A compound extracted from equisetum arvense and preparation method and application thereof

Abstract

The present application relates to a kind of compounds extracted from Eupatorium fortunei and its preparation method and application, belong to medical technology field.The present application provides the method for preparing and identifying 14 new compounds for the first time with Eupatorium fortunei whole plant as raw material, and systematically evaluates its lung protection activity, and describes its application in the development of lung inflammation related chemical prevention, treatment drug.The present application carries out lung inflammation activity test to in vitro MLE-12 cell as model, and the inhibitory activity of compound NNK induced MLE-12 cell prepared is evaluated.The results show that these new compounds have significant lung protection activity, and can be used for developing lung disease chemical prophylactic or therapeutic drug.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to a class of compounds extracted from wild horse, their preparation methods, and applications. Background Technology

[0002] The Chinese medicinal herb Yemazhui is *Eupatorium fortunei*, a plant in the Asteraceae family. Eupatorium lindleyanum The dried aerial parts of *Dendrobium nobile* (DC.) are mainly distributed in Jiangsu, Shandong, Gansu, and Hunan provinces of my country, with Xuyi County, Jiangsu Province being the authentic producing area. *Dendrobium nobile* has a bitter taste and neutral properties, and enters the lung meridian. Its main functions are to resolve phlegm, relieve cough, and alleviate asthma, and it is often used to treat various respiratory diseases. Modern pharmacological studies have shown that *Dendrobium nobile* exhibits various pharmacological activities in treating respiratory diseases, anti-inflammation, and anti-tumor effects. In recent years, Jiangsu Province has organized a research team to conduct systematic research on *Dendrobium nobile* and has developed clinical preparations such as *Dendrobium nobile* syrup, showing good therapeutic effects. *Dendrobium nobile* has also been recognized as an authentic medicinal material of Jiangsu Province, and relevant departments have vigorously promoted its artificial cultivation. It has now become one of the key varieties in the Jiangsu Province's standardized medicinal material cultivation base construction project and the pillar industry project of the northern Jiangsu local medicinal material region.

[0003] Wild horse gall (Pteris vittata) possesses expectorant, antitussive, and antiasthmatic effects, and has historically been used to treat various respiratory diseases such as chronic bronchitis, asthma, and cough, with widespread application in both folk medicine and clinical practice. With the continuous advancement of modern pharmacological research, researchers have discovered that Wild horse gall not only exerts therapeutic effects on respiratory diseases but also exhibits various pharmacological activities, including antitumor, antioxidant, antiviral, anti-inflammatory, and immunomodulatory activities, demonstrating high medicinal development value. Among these, sesquiterpene lactones, as characteristic active components of Wild horse gall, have attracted widespread attention due to their unique configuration and significant biological activity. Studies have shown that its sesquiterpene components can regulate NF-κB... k B, MAPK and other signaling pathways play an anti-inflammatory and anti-lung injury role, becoming an important material basis for supporting the efficacy of drugs in wild horses.

[0004] Currently, research on wild horse antler extract mainly focuses on the pharmacological activities of crude extracts or known components. However, existing technologies are still insufficient for in-depth exploration of the active components in wild horse antler extract, especially the discovery and systematic study of novel structural compounds. There is an urgent need to develop more efficient extraction methods and new compound discovery strategies to expand its medicinal value. Summary of the Invention

[0005] The purpose of this invention is to provide fourteen new compounds extracted from wild horse, their preparation methods, and applications.

[0006] A class of compounds or their pharmaceutically acceptable salts having the following chemical structural formula, .

[0007] Another object of the present invention is to provide a method for preparing the above-mentioned compound, comprising the following steps: (1) Wild horse chasing ( Eupatorium lindleyanum DC. is extracted by heating and reflux with pure water, ethanol solution or methanol solution, and the extract is recovered to obtain crude extract; (2) The crude extract obtained in step (1) is separated by D101 macroporous adsorption resin and eluted with a water-ethanol mixed solvent in a gradient. The fractions with a water-ethanol volume ratio of 100:0 to 20:80 are collected to obtain the target fraction A. (3) The target fraction A obtained in step (2) above is separated by ODS column chromatography, and gradient elution is performed using methanol-water or acetonitrile-water mixed solvent as mobile phase. The fractions with a volume ratio of methanol-water or acetonitrile-water of 1:9 to 8:2 are collected to obtain the target fraction B. (4) The target fraction B obtained in step (3) above is further separated and prepared by preparative high performance liquid chromatography. Gradient elution is performed using methanol-water mixed solvent or acetonitrile-water mixed solvent as the mobile phase to obtain compounds 1-14.

[0008] In the above technical solution, in step (1), the extraction method is to heat and reflux extract 2 to 5 times; the material-liquid ratio is 1:8 to 1:20 g / mL; the volume concentration of the ethanol solution is 70% to 95%; and the volume concentration of the methanol solution is 80% to 95%.

[0009] Preferably, the volume concentration of the ethanol solution is 80%~95%; the volume concentration of the methanol solution is 80%~90%; and the material-to-liquid ratio is 1:10~1:15 g / mL.

[0010] In the above technical solution, in step (2), the volume ratio of water to ethanol in the water-ethanol mixed solvent is 100:0~5:95.

[0011] Preferably, in step (2), the volume ratio of water to ethanol in the water-ethanol mixed solvent is 70:30 to 30:70.

[0012] In the above technical solution, in step (2), the crude extract is dissolved in an organic solvent, mixed with macroporous adsorption resin and dried, and then separated by D101 macroporous adsorption resin.

[0013] Preferably, the organic solvent is methanol.

[0014] In the above technical solution, in step (3), the volume ratio of methanol to water in the methanol-water mixed solvent is 1:9~9:1; the volume ratio of acetonitrile to water in the acetonitrile-water mixed solvent is 1:9~8:2.

[0015] Preferably, in step (3), the volume ratio of methanol to water in the methanol-water mixed solvent is 3:5 to 1:1; and the volume ratio of acetonitrile to water in the acetonitrile-water mixed solvent is 1:9 to 2:5.

[0016] In the above technical solution, in step (4), the volume ratio of methanol to water in the methanol-water mixed solvent is 2:8~6:4; and the volume ratio of acetonitrile to water in the acetonitrile-water mixed solvent is 1:9~6:4.

[0017] Preferably, in step (4), the volume ratio of methanol to water in the methanol-water mixed solvent is 3:7 to 5:5, and the volume ratio of acetonitrile to water in the acetonitrile-water mixed solvent is 1:9 to 3:7.

[0018] In the above technical solution, in step (4), the detection wavelength is 210 nm and the flow rate is 3 mL / min.

[0019] Another object of the present invention is to provide a pharmaceutical composition comprising the above-described compound or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier thereof.

[0020] Another object of the present invention is to provide the use of the above-described compounds or pharmaceutically acceptable salts thereof or the above-described pharmaceutical compositions in the preparation of medicaments for the prevention or treatment of lung diseases.

[0021] The beneficial effects of this invention are as follows: This invention provides for the first time a method for preparing and identifying 14 new compounds using the whole herb *Prunella vulgaris* as raw material, and systematically evaluates their lung-protective activity, elucidating their application in the development of chemopreventive and therapeutic drugs related to lung inflammation. Using MLE-12 cells as an in vitro model, this invention tested the lung-inflammatory activity of the prepared new compounds 1-14 against NNK-induced MLE-12 cells. The results show that these new compounds have significant lung-protective activity and can be used to develop chemopreventive or therapeutic drugs for lung diseases. Detailed Implementation

[0022] The following non-limiting embodiments are intended to enable those skilled in the art to more fully understand the invention, but do not limit the invention in any way.

[0023] Unless otherwise specified, the experimental methods described in the following examples are conventional methods; the reagents and materials described are commercially available unless otherwise specified.

[0024] Example 1 (1) 1 kg of the whole herb of Wild Horse Chase was extracted three times by heating and refluxing with 95% ethanol (dosage: 10 L), and the crude extract was obtained by vacuum recovery. (2) The crude extract obtained in step (1) above is dissolved in an organic solvent (methanol), then mixed with macroporous adsorption resin and dried. It is then separated by D101 macroporous adsorption resin and eluted sequentially with mixed solvents of pure water-ethanol volume ratios of 100:0, 70:30, 50:50, 30:70, and 5:95. (3) The pure water-ethanol fraction with a volume ratio of 100:0 to 50:50 obtained in step (2) above was separated by ODS chromatography and eluted with a methanol-water volume ratio of 1:9, 3:7, 5:5, 7:3, 9:1. (4) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 30:70, to obtain compound 1 (t R = 21 min (yield 0.0028%); (5) The methanol-water fraction with a volume ratio of 3:7 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 33:67, to obtain compound 2 (t R = 31 min (yield 0.0005%); (6) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 25:75, to obtain compound 3 (t R = 33 min (yield 0.0014%); (7) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 16:84, to obtain compound 4 (t R = 36 min (yield 0.0017%); (8) The methanol-water fraction with a volume ratio of 3:7 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 30:70, to obtain compound 5 (t R = 25 min (yield 0.0010%); (9) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 25:75, to obtain compound 6 (t R= 45 min (yield 0.0017%); (10) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile-water with a volume ratio of 25:75, to obtain compound 7 (t R = 33 min (yield 0.0020%); (11) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 42:58, to obtain compound 8 (t R = 35 min (yield 0.0004%); (12) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 25:75, to obtain compound 9 (t R = 21 min (yield 0.0005%).

[0025] (13) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 22:78, to obtain compound 10 (t R = 31 min (yield 0.0007%).

[0026] (14) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 20:80, to obtain compound 11 (t R = 23 min (yield 0.0014%).

