6,7.3Hz,H-15),3.12(1H,dd,J=8.6,7.3Hz,H-15),2.63(3H,s,H-14),1.70(3H,s,H-12),10.27(1H,s,-OH); 13C NMR (100 MHz, DMSO-d6) δ C :203.9(C-13),161.3(C-6),158.7(C-8),158.3(C-2),128.0(C-4),121.1(C-9),116.9(C-5),104.4(C-3),96.6(C-7),77.1(C-10),65.3(C-11),57.0(C-15),26.0(C-14),18.9(C-12),17.0(C-16). 1 H-NMR spectrum and 13 The C-NMR spectrum was consistent with that in the literature (Díaz, CE, Fraga, B., Portero, AG, et al. Insect Antifeedant Benzofurans from Pericallis Species[J]. Molecules, 2023, 28(3): 975.), so the compound was identified as 10-ethoxy-11-hydroxy-10,11-dihydroeuparin, a yellow oil.

[0052] Structural analysis of compound 3:

[0053] The NMR spectroscopy data and mass spectrometry data are as follows:

[0054] Compound 3: C 13 H 14 O4,ESI-MS:m / z 235.1[M+H] + , 1 H-NMR (400 MHz, DMSO-d6) δ H :7.60(1H,s,H-4),7.39(1H,s,H-6),5.33(1H,d,J=3.6Hz,H-3),5.17(1H,s,H-2'-a), 4.82(1H,d,J=3.6Hz,H-2),4.77(1H,s,H-2'-b),2.36(1H,s,H-9),1.65(1H,s,H-3'); 13 C-NMR (100 MHz, DMSO-d6) δ C :198.6(C-8),152.4(C-7a),140.7(C-1'),140.6(C-5),131.3(C-7),129.1(C-4a),119.1(C-4),117.7(C-6),113.0(C-2'),95.1(C-2),76.3(C-3),26.3(C-9),17.2(C-3').1 H-NMR spectrum and 13 The C-NMR spectrum was consistent with that in the literature (Zhou, ZY, Liu, WX, Pei, G., et al. Phenolics from Ageratinaadenophora Roots and Their Phytotoxic Effects on Arabidopsis thaliana Seed Germination and Seedling Growth[J]. Journal of Agricultural & Food Chemistry, 2013, 61(48): 11792-11799.), so the compound was identified as (2S, 3R)-7-Hydroxytoxol, a colorless oil.

[0055] Example 2: Test of α-glucosidase inhibitory activity of compounds 1 to 3

[0056] (1) Experimental methods

[0057] α-glucosidase activity assay

[0058] The α-glucosidase assay was modified from the method of Tao et al. (Biomedical chromatography: BMC. 2013, 27(2): 148-155). The p-nitrophenyl-β-galactopyranoside method was used. pNPG was used as a substrate and decomposed into pNP and glucose under the catalysis of α-glucosidase. PNP was yellow and had strong UV absorption at 405 nm. Acarbose was used as a positive control. The amount of product after adding the sample was detected to determine the inhibition rate of the sample. 80 μL of sample (n-butanol extract of Huazelan or compounds 1-3) solution (pH 7.2 PBS buffer) and 20 μL of α-glucosidase (2 U / mL, pH 7.2 PBS buffer) were added to the test wells of a 96-well plate. After mixing, the mixture was incubated at 37°C for 15 min. 20 μL of 5 mmol / L pNPG was added to initiate the reaction. After incubation at 37°C for 15 minutes, the reaction was terminated by adding 80 μL of 1 mol / L Na2CO3. To the blank control group, 80 μL of 1% PBS was added instead of the sample solution. The absorbance (A) at 405 nm was used to quantify the amount of pNPG released. The following groups were set up: an enzyme activity group (20 μL enzyme + 80 μL reaction buffer + 20 μL substrate), an enzyme blank group (100 μL reaction buffer + 20 μL substrate), a sample group (80 μL sample + 20 μL enzyme + 20 μL substrate), and a sample blank group (80 μL sample + 20 μL reaction buffer + 20 μL substrate). Acarbose solution was used as a positive control. IC50 values ​​of the compounds were calculated using Origin software. The inhibition rate was calculated using the following formula: Inhibition rate = 1 - (A sample - A sample blank) / (A enzyme activity - A enzyme blank).

[0059] (2) Experimental results: Compounds 1 to 3 were tested for α-glucosidase inhibitory activity. The results are shown in Table 2:

[0060] Table 2. Results of the inhibition of α-glucosidase activity by compounds 1 to 3

[0061]

[0062] The test results show that the above-mentioned benzofuran compounds from the roots of Huazelan have good α-glucosidase inhibitory activity.

[0063] Example 3: Docking of compounds 1 to 3 with protein target molecules

[0064] (1) Experimental methods

[0065] Compounds 1 to 3 for this docking 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 optimization and exported as PDBQT files. The protein structure of α-glucosidase (PDB ID: 5ZCE) was obtained from the RCSB database ( https: / / www.rcsb.org / ), the protein structure was treated with Pymol and Autodock platforms to remove water molecules, remove ligands, and add hydrogen. The enzyme protein was then energy minimized, the geometry optimized, and exported as a PDBQT file. The Grid module in the Autodock software was used to perform computer simulation molecular docking calculations on the enzyme protein target and the compound after software optimization. The PDBQT file in Example 2 was imported into the software, and the box was allowed to fully wrap the protein, and then molecular docking was performed. Pymol 2.1 was used to visualize the enzyme protein and the molecular complex, and the interaction between the compound and the protein residue was analyzed. Finally, the binding mode was scored, and a reasonable inference was made on the active mechanism of the compound.