[0027] (15) The methanol-water fraction with a volume ratio of 5:5 to 7:3 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 33:67, to obtain compound 12 (t R = 35 min (yield 0.0012%).

[0028] (16) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 21:79, to obtain compound 13 (t R = 27 min (yield 0.0014%).

[0029] (17) The methanol-water fraction with a volume ratio of 5:5 to 7:3 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 39:61, to obtain compound 14 (t R = 31 min (yield 0.0002%).

[0030] The structures of compounds 1-14 were identified based on their physicochemical properties and spectroscopic data.

[0031] The structural identification data of compound 1 are as follows: Transparent oil (methanol) -24.7 ( c 0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 445.1838 [M+Na] + (calcd. 445.1845 for C 22 H 30 O8Na). The molecular formula of the compound is presumed to be C. 22 H 30 O8 has an unsaturation degree of 8. 1 The H-NMR (600 MHz, CD3OD) spectrum gives three olefin hydrogen signals: d H 5.46 (1H, d, J =4.7 Hz, H-4), 6.44 (1H, d, J = 1.1 Hz, H-13a), 5.63 (1H, s, H-13b); 2 sets of oxygen-hydrogen signals: d H 5.44 (1H, t, J = 8.7 Hz, H-2), 5.27 (1H, q, J = 3.1 Hz, H-7); 3 sets of methylene hydrogen signals: d H 2.61 (1H, t, J = 12.0 Hz, H-5), 2.74 (1H, dd, J= 12.7, 2.8 Hz, H-6), 1.66 (1H, m, H-10); 2 sets of methylene hydrogen signals: d H 2.49 (1H, ddd, J = 11.8, 6.3, 1.9 Hz, H-1a),1.43 (1H, ddd, J = 12.8, 11.9, 10.1 Hz, H-1b), 2.10 (1H, dd, J = 14.5, 3.0 Hz, H-8a), 1.86 (1H, dd, J = 14.4, 3.4 Hz, H-8b); 2 sets of methyl hydrogen signals: d H 1.17 (3H, s, C H 3-14), 1.65 (3H, t, J = 1.8, 1.8 Hz, C H 3-15) indicates that compound 1 is a juniperane-type sesquiterpene lactone. Additionally, a set of acetyl hydrogen signals is given: d H 2.06 (3H, s, C H 3-2'') and a set of side-chain hydrogen signals: d H 6.78(1H, t, J = 5.9 Hz, H-3'), 4.28 (2H, dd, J = 6.0 Hz, H-4'), 1.80 (3H, d, J = 1.3Hz, C H 3-5'). 13 C-NMR (150 MHz, CD3OD) yielded 22 carbon signals, including 15 carbon signals from the parent nuclei of juniperane-type sesquiterpene lactones: d C 29.7 (C-1), 75.0 (C-2), 136.1 (C-3), 129.1 (C-4), 37.2 (C-5), 47.2 (C-6), 71.8 (C-7), 45.7 (C-8), 70.9 (C-9), 50.1 (C-10), 139.4 (C-11), 170.0 (C-12), 128.9 (C-13), 23.4 (C-14), 19.3 (C-15). Additionally, a set of side-chain carbon signals is given: d C170.0 (C-1'), 128.3 (C-2'), 142.8 (C-3'), 59.1 (C-4'), 12.8 (C-5') and one set of acetyl carbon signals: d C 172.7 (C-1''), 21.0 (C-2''). In summary, this suggests that compound 1 is a juniperane-type sesquiterpene lactone with one side chain and one acetyl-substituted group. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, [the following can be observed]... d H 2.21 (Ha-8) and d C 23.4 (C-14), 47.2 (C-6), and 70.9 (C-9) are relevant. d H 1.66 (H-10) and d C 23.3 (C-14), 29.7 (C-1), and 70.9 (C-9) are relevant. d H 2.49 (H-1a) and d C The values ​​37.2 (C-5), 50.1 (C-10), 75.0 (C-2), and 136.1 (C-3) are relevant. d H 2.61 (H-5) and d C The related proofs for 47.2 (C-6), 50.1 (C-10), 129.1 (C-4), and 136.1 (C-3) demonstrate the existence of two hexacyclic rings that are coupled at positions 5 and 10. d H 6.44 (H-13a) and d C The correlation between 47.2 (C-6) and 170.0 (C-12) proves that the unsaturated lactone is attached at the C-6 position; d H 5.27 (H-7) and d C The correlation at 170.0 (C-1') proves that the sidechain is linked at position C-7. d H 1.25 (C H 3-14) and d C The correlation between 45.7 (C-8), 50.1 (C-10), and 70.9 (C-9) proves that CH3-14 is attached to the C-9 position. Combined with HSQC, it can be known that C-9 is an oxygen-bound quaternary carbon, and therefore it is also attached to a hydroxyl group. d H 1.61 (C H 3-15) and d C The correlation between 75.0 (C-2), 129.0 (C-4), and 136.1 (C-3) proves that CH3-15 is connected at position C-3; d H 5.44 (H-2) and d C 172.7 (C-1'') related, d H 2.06 (C H 3-2'') and d C The correlation at 172.7 (C-1'') indicates that the acetyl group is attached at the C-2 position. In the NOESY spectrum, d H 1.25 (C H 3-14) and d H Related to 2.61 (H-5) and 2.10 (H-8a), d H 1.66 (H-10) and d H The correlation between 2.61 (H-6), 5.44 (H-2), and 1.86 (H-8b) proves that H-5 and C... H 3-14 have the same orientation, H-2, H-6, H-10 and H-5, C H The orientations of positions 3-14 are opposite. The relative configuration at position H-7 cannot be determined based on the NOESY spectrum. The relative configuration of this compound was ultimately determined by calculating the carbon NMR spectrum, comparing the two relative configurations. R , 5 R , 6 S , 7 R 9 S , 10 S (1a) and 2 S , 5 S , 6 R , 7 S 9R , 10 R (1b) Perform DP4+ carbon spectrum calculations. Measured data and 2 S , 5 S , 6 R , 7 S 9R , 10 R (1b) Linear correlation coefficient R of the calculated data of the configuration 2 The value is 0.9984, and the DP4+ probability is 99.97%, thus determining the relative configuration of compound 1 to be 2. S , 5 S , 6 R , 7 S 9R , 10 R The NOESY spectral analysis results were verified. The absolute configuration was determined by comparing the calculated ECD with the measured spectrum, confirming that the absolute configuration of compound 1 was 2. S , 5 S , 6 R , 7 S , 9R, 10 R In summary, based on Sci-finder searches, this compound is a novel compound that has not been previously reported, and it has been named eupacadinolide G.

[0032] The structural identification data of compound 2 are as follows: Transparent oil (methanol) -15.3 ( c 0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 445.1837 [M+Na] + (calcd. 445.1833 for C 20 H 30 O8Na). The molecular formula of the compound is presumed to be C. 22 H 30 O8 has an unsaturation degree of 8. 1 Two sets of double bond hydrogen signals are given in the H-NMR (600 MHz, CD3OD) spectrum: d H 5.47 (1H, q, J=1.6 Hz, H-4), 6.44 (1H, d, s, H-13a), 5.59 (1H, s, H-13b); 2 oxygen-hydrogen signals: d H 5.43 (1H, t, J = 7.2 Hz, H-2), 5.36 (1H, q, J = 3.2 Hz, H-7); 3 sets of methylene hydrogen signals: d H 2.54 (1H, t, J = 11.4 Hz, H-5), 2.74 (1H, dd, J = 12.8, 2.8 Hz, H-6), 1.64 (1H,ddd, J = 12.7, 10.4, 1.8 Hz, H-10); 2 sets of methylene hydrogen signals: d H 2.48 (1H, ddd, J = 11.9,6.4, 1.9 Hz, H-1a), 1.41 (1H, td, J = 12.5, 12.4, 10.2 Hz, H-1b), 2.11 (1H, dd, J = 14.4, 3.0 Hz, H-8a), 1.87 (1H, dd, J = 14.4, 3.4 Hz, H-8b); 2 sets of methyl hydrogen signals: d H 1.25 (3H, s, C H 3-14), 1.60 (3H, s, C H 3-15) suggests that compound 2 is a juniperane-type sesquiterpene lactone. Additionally, a set of acetyl hydrogen signals is given: d H 2.07 (3H, s, C H 3-2'') and a set of side-chain hydrogen signals: d H 6.32(1H, q J = 7.3 Hz, H-3'), 1.98 (3H, d, J = 7.3 Hz, H-4'), 4.19 (2H, d, J = 4.6 Hz, H-5'). 13C-NMR (150 MHz, CD3OD) yielded 22 carbon signals, including 15 carbon signals from the parent nuclei of juniperane-type sesquiterpene lactones: d C 29.8 (C-1), 75.0 (C-2), 136.0 (C-3), 129.2 (C-4), 37.1 (C-5), 47.2 (C-6), 71.3 (C-7), 45.8 (C-8), 70.9 (C-9), 50.1 (C-10), 139.4 (C-11), 170.4 (C-12), 128.1 (C-13), 23.2 (C-14), 19.3 (C-15). Additionally, a set of side-chain carbon signals is given: d C 167.1 (C-1'), 133.7 (C-2'), 139.8 (C-3'), 15.6 (C-4'), 63.7 (C-5') and one set of acetyl carbon signals: d C 172.7 (C-1''), 21.0 (C-2''). In summary, this suggests that compound 2 is a juniperane-type sesquiterpene lactone with one side chain and one acetyl-substituted group. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, it can be observed that... d H 2.11 (H-8a) and d C 47.2 (C-6) related, d H 1.64 (H-10) and d C 23.2 (C-14) and 70.9 (C-9) are relevant. d H 2.48 (H-1a) and d C 37.1 (C-5) and 136.0 (C-3) are relevant. d H 2.61 (H-5) and d C 136.0 (C-3) proves the existence of two six-membered rings that are coupled at bits 5 and 10; d H 6.44 (H-13a) and d C The correlation between 47.2 (C-6) and 170.4 (C-12) proves that the unsaturated lactone is attached at the C-6 position; d H 5.36 (H-7) and d C The correlation at 167.1 (C-1') proves that the sidechain is linked at position C-7. d H 1.25 (C H 3-14) and d C The correlation between 45.8 (C-8), 50.1 (C-10), and 70.9 (C-9) proves that CH3-14 is attached to the C-9 position. Combined with HSQC, it can be known that C-9 is an oxygen-bound quaternary carbon, and therefore it is also attached to a hydroxyl group. d H 1.60 (C H 3-15) and d C The correlation between 75.0 (C-2), 129.2 (C-4), and 136.0 (C-3) proves that CH3-15 is connected at position C-3; d H 5.43 (H-2) and d C The correlation at 172.7 (C-1'') proves that the acetyl group is attached at the C-2 position. In the NOESY spectrum, d H 1.25 (C H 3-14) and d H 2.54 (H-5) and 2.10 (H-8a) are related. d H 2.54 (H-5) and d H 5.36 (H-7) related, d H 1.64 (H-10) and d H The correlations of 2.74 (H-6), 5.43 (H-2), and 1.87 (H-8b) prove the existence of H-5, H-7, and C. H 3-14 have the same orientation, H-2, H-6, H-10 and H-5, H-7, C H Since 3-14 have opposite orientations, it is inferred that the relative configuration of the compound is 2. S , 5 S , 6 R , 7 S , 9 R , 10 R The absolute configuration of compound 2 was determined by comparing calculated ECD with measured spectra. S , 5 S , 6 R , 7 S , 9 R 10 R In summary, based on Sci-finder searches, this compound is a novel compound that has not been previously reported, and it has been named eupacadinolide H.

[0033] Table 1. Attribution of 1H NMR and 1C NMR data for compounds 1-2

[0034] The structural identification data of compound 3 are as follows: White powder (methanol) -120.7 ( c 0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 461.1786 [M+Na] + (calcd. 461.1782 for C 22 H 30 O9Na). The molecular formula of the compound is presumed to be C. 22 H 30 O9 has an unsaturation degree of 8. 1 Two sets of double bond hydrogen signals are given in the H-NMR (600 MHz, CD3OD) spectrum: d H 5.76 (1H, dd,m, H-4), 6.41 (1H, d, J = 1.2 Hz, H-13a), 5.76 (1H, m, H-13b); 2 oxygen-hydrogen signals: d H 5.37 (1H, dd, J = 7.2, 9.6 Hz, H-2), 5.20 (1H, q, J = 2.8, 2.8, 2.9 Hz, H-7); 3 sets of methylene hydrogen signals: d H 3.05 (1H, m, H-5), 3.05 (1H, m, H-6), 1.98 (1H, dd, J = 2.4, 14.3 Hz, H-10); 2 sets of methylene hydrogen signals: d H2.11 (1H, m, H-1a), 1.59 (1H, m, H-1b), 1.91 (1H, dt, J = 2.3, 2.3, 15.0 Hz, H-8a), 1.79 (1H, dd, J = 3.4, 15.3 Hz, H-8b); 2 sets of methyl hydrogen signals: d H 1.21 (3H, s, C H 3-14), 1.59 (3H, s, C H 3-15) indicates that compound 3 is a juniperane-type sesquiterpene lactone. Additionally, a set of acetyl hydrogen signals is given: d H 2.06 (3H, s, C H 3-2'') and a set of side-chain hydrogen signals: d H 6.87 (1H, t, J = 6.0, 6.0 Hz, H-3'), 4.34 (2H, m, H-4'), 4.28 (2H,s, H-5'). 13 C-NMR (150 MHz, CD3OD) yielded 22 carbon signals, including 15 carbon signals from the parent nuclei of juniperane-type sesquiterpene lactones: d C 29.8 (C-1), 75.0 (C-2), 136.0 (C-3), 129.2 (C-4), 37.1 (C-5), 47.2 (C-6), 71.3 (C-7), 45.8 (C-8), 70.9 (C-9), 50.1 (C-10), 139.4 (C-11), 170.4 (C-12), 128.1 (C-13), 23.2 (C-14), 19.3 (C-15). Additionally, a set of side-chain carbon signals is given: d C 167.1 (C-1'), 133.7 (C-2'), 139.8 (C-3'), 15.6 (C-4'), 63.7 (C-5') and one set of acetyl carbon signals: d C 172.7 (C-1''), 21.0 (C-2''). In summary, this suggests that compound 3 is a juniperane-type sesquiterpene lactone with one side chain and one acetyl-substituted group. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, it can be observed that... d H2.11 (H-1a) and d C 31.3 (C-5) and 134.9 (C-3) are relevant. d H 1.91 (H-8a) and d C Correlation between 43.4 (C-6) and 45.6 (C-10) proves the existence of two hexacyclic rings, which are concatenated at positions 5 and 10. d H 6.41 (H-13a) and d C The correlation between 43.4 (C-6) and 171.4 (C-12) proves that the unsaturated lactone is attached at the C-6 position; d H 5.20 (H-7) and d C The correlation at 167.1 (C-1') proves that the sidechain is linked at position C-7. d H 1.21 (C H 3-14) and d C The correlation between 37.9 (C-8), 45.6 (C-10), and 72.5 (C-9) proves that CH3-14 is attached to the C-9 position. Combined with HSQC, it can be known that C-9 is an oxygen-bound quaternary carbon, and therefore it is also attached to a hydroxyl group. d H 1.59 (C H 3-15) and d C The correlation between 74.6 (C-2), 130.5 (C-4), and 134.9 (C-3) proves that CH3-15 is connected at position C-3; d H 5.37 (H-2) and d C The correlation at 172.7 (C-1'') indicates that the acetyl group is attached at the C-2 position. In the NOESY spectrum, d H 1.21(C H 3-14) and d H 1.98 (H-10) related, d H 5.37 (H-2) and d H 1.98 (H-10) correlation, prove H-2, H-10, C HPositions 3-14 have the same orientation. The relative configurations at positions H-6, H-7, and H-8 could not be determined using NOESY spectroscopy. The relative configurations of this compound were ultimately determined by calculating carbon nanotube spectra, and the four relative configurations were... R , 5 S , 6 S , 7 S , 9 R , 10 S (3a), 2 R , 5 S , 6 S , 7 R 9R , 10 S (3b), 2 R , 5 S , 6 R , 7 S , 9 R , 10 S (3c) and 2 R , 5 R , 6 S , 7 S 9R , 10 S (3d) Perform DP4+ carbon spectrum calculations. Measured data and 2 R 5 S , 6 S , 7 S , 9 R , 10 S (3a) Linear correlation coefficient R of the calculated data of the configuration 2The value is 0.9985, and the DP4+ probability is 99.91%, thus determining the relative configuration of compound 3 to be 2. R , 5 S , 6 S , 7 S , 9 R , 10 S The NOESY spectral analysis results were verified. The absolute configuration was determined by comparing the calculated ECD with the measured spectrum, confirming that the absolute configuration of compound 3 is 2. R , 5 S , 6 S , 7 S , 9 R , 10 S In summary, based on Sci-finder search results, this compound is a novel compound that has not been previously reported, and it has been named eupacadinolide I.