[0066] (2) Docking results

[0067] Molecular docking results

[0068] Table 3. Docking results of compounds 1 to 3 with target proteins

[0069]

[0070] Compound-protein interaction analysis

[0071] In this experiment, compounds 1 to 3 were molecularly docked with α-glucosidase protein. The molecular docking results showed that the compounds had a good binding effect with the target protein and a high degree of match, and the binding energy was less than -5kcal / mol. The complex formed by the docked compound and the protein was visualized using Pymol2.1 software to obtain the binding mode of the compound and the protein. According to the binding mode, the amino acid residues that bind to the compound and the protein pocket can be clearly seen. Compound 1 formed hydrophobic interactions with the PHE-144, PHE-282, and ASP-327 amino acids in the active site of the α-glucosidase protein, and formed multiple hydrogen bond interactions with ASP-60, GLN-256, ASP-327, GLN-328, and ARG-411 amino acids; Compound 2 formed hydrophobic interactions with α-glucosidase The PHE-163 amino acid in the active site of the protein formed a hydrophobic interaction, and formed a hydrogen bond interaction between the compound and the protein with the ASP-199, ALA-200, GLN-256, and ASN-258 amino acids; compound 3 formed a hydrophobic bond interaction with the ILE-143, PHE-225, and TYR-63 amino acid positions in the active site of the α-glucosidase protein, and formed a hydrogen bond interaction with the ALA-200, HIS-203, GLN-256, ASN-258, ASP-327, and ARG-411 amino acids.

[0072] The above detailed description of the analytical methods involved in the present invention provides a detailed introduction. It should be noted that the above description is intended solely to help those skilled in the art better understand the methods and concepts of the present invention, and is not intended to limit the relevant content. Without departing from the principles of the present invention, those skilled in the art may make appropriate adjustments or modifications to the present invention, and such adjustments and modifications shall also fall within the scope of protection of the present invention.

Claims

1. Benzofuran compounds from the root of Herba Cynanchifoliae, characterized in that: The structural formula is as follows:

2. The method for separating and extracting benzofuran compounds from the root of Herba Cynanchifoliae according to claim 1, characterized in that: The steps include: A1. The root of Herba Cynanchifoliae was crushed and extracted with ethanol, petroleum ether, ethyl acetate, and n-butanol in sequence, and the n-butanol extract was concentrated to obtain extract a; B1. Take extract a, add deionized water and ultrasonically dissolve it, soak the macroporous adsorption resin in deionized water and load it onto the column, rinse it, wet-load it, let it stand overnight for adsorption, elute it with ethanol / deionized water system, and concentrate the eluate under reduced pressure to obtain extract b; C1. Dissolve the extract b in methanol and perform crude fractionation using reverse-phase silica gel column chromatography. Wet-pack the column, dry-load the sample, and perform gradient elution using a methanol / deionized water system. Collect the eluate. D1. Separate the eluate using an HW-40F gel column chromatography to obtain a separated product. E1. The separated product was subjected to preliminary separation by reverse-phase silica gel column chromatography, and the obtained fragments were prepared by semi-preparative HPLC to obtain compounds 1-3.

3. The method for separating and extracting benzofuran compounds from the root of Herba Cynanchifoliae according to claim 2, wherein: In step C1, 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 were used for gradient elution, and the eluate from the 20% methanol / 80% water portion, the eluate from the 40% methanol / 60% water portion, or the eluate from the 60% methanol / 40% water portion was collected.

4. The method for separating and extracting benzofuran compounds from the root of Herba Cynanchifoliae according to claim 3, wherein: The eluate in step D1 was separated by gel column chromatography HW-40F with 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 root of Herba Cynanchifoliae according to claim 4, wherein: In step E1, the separated product is separated by reverse silica gel column chromatography using methanol-water at a volume fraction of 55-70:55-70; In the semi-preparative HPLC, the mobile phase was acetonitrile-water 20:80, and the flow rate was 2 mL / min.

6. The method for separating and extracting benzofuran compounds from the root of Herba Cynanchifoliae according to claim 5, characterized in that: Compound 1 and compound 3 were separated from the eluate of 20% methanol / 80% water; Compound 2 was isolated from the 60% methanol / 40% water eluate.

7. A drug for treating diabetes and obesity, characterized in that: Compounds 1-3 according to claim 1.

8. The drug for treating diabetes and obesity according to claim 7, characterized in that: The compounds 1-3 are used to prepare drugs for treating diabetes and obesity by inhibiting the activity of α-glucosidase.

9. The drug for treating diabetes and obesity according to claim 7, characterized in that: The compounds 1-3 combine with the amino acids in the active site of the α-glucosidase protein to form sites containing hydrophobic interactions and hydrogen bond interactions, thereby inhibiting the activity of the α-glucosidase.

10. The drug for treating diabetes and obesity according to any one of claims 7 to 9, characterized in that: The medicament includes a pharmaceutically acceptable excipient.

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