[0035] The structural identification data of compound 4 are as follows: Needle crystals (methanol) -118.7 ( c 0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 461.1783 [M+Na] + (calcd. 461.1782 for C 22 H 30 O9Na). The molecular formula of the compound is presumed to be C. 22 H 30 O9 has an unsaturation degree of 8. 1 Two sets of double bond hydrogen signals are given in the H-NMR (600 MHz, CD3OD) spectrum: d H 5.60 (1H, t, J =1.7, 1.7 Hz, H-4), 6.42 (1H, s, H-13a), 5.62 (1H, s, H-13b); 2 hydroxymethyl hydrogen signals: d H 5.23 (1H, d, J = 4.0 Hz, H-2), 5.31 (1H, q, J = 3.1, 3.1, 3.0 Hz, H-7); 3 sets of methylene hydrogen signals: dH 1.87 (1H, dd, J = 3.4, 14.4 Hz, H-10), 2.47 (1H, m, H-5), 2.77(1H, dd, J = 2.7, 12.7 Hz, H-6); 2 sets of methylene hydrogen signals: d H 2.14 (1H, dd, J = 3.0, 14.4 Hz,H-1a), 1.63 (1H, m, H-1b), 1.56 (1H, td, J = 4.3, 13.3, 13.5 Hz, H-8a) , 2.23(1H, dt, J = 1.9, 1.9, 13.9 Hz, H-8b); 2 sets of methyl hydrogen signals: d H 1.24 (3H, s, C H 3-14), 1.66(3H, t, J = 1.8, 1.8 Hz, C H (3-15) The above information suggests that compound 4 is a juniperane-type sesquiterpene lactone compound. In addition, a set of acetyl hydrogen signals is given: d H 2.06 (3H, s, C H 3-2''), a set of side-chain hydrogen signals: d H 6.89 (1H,t, J = 6.0 Hz, H-3'), 4.39 (2H, d, J = 6.0 Hz, H-4'), 4.27 (2H, s, H-5'). 13 C-NMR (150 MHz, CD3OD) yielded 22 carbon signals, including 15 carbon signals from the parent nucleus of juniperane-type sesquiterpene lactones: d C 45.8 (C-1), 72.2 (C-2), 134.1 (C-3), 130.5 (C-4), 37.2 (C-5), 47.1 (C-6), 71.6 (C-7), 29.6 (C-8), 71.2 (C-9), 45.7 (C-10), 139.8 (C-11), 170.7 (C-12), 128.1 (C-13), 23.6 (C-14), 20.9 (C-15). In addition, a set of side-chain carbon signals is also given: d C 167.1 (C-1'), 132.8 (C-2'), 146.2 (C-3'), 59.4 (C-4'), 56.9 (C-5') and one set of acetyl carbon signals: d C 172.8 (C-1''), 21.2 (C-2''). In summary, this suggests that compound 4 is a juniperane-type sesquiterpene lactone compound with an acetyl group and one side chain substitution. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, [the following can be observed]. d H 2.23(H-1b) and d C 37.2 (C-5) and 134.1 (C-3) are relevant. d H 5.60 (H-4) and d C 72.2 (C-2) related, d H 5.31 (H-7) and d C Correlation between 37.2 (C-5) and 71.2 (C-9) proves the existence of two hexacyclic rings, which are concatenated at positions 5 and 10. d H 6.42 (H-13a) and d C 47.1 (C-6) correlation, proving that the unsaturated lactone is attached at the C-6 position; d H 5.31 (H-7) and d C 167.1 (C-1') is relevant, proving that the sidechain is linked at position C-7; d H 1.24 (C H 3-14) and d C The correlation between 45.7 (C-10) and 71.2 (C-9) proves that CH3-14 is attached to the C-9 position. Combined with HSQC, it can be known that C-9 is an oxygen-bound quaternary carbon, and therefore it is also attached to a hydroxyl group. d H 1.66 (C H 3-15) and d C The correlation between 72.2 (C-2), 130.5 (C-4), and 134.1 (C-3) proves that CH3-15 is connected at position C-3; d H 5.23(H-2) and d C The correlation at 172.8 (C-1'') indicates that the acetyl group is attached at the C-2 position. In the NOESY spectrum, d H 1.19 (C H 3-14) and d H 2.14 (H-1a), 2.47 (H-5), and 1.56 (H-8a) are related, and 1.56 (H-8a) is related to... d H 5.22 (H-2) related, d H 2.14(H-1a) and d H 1.87 (H-10) and 5.30 (H-7) are relevant. d H 2.77 (H-6) and d H 1.63 (H-1b) related, prove H-2, H-5, H-7, H-10, C H Compounds 3-14 have the same orientation, while H-6 has the opposite orientation. The absolute configuration was determined by comparing calculated ECD with measured spectra, confirming that the absolute configuration of compound 4 is 2. S , 5 S , 6 R , 7 S , 9R, 10 S In summary, based on Sci-finder search results, this compound is a novel compound that has not been previously reported, and it has been named eupacadinolide J.

[0036] The structural identification data of compound 5 are as follows: Yellow powder (methanol) -116.0 ( c 0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 461.1786 [M+Na] + (calcd. 461.1782 for C 22 H 30 O9Na). The molecular formula of the compound is presumed to be C. 22 H 30 O9 has an unsaturation degree of 8. 1 Two sets of double bond hydrogen signals are given in the H-NMR (600 MHz, CD3OD) spectrum: d H 5.64 (1H, t, J=1.7, 1.7 Hz, H-4), 6.44 (1H, s, H-13a), 5.60 (1H, s, H-13b), 6.87 (1H, t, J =6.0 Hz, H-3'); 2 hydroxymethyl hydrogen signals: d H 5.26 (1H, d, J = 4.15 Hz, H-2), 5.28 (1H,d, J = 2.8 Hz, H-7); 3 sets of methylene hydrogen signals: d H 1.51 (1H, ddd, J = 2.1, 10.3, 12.8 Hz, H-10), 2.71 (1H, t, J = 11.6, 11.6 Hz, H-5), 2.81 (1H, dd, J = 2.5, 12.7 Hz, H-6); 2 sets of methylene hydrogen signals: d H 2.09 (1H, m, H-1a), 1.74 (1H, m, H-1b), 1.85 (1H, dd, J = 3.2, 15.1 Hz, H-8a), 2.06 (1H, m, H-8b); 2 sets of methyl hydrogen signals: d H 1.17 (3H, s, C H 3-14), 1.65(3H, t, J = 1.8, 1.8 Hz, C H 3-15). The above information suggests that compound 5 is a juniperane-type sesquiterpene lactone compound. In addition, a set of acetyl hydrogen signals is given: d H 2.06 (3H, s, C H 3-2''), a set of side-chain hydrogen signals: d H 6.87 (1H,t, J = 6.0 Hz, H-3'), 4.34 (2H, m, H-4'), 4.27 (2H, m, H-5'). 13 C-NMR (150 MHz, CD3OD) yielded 22 carbon signals, including 15 carbon signals from the parent nucleus of juniperane-type sesquiterpene lactones: d C30.0 (C-1), 71.7 (C-2), 133.7 (C-3), 130.9 (C-4), 34.6 (C-5), 46.7 (C-6), 71.9 (C-7), 43.3 (C-8), 70.8 (C-9), 44.5 (C-10), 140.3 (C-11), 171.2 (C-12), 127.8 (C-13), 28.2 (C-14), 20.8 (C-15). Additionally, a set of side-chain carbon signals is given: d C 167.2 (C-1'), 133.0 (C-2'), 145.3 (C-3'), 59.1 (C-4'), 57.0 (C-5') and one set of acetyl carbon signals: d C 172.7 (C-1''), 21.2 (C-2''). In summary, this suggests that compound 5 is a juniperane-type sesquiterpene lactone compound with an acetyl group and one side chain substitution. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, [the following can be observed]. d H 2.09 (H-1a) and d C 34.6 (C-5) and 133.7 (C-3) are relevant. d H 5.64 (H-4) and d C 44.5 (C-10) and 71.7 (C-2) are relevant. d H 5.28 (H-7) and d C 34.6 (C-5) and 70.8 (C-9) are relevant. d H 2.21 (H-8b) and d C Correlation between 44.5 (C-10) and 46.7 (C-6) proves the existence of two hexacyclic rings, which are concatenated at positions 5 and 10. d H 6.44 (H-13a) and d C 46.7 (C-6) correlation, proving that the unsaturated lactone is attached at the C-6 position; d H 5.28 (H-7) and d C The correlation at 167.2 (C-1') proves that the sidechain is linked at position C-7. dH 1.17 (C H 3-14) and d C The correlation between 43.3 (C-8) and 70.8 (C-9) proves that CH3-14 is attached to the C-9 position. Combined with HSQC, it can be known that C-9 is an oxygen-bound quaternary carbon, and therefore it is also attached to a hydroxyl group. d H 1.65 (C H 3-15) and d C The correlation between 71.7 (C-2), 130.9 (C-4), and 133.7 (C-3) proves that CH3-15 is connected at position C-3; d H 5.26 (H-2) and d C The correlation at 172.7 (C-1'') indicates that the acetyl group is attached at the C-2 position. In the NOESY spectrum, d H 1.19 (C H 3-14) and d H 1.51 (H-10) and 2.09 (H-1a) are relevant. d H 1.51 (H-10) and d H 2.81 (H-6) related, d H 2.71 (H-5) and d H 1.73 (H-1b) related, prove H-6, H-10, C H Positions 3-14 have the same orientation, while H-5 has the opposite orientation. The relative configurations of positions H-2 and H-7 cannot be determined based on the NOESY spectrum. The relative configurations of this compound were ultimately determined by calculating the carbon nanotube spectrum, and the two relative configurations were compared. R , 5 R , 6 S 7 R , 9 R , 10 S (5a) and 2 S , 5 S , 6 R , 7 S 9S , 10 R (5b) Perform DP4+ carbon spectrum calculations. Measured data and 2 S , 5 S , 6 R , 7 S 9S , 10 R (5b) Linear correlation coefficient R of the calculated data of the configuration 2 The value is 0.9985, and the DP4+ probability is 100.00%, thus determining the relative configuration of compound 5 to be 2. S , 5 S , 6 R 7 S 9S , 10 R The NOESY spectral analysis results were verified. The absolute configuration was determined by comparing the calculated ECD with the measured spectrum, confirming that the absolute configuration of compound 5 was 2. S , 5 S , 6 R , 7 S , 9S, 10 R In summary, based on Sci-finder searches, this compound is a novel compound that has not been previously reported, and it has been named eupacadinolide K.

[0037] Table 2. Attribution of 1H NMR and 1C NMR data for compounds 3-5

[0038] The structural identification data for compound 6 are as follows: Needle crystals (methanol) -21.3 ( c 0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 475.1938 [M+Na] + (calcd. 475.1940 for C 23 H 32 O9Na). The molecular formula of the compound is presumed to be C. 23 H 32O9 has an unsaturation degree of 8. 1 Two sets of double bond hydrogen signals are given in the H-NMR (600 MHz, CD3OD) spectrum: d H 5.62 (1H, t, J =1.7, 1.7 Hz, H-4), 6.40 (1H, s, H-13a), 5.59 (1H, s, H-13b); 2 sets of hydroxymethyl hydrogen signals: d H 5.21 (1H, d, J = 3.95 Hz, H-2), 5.37 (1H, d, J = 3.1 Hz, H-7); 3 sets of methylene hydrogen signals: d H 1.77 (1H, ddd, J = 2.1, 10.7, 13.0 Hz, H-10), 2.53 (1H, t, J = 11.8, 11.8Hz, H-5), 2.76 (1H, dd, J = 2.6, 12.9 Hz, H-6); 2 sets of methylene hydrogen signals: d H 2.11 (1H, dt, J = 2.0, 2.0, 14.2 Hz, H-1a), 1.53 (1H, ddd, J = 4.3, 12.8, 14.2 Hz, H-1b), 2.22(1H, dd, J = 3.0, 14.1 Hz, H-8a), 1.90 (1H, dd, J = 3.3, 14.1 Hz, H-8b); 2 sets of methyl hydrogen signals: d H 1.26 (3H, s, C H 3-14), 1.66 (3H, d, J = 1.9 Hz, C H 3-15) indicates that compound 6 is a juniperane-type sesquiterpene lactone. Additionally, a set of acetyl hydrogen signals is given: d H 2.06 (3H, s, C H 3-2''), a set of side-chain hydrogen signals: d H 6.90 (1H, t, J= 6.0, 6.0 Hz, H-3'), 4.40 (2H, d, J = 5.9 Hz, H-4'), 4.28 (2H, s, H-5'), and a set of methoxy hydrogen signals: d H 3.21 (3H, s, OC H 3-9). 13 C-NMR (150MHz, CD3OD) yielded 23 carbon signals, including 15 carbon signals from the parent nucleus of juniperane-type sesquiterpene lactones: d C 29.5 (C-1), 71.6 (C-2), 134.0 (C-3), 130.6 (C-4), 37.0 (C-5), 47.1 (C-6), 72.0 (C-7), 40.0 (C-8), 75.9 (C-9), 42.9 (C-10), 140.3 (C-11), 169.5 (C-12), 127.6 (C-13), 21.0 (C-14), 20.9 (C-15). Additionally, a set of side-chain carbon signals is given: d C 167.1 (C-1'), 132.8 (C-2'), 146.2 (C-3'), 59.5 (C-4'), 56.9 (C-5') and a methoxy carbon signal: d C 49.9(O C H3-9), Group 1 acetyl carbon signal: d C 172.7 (C-1''), 21.2 (C-2''). In summary, this suggests that compound 6 is a juniperane-type sesquiterpene lactone compound with acetyl, methoxy, and one side chain substitution. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, [the following can be observed]... d H 2.11 (H-1a) and d C 37.0 (C-5) and 134.0 (C-3) are relevant. d H 2.22(H-8a) and d C Related to 42.9 (C-10) and 47.1 (C-6) d H 1.77 (H-10) and d C71.6 (C-2) related, proves the existence of two six-membered rings through 5- and 10-bit concatenation; d H 6.40 (H-13a) and d C 47.1 (C-6) correlation, proving that the unsaturated lactone is attached at the C-6 position; d H 5.37 (H-7) and d C The correlation at 167.1 (C-1') proves that the sidechain is linked at position C-7. d H 1.26 (C H 3-14) and d C The relevant proofs for 40.0 (C-8) and 75.9 (C-9) show that CH3-14 is connected at position C-9; d H 3.21 (OC H 3-9) and d C The relevant evidence for 75.9 (C-9) shows that the methoxy group is also attached at the C-9 position; d H 1.66 (C H 3-15) and d C The correlation between 71.6 (C-2) and 134.0 (C-3) proves that CH3-15 is connected at position C-3; d H 5.21 (H-2) and d C The correlation at 172.7 (C-1'') indicates that the acetyl group is attached at the C-2 position. In the NOESY spectrum, d H 1.26 (C H 3-14) and d H 1.53 (H-1b) and 2.53 (H-5) are related. d H 1.77 (H-10) and d H 2.11 (H-1a) and 2.76 (H-6) are related, proving H-5 and C H Positions 3-14 have the same orientation, while H-10 and H-6 have opposite orientations. The relative configurations of positions H-2 and H-7 cannot be determined based on the NOESY spectrum. The relative configurations of this compound were ultimately determined by calculating the carbon NMR spectrum, and the two relative configurations were compared. S , 5 S , 6 R , 7 R , 9 R , 10 R (6a) and 2 S , 5 S , 6 R , 7 S 9R , 10 R (6b) Perform DP4+ carbon spectrum calculations. Measured data and 2 S , 5 S , 6 R , 7 S 9R , 10 R (6b) Linear correlation coefficient R of the calculated data of the configuration 2 The value is 0.9986, and the DP4+ probability is 100.00%, thus determining the relative configuration of compound 6 to be 2. S , 5 S , 6 R , 7 S 9R , 10 R The NOESY spectral analysis results were verified. The absolute configuration was determined by comparing the calculated ECD with the measured spectrum, confirming that the absolute configuration of compound 6 was 2. S , 5 S , 6 R , 7 S ,9R, 10 R In summary, based on Sci-finder search results, this compound is a novel compound that has not been previously reported, and it has been named eupacadinolide L.

[0039] The structural identification data of compound 7 are as follows: Yellow-green crystals (methanol) +42.0 ( c0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 475.1940 [M+Na] + (calcd. 475.1938 for C 23 H 32 O9Na). The molecular formula of the compound is presumed to be C. 23 H 32 O9 has an unsaturation degree of 8. 1 Two sets of double bond hydrogen signals are given in the H-NMR (600 MHz, CD3OD) spectrum: d H 5.59 (1H, dd, m, H-4), 6.44 (1H, s, H-13a), 5.65 (1H, s, H-13b); 2 hydroxymethyl hydrogen signals: d H 5.21(1H, m, H-2), 5.36 (1H, d, J = 3.1 Hz, H-7); 3 sets of methylene hydrogen signals: d H 1.76 (1H, m, H-10), 2.53 (1H, t, J = 11.8, 11.8 Hz, H-5), 2.75 (1H, dd, J = 2.6, 12.3 Hz, H-6); 2 sets of methylene hydrogen signals: d H 2.12 (1H, d, J = 2.5 Hz, H-1a), 1.53 (1H, td, J = 4.3, 13.5,13.6 Hz, H-1b), 1.89 (1H, dd, J = 3.3, 14.5 Hz, H-8a), 2.21 (1H, dd, J = 3.0, 14.2 Hz, H-8b); 2 sets of methyl hydrogen signals: d H 1.26 (3H, s, C H 3-14), 1.65 (3H, s, C H 3-15); This suggests that compound 7 is a juniperane-type sesquiterpene lactone. Additionally, a set of acetyl hydrogen signals is given: d H 2.05 (3H, s, C H 3-2''), a set of side-chain hydrogen signals: dH 6.89 (1H, t, J = 5.8, 5.8 Hz, H-3'), 4.39 (2H, d, J = 5.8,H-4'), 4.27 (2H, s, H-5'); and a set of methoxy hydrogen signals: d H 3.20 (3H, s, OC H 3-9). 13 C-NMR (150 MHz, CD3OD) yielded 23 carbon signals, including 15 carbon signals from the parent nucleus of juniperane-type sesquiterpene lactones: d C 21.2 (C-1), 71.9 (C-2), 134.1 (C-3), 130.8 (C-4), 36.9 (C-5), 47.0 (C-6), 71.6 (C-7), 40.0 (C-8), 75.8 (C-9), 42.9 (C-10), 139.4 (C-11), 170.3 (C-12), 128.5 (C-13), 29.5 (C-14), 20.9 (C-15). Additionally, a set of side-chain carbon signals is given: d C 167.1 (C-1'), 132.8 (C-2'), 146.3 (C-3'), 59.4 (C-4'), 56.9 (C-5') and a methoxy carbon signal: d C 49.6 (O C H3-9), Group 1 acetyl carbon signal: d C 172.7 (C-1''), 21.0 (C-2''). In summary, this suggests that compound 7 is a juniperane-type sesquiterpene lactone compound with acetyl, methoxy, and one side chain substitution. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, [the following can be observed]... d H 2.12 (H-1a) and d C 36.9 (C-5), 75.8 (C-9), and 134.1 (C-3) are relevant. d H 2.53 (H-5) and d C 134.1 (C-3) related, d H 2.21 (H-8b) and dC 42.9 (C-10) and 47.0 (C-6) are relevant. d H 1.76 (H-10) and d C 47.0 (C-6) is relevant, proving the existence of two six-membered rings through 5- and 10-bit concatenation; d H 6.44 (H-13a) and d C 47.0 (C-6) correlation, proving that the unsaturated lactone is linked at the C-6 position; d H 5.37 (H-7) and d C The correlation at 167.1 (C-1') proves that the sidechain is linked at position C-7. d H 1.26 (C H 3-14) and d C The relevant proofs for 40.0 (C-8) and 75.8 (C-9) show that CH3-14 is connected at position C-9; d H 3.20 (OC H 3-9) and d C The relevant evidence for 75.8 (C-9) shows that the methoxy group is also attached at the C-9 position; d H 1.65 (C H 3-15) and d C The correlation between 71.9 (C-2), 130.8 (C-4), and 134.1 (C-3) proves that CH3-15 is connected at position C-3; d H 5.21 (H-2) and d C The correlation at 172.7 (C-1'') indicates that the acetyl group is attached at the C-2 position. In the NOESY spectrum, d H 1.26 (C H 3-14) and d H 2.21 (H-8a) and 2.53 (H-5) are related. d H 5.36 (H-7) and d H 2.53(H-5) related, d H 1.76 (H-10) and dH 2.75 (H-6), 3.20 (OC) H 3-9) related, d H 1.89 (H-8b) and d H 2.75(H-6) related, prove H-5, H-7, C H 3-14 have the same orientation, H-6, OC H The orientations of H-9 and H-10 are opposite. The relative configuration at the H-2 position cannot be determined based on the NOESY spectrum. The relative configuration of this compound was ultimately determined by calculating the carbon NMR spectrum, comparing the two relative configurations. R , 5 S , 6 S , 7 S , 9 S , 10 S (7a) and 2 S , 5 S , 6 S , 7 S 9 S , 10 S (7b) Perform DP4+ carbon spectrum calculations. Measured data and 2 R , 5 S , 6 S , 7 S , 9 S , 10 S (7a) Linear correlation coefficient R of the calculated data of the configuration 2 The value is 0.9984, and the DP4+ probability is 100.00%, thus determining the relative configuration of compound 7 to be 2. R , 5 S , 6 S , 7 S , 9 S , 10 S The NOESY spectral analysis results were verified. The absolute configuration was determined by comparing the calculated ECD with the measured spectrum, confirming that the absolute configuration of compound 7 is 2. R , 5 S , 6 S , 7 S , 9 S 10 S In summary, based on Sci-finder search results, this compound is a novel compound that has not been previously reported, and it is named eupacadinolide M.

[0040] The structural identification data for compound 8 are as follows: Transparent oil (methanol) +36.7 ( c 0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 409.1867 [MH] - (calcd. 409.1857 for [C 21 H 29 O8] - The molecular formula of the compound is presumed to be C. 21 H 30 O8 has an unsaturation degree of 7. 1 Two sets of double bond hydrogen signals are given in the H-NMR (600 MHz, CD3OD) spectrum: d H 5.50 (1H, m, H-4), 6.34 (1H, s, H-13a), 5.50 (1H, m, H-13b); 2 hydroxymethyl hydrogen signals: d H 3.96 (1H,m, H-2), 5.39 (1H, d, J = 3.1 Hz, H-7); 3 sets of methylene hydrogen signals: d H 1.91 (1H, ddd, J = 2.0,10.8, 12.8 Hz, H-10), 2.46 (1H, t, J = 11.7, 11.7 Hz, H-5), 2.80 (1H, dd, J =2.7, 12.6 Hz, H-6); 2 sets of methylene hydrogen signals: d H 2.08 (1H, dt, J= 2.0, 2.0, 13.5 Hz, H-1a), 1.46 (1H, td, J = 4.2, 13.3, 13.4 Hz, H-1b), 2.12 (1H, dd, J = 2.9, 14.1 Hz, H-8a), 1.99 (1H, m, H-8b); 2 sets of methyl hydrogen signals: d H 1.26 (3H, s, C H 3-14), 1.75 (3H, t, C) H 3-15); This suggests that compound 8 is a juniperane-type sesquiterpene lactone. Additionally, a set of side-chain hydrogen signals is given: d H 6.90 (1H,t, J = 5.9, 5.9 Hz, H-3'), 4.40 (2H, d, J = 5.9 Hz, H-4'), 4.27 (2H, s, H-5'); and a set of methoxy hydrogen signals: d H 3.24 (3H, s, OC H 3-9). 13 C-NMR (150 MHz, CD3OD) yielded 21 carbon signals, including 15 carbon signals from the parent nucleus of juniperane-type sesquiterpene lactones: d C 32.5 (C-1), 68.8 (C-2), 137.3 (C-3), 128.0 (C-4), 37.4 (C-5), 47.2 (C-6), 72.5 (C-7), 39.8 (C-8), 76.1 (C-9), 41.2 (C-10), 141.9 (C-11), 173.2 (C-12), 126.2 (C-13), 21.7 (C-14), 21.3 (C-15). Additionally, a set of side-chain carbon signals is given: d C 167.2 (C-1'), 132.9 (C-2'), 146.1 (C-3'), 59.5 (C-4'), 56.9 (C-5') and a methoxy carbon signal: d C 49.6 (O C H3-9). In summary, this suggests that compound 8 is a juniperane-type sesquiterpene lactone compound with one methoxy group and one side chain substitution. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, it can be observed that... d H 2.08 (H-1a) and d C 37.4 (C-5) related, d H 5.50 (H-4) and d C 47.2 (C-6) and 68.8 (C-2) are relevant. d H 2.46 (H-5) and d C 137.3 (C-3) related, proves the existence of two six-membered rings through 5- and 10-bit concatenation; d H 6.34 (H-13a) and d C 47.2 (C-6) correlation, proving that the unsaturated lactone is attached at the C-6 position; d H 5.39 (H-7) and d C The correlation at 167.2 (C-1') proves that the sidechain is linked at position C-7. d H 1.26 (C H 3-14) and d C The relevant proofs for 39.8 (C-8) and 76.1 (C-9) show that CH3-14 is connected at position C-9; d H 3.24 (OC H 3-9) and d C 76.1 (C-9) provides evidence that the methoxy group is also attached at the C-9 position; d H 1.75 (C H 3-15) and d C Correlation between 68.8 (C-2), 128.0 (C-4), and 137.3 (C-3) indicates that CH3-15 is attached at the C-3 position. In the NOESY spectrum, d H 1.26 (C H 3-14) and d H 2.46 (H-5) related, d H 1.91 (H-10) and d H 2.80 (H-6), 3.24 (OC) H3-9) Related, prove H-5, C H 3-14 have the same orientation, H-6, OC H The orientations of H-9 and H-10 are opposite. The relative configurations of H-2 and H-7 cannot be determined based on the NOESY spectrum. The relative configuration of this compound was ultimately determined by calculating the carbon NMR spectrum, and the two relative configurations were compared. S , 5 S , 6 R , 7 R , 9 R , 10 R (8a) and 2 S , 5 S , 6 R , 7 S 9R , 10 R (8b) Perform DP4+ carbon spectrum calculations. Measured data and 2 S 5 S , 6 R , 7 S 9R , 10 R (8b) Linear correlation coefficient R of the calculated data of the configuration 2 The value is 0.9983, and the DP4+ probability is 100.00%, thus determining the relative configuration of compound 8 to be 2. S , 5 S , 6 R , 7 S 9R , 10 R The NOESY spectral analysis results were verified. The absolute configuration was determined by comparing the calculated ECD with the measured spectrum, confirming that the absolute configuration of compound 8 is 2. R , 5 R , 6 S , 7 R , 9S, 10S In summary, based on Sci-finder search results, this compound is a novel compound that has not been previously reported, and it is named eupacadinolide N.

[0041] Table 3. Attribution of 1H NMR and 1C NMR data for compounds 6-8

[0042] The structural identification data of compound 9 are as follows: White powder (methanol) +144.0 ( c 0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 395.1709 [MH] - (calcd. 395.1700 for [C 20 H 27 O8] - The molecular formula of the compound is presumed to be C. 20 H 28 O8 has an unsaturation degree of 7. 1 Two sets of double bond hydrogen signals are given in the H-NMR (600 MHz, CD3OD) spectrum: d H 5.34 (1H, q, J =1.6, 1.6, 1.6 Hz, H-4), 6.40 (1H, d, J = 1.2 Hz, H-13a), 5.50 (1H, s, H-13b); 2 hydroxymethyl hydrogen signals: d H 4.15 (1H, dd, J = 6.1, 10.7 Hz, H-2), 5.28 (1H, q, J = 3.1, 3.1, 3.0 Hz, H-7); 3 sets of methylene hydrogen signals: d H 1.57 (1H, ddd, J = 1.8, 10.4, 12.5 Hz, H-10), 2.56 (1H, t, J = 11.8, 11.8 Hz, H-5), 2.72 (1H, dd, J = 2.7, 12.7 Hz, H-6); 2 sets of methylene hydrogen signals: d H 2.41 (1H, ddd, J= 1.8, 6.1, 12.0 Hz, H-1a) , 1.37 (1H, td, J =10.3, 12.3, 12.3 Hz, H-1b), 2.12 (1H, dd, J = 3.0, 14.4 Hz, H-8a), 1.84 (1H,dd, J = 3.4, 14.4 Hz, H-8b); 2 sets of hydroxymethylene hydrogen signals: 4.40 (2H, d, J = 5.9 Hz, H-4'), 4.27 (2H, s, H-5'); 2 sets of methyl hydrogen signals: d H 1.28 (3H, s, C H 3-14), 1.71 (3H, m, C H 3-15) indicates that compound 9 is a juniperane-type sesquiterpene lactone. Furthermore, a set of side-chain hydrogen signals is also given: d H 6.90 (1H, t, J =5.9, 5.9 Hz, H-3'), 4.40 (2H, d, J = 5.9 Hz, H-4'), 4.27 (2H, s, H-5'). 13 C-NMR (150 MHz, CD3OD) yielded 20 carbon signals, including 15 carbon signals from the parent nucleus of juniperane-type sesquiterpene lactones: d C 33.5 (C-1), 72.0 (C-2), 139.9 (C-3), 126.7 (C-4), 37.6 (C-5), 47.4 (C-6), 72.2 (C-7), 45.8 (C-8), 71.1 (C-9), 50.5 (C-10), 140.2 (C-11), 171.0 (C-12), 127.8 (C-13), 23.5 (C-14), 19.6 (C-15). Additionally, a set of side-chain carbon signals is given: d C 167.1 (C-1'), 132.9 (C-2'), 146.2 (C-3'), 59.5 (C-4'), 56.9 (C-5'). In summary, this suggests that compound 9 is a juniperane-type sesquiterpene lactone compound with one side chain substitution. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, [the following can be observed]. d H 2.41 (H-1a) and d C 37.6 (C-5) and 139.9 (C-3) are relevant. d H 5.34 (H-4) and d C 72.0 (C-2) related, d H 5.28 (H-7) and d C 37.6 (C-5) related, d H 2.12 (H-9a) and d C 47.4 (C-6) related, proves the existence of two six-membered rings through 5- and 10-bit concatenation; d H 6.40 (H-13a) and d C 47.4 (C-6) correlation, proving that the unsaturated lactone is attached at the C-6 position; d H 5.28 (H-7) and d C The correlation at 167.1 (C-1') proves that the sidechain is linked at position C-7. d H 1.28 (C H 3-14) and d C The relevant proofs for 45.8 (C-8) and 71.1 (C-9) show that CH3-14 is connected at position C-9; d H 1.71 (C H 3-15) = with d C Correlation between 72.0 (C-2), 126.7 (C-4), and 139.9 (C-3) indicates that CH3-15 is attached at the C-3 position. In the NOESY spectrum, d H 1.28(C H 3-14) and d H 2.56 (H-5) and 2.12 (H-8a) are related. d H 1.57 (H-10) and d H The correlations of 2.72 (H-6), 4.15 (H-2), and 1.84 (H-8b) prove that H-5 and C... HPositions 3-14 have the same orientation, while H-6 and H-2 have opposite orientations. The relative configuration of position H-7 cannot be determined based on the NOESY spectrum. The relative configuration of this compound was ultimately determined by calculating the carbon NMR spectrum, comparing the two relative configurations. S , 5 S , 6 R , 7 R , 9 R , 10 R (9a) and 2 S , 5 S , 6 R , 7 S 9R , 10 R (9b) Perform DP4+ carbon spectrum calculations. Measured data and 2 S , 5 S , 6 R , 7 S 9R , 10 R (9b) Linear correlation coefficient R of the calculated data of the configuration 2 The value is 0.9985, and the DP4+ probability is 100.00%, thus determining the relative configuration of compound 9 to be 2. S , 5 S , 6 R , 7 S 9R , 10 R The NOESY spectral analysis results were verified. The absolute configuration was determined by comparing the calculated ECD with the measured spectrum, confirming that the absolute configuration of compound 9 was 2. S , 5 S , 6 R , 7 S , 9 R , 10 RIn summary, based on Sci-finder searches, this compound is a novel compound that has not been previously reported, and it has been named eupacadinolide O.

[0043] The structural identification data of compound 10 are as follows: Transparent oil (methanol) -146.0 ( c 0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 419.1678 [M+Na] + (calcd. 419.1676 for C 20 H 28 O8Na). The molecular formula of the compound is presumed to be C. 20 H 28 O8 has an unsaturation degree of 7. 1 Two sets of double bond hydrogen signals are given in the H-NMR (600 MHz, CD3OD) spectrum: d H 5.47 (1H, t, J =1.7, 1.7 Hz, H-4), 6.36 (1H, d, J = 1.4 Hz, H-13a), 5.54 (1H, s, H-13b); 2 hydroxymethyl hydrogen signals: d H 3.98 (1H, d, J = 3.96 Hz, H-2), 5.32 (1H, q, J = 3.1, 3.1, 3.0 Hz, H-7); 3 sets of methylene hydrogen signals: d H 1.71 (1H, ddd, J = 2.1, 10.6, 12.9 Hz, H-10), 2.39 (1H,t, J = 11.8, 11.8 Hz, H-5), 2.82 (1H, dd, J = 2.7, 12.6 Hz, H-6); 2 sets of methylene hydrogen signals: d H 2.19 (1H, dt, J = 2.0, 2.0, 13.4 Hz, H-1a), 1.50 (1H, td, J = 4.25, 13.2, 13.2Hz, H-1b), 2.12 (1H, dd, J= 3.0, 14.3 Hz, H-8a), 1.89 (1H, dd, J = 3.3, 14.4 Hz, H-8b); 2 sets of methyl hydrogen signals: d H 1.24 (3H, s, C H 3-14), 1.71 (3H, t, J = 1.8, 1.8Hz, C H 3-15). The above information suggests that compound 10 is a juniperane-type sesquiterpene lactone compound. In addition, a set of side-chain hydrogen signals is given: d H 6.89 (1H, t, J = 5.9, 5.9 Hz, H-3'), 4.39 (2H, d, J = 5.9 Hz, H-4'), 4.27 (2H,s, H-5'). 13 C-NMR (150 MHz, CD3OD) yielded 20 carbon signals, including 15 carbon signals from the parent nucleus of juniperane-type sesquiterpene lactones: d C 32.5 (C-1), 68.6 (C-2), 137.4 (C-3), 127.8 (C-4), 37.6 (C-5), 47.3 (C-6), 72.6 (C-7), 45.9 (C-8), 71.5 (C-9), 44.9 (C-10), 141.2 (C-11), 172.4 (C-12), 126.8 (C-13), 23.7 (C-14), 21.3 (C-15). Additionally, a set of side-chain carbon signals is given: d C 167.2 (C-1'), 132.9 (C-2'), 146.1 (C-3'), 59.5 (C-4'), 56.9 (C-5'). All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, [the following can be observed]... d H 2.19 (H-1a) and d C 37.6 (C-5) and 137.4 (C-3) are relevant. d H 5.47 (H-4) and d C 47.3 (C-6), 44.9 (C-10), and 68.6 (C-2) are relevant. d H5.32(H-7) and d C 37.6 (C-5) and 71.5 (C-9) are relevant. d H 2.12 (H-8a) and d C Correlation between 47.3 (C-6) and 44.9 (C-10) proves the existence of two six-membered rings, which are concatenated at positions 5 and 10. d H 6.36 (H-13a) and d C 47.3 (C-6) correlation, proving that the unsaturated lactone is attached at the C-6 position; d H 5.32 (H-7) and d C The correlation at 167.2 (C-1') proves that the sidechain is linked at position C-7. d H 1.24 (C H 3-14) and d C The correlation between 44.9 (C-10) and 71.5 (C-9) proves that CH3-14 is connected at position C-9; d H 1.75 (C H 3-15) and d C A correlation of 68.6 (C-2) indicates that CH3-15 is attached at the C-3 position. In the NOESY spectrum, d H 1.24 (C H 3-14) and d H 2.39(H-5) related, d H 1.71 (H-10) and d H 2.82 (H-6) correlation, prove H-5, C H Positions 3-14 have the same orientation, while H-6 and H-10 have opposite orientations. The relative configurations of positions H-2 and H-7 cannot be determined based on the NOESY spectrum. The relative configurations of this compound were ultimately determined by calculating the carbon NMR spectrum, and the two relative configurations were compared. S , 5 S , 6 R , 7 R , 9 R , 10 R (10a) and 2 S , 5 S , 6 R , 7 S 9R , 10 R (10b) Perform DP4+ carbon spectrum calculations. Measured data and 2 S , 5 S , 6 R , 7 S 9R , 10 R (10b) Linear correlation coefficient R of the calculated data of the configuration 2 The value is 0.9985, and the DP4+ probability is 99.99%, thus determining the relative configuration of compound 10 to be 2. S , 5 S , 6 R , 7 S 9R , 10 R The NOESY spectral analysis results were verified. The absolute configuration was determined by comparing the calculated ECD with the measured spectrum, confirming that the absolute configuration of compound 10 is 2. R , 5 R , 6 S , 7 R , 9 S , 10 S In summary, based on Sci-finder searches, this compound is a novel compound that has not been previously reported, and it has been named eupacadinolide P.

[0044] Table 4. Attribution of 1H NMR and 1C NMR data for compounds 9-10

[0045] The structural identification data of compound 11 are as follows: Transparent oil (methanol) +183.3 ( c0.10, MeOH). Quasi-molecular ion peaks were given by HR-ESI-MS. m / z 401.1573 [M+Na] + (calcd. 401.1571 for C 20 H 26 O7Na). The molecular formula of the compound is presumed to be C. 20 H 26 O7 has an unsaturation degree of 8. 1 Two sets of double bond hydrogen signals are given in the H-NMR (600 MHz, CD3OD) spectrum: d H 5.40 (1H, m, H-4), 6.36 (1H, s, H-13a), 5.65 (1H, s, H-13b); 2 hydroxymethyl hydrogen signals: d H 4.0 (1H,d, J = 3.0 Hz, H-2), 5.28 (1H, m, H-7); 2 sets of methylene hydrogen signals: d H 3.17 (1H, d, J = 11.4Hz, H-5), 2.87 (1H, d, J = 12.0 Hz, H-6); 2 sets of methylene hydrogen signals: d H 2.94 (1H, m, H-1a) ,2.87 (1H, d, J = 12.0 Hz, H-1b), 2.53 (1H, d, J = 16.4 Hz, H-8a), 2.16 (1H, d, J =16.4 Hz, H-8b); 2 sets of methyl hydrogen signals: d H 1.71 (3H, s, C H 3-14), 1.74 (3H, s, C H 3-15). The above information suggests that compound 11 is a juniperane-type sesquiterpene lactone compound. In addition, a set of side-chain hydrogen signals is given: d H 6.84(1H, t, J = 5.9, 5.9 Hz, H-3'), 4.37 (2H, d, J = 5.9 Hz, H-4'), 4.25 (2H, s, H-5'). 13C-NMR (150 MHz, CD3OD) yielded 20 carbon signals, including 15 carbon signals from the parent nucleus of juniperane-type sesquiterpene lactones: d C 36.1 (C-1), 70.9 (C-2), 136.8 (C-3), 128.0 (C-4), 37.3 (C-5), 44.4 (C-6), 71.8 (C-7), 38.2 (C-8), 124.1 (C-9), 126.1 (C-10), 142.3 (C-11), 172.4 (C-12), 125.5 (C-13), 18.7 (C-14), 21.2 (C-15). Additionally, a set of side-chain carbon signals is given: d C 167.7 (C-1'), 133.0 (C-2'), 145.8 (C-3'), 59.4 (C-4'), 56.9 (C-5'). In summary, this suggests that compound 11 is a juniperane-type sesquiterpene lactone compound with one side chain substitution. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, it can be observed that... d H 2.94 (H-1a) and d C 37.3 (C-5), 124.1 (C-9), and 136.8 (C-3) are relevant. d H 5.40 (H-4) and d C 70.9 (C-2) and 126.1 (C-10) are relevant. d H 5.28 (H-7) and d C 37.3 (C-5) and 124.1 (C-9) are relevant. d H 2.12 (H-8b) and d C Correlation between 44.4 (C-6) and 124.1 (C-9) proves the existence of two hexacyclic rings, which are concatenated at positions 5 and 10. d H 6.36 (H-13a) and d C 44.4 (C-6) correlation, proving that the unsaturated lactone is attached at the C-6 position; d H 1.71 (C H 3-14) and d CThe relevant proofs for 38.2 (C-8) and 124.1 (C-9) show that CH3-14 is connected at position C-9; d H 1.74(C H 3-15) and d C Correlation between 70.9 (C-2), 128.0 (C-4), and 136.8 (C-3) indicates that CH3-15 is attached at the C-3 position. In the NOESY spectrum, d H 2.16 (H-8b) and d H 3.17 (H-5) and 4.01 (H-2) are relevant. d H 2.16 (H-8a) and d H 2.87 (H-6) related, prove H-2, H-5, C H Positions 3-14 have the same orientation, while H-6 has the opposite orientation. The relative configuration of position H-7 cannot be determined based on the NOESY spectrum. The relative configuration of this compound was ultimately determined by calculating the carbon NMR spectrum, comparing the two relative configurations. S , 5 R 6 S , 7 R (11a) and 2 S , 5 S , 6 S , 7 R (11b) Perform DP4+ carbon spectrum calculations. Measured data and 2 S 5 R , 6 S , 7 R (11a) Linear correlation coefficient R of the calculated data of the configuration 2 The value is 0.9988, and the DP4+ probability is 98.03%, thus determining the relative configuration of compound 11 to be 2. S , 5 R , 6 S , 7 R The NOESY spectral analysis results were verified. The absolute configuration was determined by comparing the calculated ECD with the measured spectrum, confirming that the absolute configuration of compound 11 is 2. S , 5 R , 6 S , 7 R In summary, based on Sci-finder search results, this compound is a novel compound that has not been previously reported, and it has been named eupacadinolideQ.

[0046] The structural identification data of compound 12 are as follows: Yellow oily substance (methanol). -20.7 ( c 0.10, MeOH). HR-ESI-MS yielded quasi-molecular ion peaks. m / z 383.1468 [M+Na] + (calcd. 383.1465 for C 20 H 24 O6Na). The molecular formula of the compound is presumed to be C. 20 H 24 O6 has an unsaturation degree of 9. 1 The H-NMR (600 MHz, CD3OD) spectrum gives a set of hydrogen signals for the ABX spin-coupled system: d H 7.14(1H, d, J = 7.92 Hz, H-1), 7.03 (1H, dd, J = 8.0, 1.9 Hz, H-2), 6.79 (1H, m, H-4); 1 set of double bond hydrogen signals: d H 6.38 (1H, d, J = 1.3 Hz, H-13a), 4.98 (1H, d, J = 1.2 Hz, H-13b); 1 hydroxymethyl hydrogen signal: d H 5.54 (1H, ddd, J = 2.9, 5.6, 11.1 Hz, H-7); 2 sets of methylene hydrogen signals: d H 4.62 (1H, d, J = 5.6 Hz, H-6), 3.17 (1H, m, H-9); 1 group of methylene hydrogen signals: d H2.25(1H, m, H-8a), 1.67 (1H, dt, J = 3.5, 3.5, 13.0 Hz, H-8b); 2 sets of methyl hydrogen signals: d H 1.34(3H, d, J = 7.2 Hz, C H 3-14), 2.26 (3H, s, C H 3-15). The above information suggests that compound 12 is a juniperane-type sesquiterpene lactone compound. In addition, a set of side-chain hydrogen signals is given: d H 6.84 (1H, t, J = 5.9, 5.9 Hz, H-3'), 4.40 (2H, d, J = 2.2, 6.0 Hz, H-4'), 4.29 (2H, d, J = 1.4 Hz, H-5'). 13 C-NMR (150 MHz, CD3OD) yielded 20 carbon signals, including 15 carbon signals from the parent nucleus of juniperane-type sesquiterpene lactones: d C 129.0 (C-1), 129.2 (C-2), 137.0 (C-3), 131.5 (C-4), 136.3 (C-5), 44.4 (C-6), 70.4 (C-7), 32.1 (C-8), 32.6 (C-9), 139.8 (C-10), 144.2 (C-11), 170.8 (C-12), 129.9 (C-13), 24.5 (C-14), 20.9 (C-15). Additionally, a set of side-chain carbon signals is also given: d C 167.5 (C-1'), 132.9 (C-2'), 145.9 (C-3'), 59.5 (C-4'), 57.0 (C-5'). In summary, this suggests that compound 12 is a juniperane-type sesquiterpene lactone compound with one side chain substitution. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, it can be observed that... d H 7.14 (H-1) and d C 32.6 (C-9) and 136.3 (C-5) are relevant. d H 7.03 (H-2) and d CRelated to 131.5 (C-4) and 139.8 (C-10), d H 6.79 (H-4) and d C 44.4 (C-6), 129.0 (C-1), and 139.8 (C-10) are relevant. d H 1.67 (H-8b) and d C 44.4 (C-6) and 39.8 (C-10) are relevant. d H 3.17 (H-9) and d C The correlation at 70.4 (C-7) confirms the presence of a benzene ring and an aliphatic ring in the compound, fused at positions 5 and 10. d H 6.38 (H-13a) and d C 44.4 (C-6) correlation, proving that the unsaturated lactone is attached at the C-6 position; d H 1.34 (C H 3-14) and d C The correlation between 32.1 (C-8) and 139.8 (C-10) proves that CH3-14 is connected at position C-9; d H 2.26 (C H 3-15) and d C The correlation between 31.5 (C-4) and 137.0 (C-3) proves that CH3-15 is attached at the C-3 position. In the NOESY spectrum, d H 1.34 (C H 3-14) and d H 1.68 (H-8b) and 5.56 (H-7) are relevant. d H 4.62 (H-6) and d H 1.68 (H-8a) related, prove H-6, H-7, C H Compounds 3-14 have the same orientation, while H-10 has the opposite orientation, thus determining the relative configuration of compound 12 to be 6. R , 7 R , 9 R The absolute configuration of compound 12 was determined by comparing calculated ECD with measured spectra, and was found to be 6. R , 7 R , 9 R In summary, based on Sci-finder search results, this compound is a novel compound that has not been previously reported, and it is named eupacadinolide R.

[0047] The structural identification data of compound 13 are as follows: Transparent oil (methanol) -168.0 ( c 0.10, MeOH). HR-ESI-MS yielded quasi-molecular ion peaks. m / z 401.1574 [M+Na] + (calcd. 401.1571 for C 20 H 24 O6Na). The molecular formula of the compound is presumed to be C. 20 H 26 O7 has an unsaturation degree of 8. 1 The H-NMR (600 MHz, CD3OD) spectrum gives three sets of double bond hydrogen signals: d H 5.46 (1H, t, J =1.6, 1.6 Hz, H-4), 6.40 (1H, d, J = 1.2 Hz, H-13a), 5.55 (1H, s, H-13b), 4.76(1H, d, J = 1.8 Hz, H-14a), 4.73 (1H, d, J = 1.9 Hz, H-14b); 2 hydroxymethyl hydrogen signals: d H 4.02 (1H, d, J = 4.2 Hz, H-2), 5.27 (1H, q, J = 2.8 Hz, H-7); 3 sets of methylene hydrogen signals: d H 2.23 (1H, t, J = 11.4 Hz, H-10), 2.31 (1H, m, H-5), 2.96 (1H, dd, J = 2.3, 12.4Hz, H-6); 2 sets of methylene hydrogen signals: d H 2.12 (1H, dt,J = 2.0, 2.0, 13.6 Hz, H-1a), 1.74(1H, m, H-1b), 2.60 (1H, dd, J = 3.1, 14.1 Hz, H-8a), 2.52 (1H, dd, J = 2.5, 14.1Hz, H-8b); 1 group of methyl hydrogen signals: d H 1.77 (3H, t, J = 1.9 Hz, C H 3-15). The above information suggests that compound 13 is a juniperane-type sesquiterpene lactone compound. In addition, a set of side-chain hydrogen signals is given: d H 6.86 (1H, t, J = 5.9,5.9 Hz, H-3'), 4.37 (2H, dd, J = 5.9 Hz, H-4'), 4.25 (2H, s, H-5'). 13 C-NMR (150MHz, CD3OD) yielded 20 carbon signals, including 15 carbon signals from the parent nucleus of juniperane-type sesquiterpene lactones: d C 35.7 (C-1), 68.5 (C-2), 137.5 (C-3), 127.9 (C-4), 42.0 (C-5), 47.0 (C-6), 73.3 (C-7), 41.3 (C-8), 148.0 (C-9), 39.6 (C-10), 140.3 (C-11), 171.2 (C-12), 127.8 (C-13), 108.2 (C-14), 21.2 (C-15). Additionally, a set of side-chain carbon signals is given: d C 167.2 (C-1'), 132.9 (C-2'), 145.9 (C-3'), 59.4 (C-4'), 56.9 (C-5'). In summary, this suggests that compound 13 is a juniperane-type sesquiterpene lactone compound with one side chain substitution. All hydrogen and carbon data were assigned according to HSQC. In the HMBC spectrum, it can be observed that... d H 2.12 (H-1a) and d C 42.0 (C-5) and 37.5 (C-3) are relevant. d H 4.02 (H-2) and d C 39.6 (C-10), 127.8 (C-4) are the thresholds. d H 5.46 (H-4) and d C 39.6 (C-10), 47.0 (C-6), and 68.5 (C-2) are relevant. d H 5.27 (H-7) and d C Related to 42.0 (C-5) and 148.0 (C-9) d H 2.60 (H-8a) and d C The correlations of 39.6 (C-10), 47.0 (C-6), and 108.2 (C-14) prove the existence of two six-membered rings, which are concatenated at positions 5 and 10. d H 6.40 (H-13a) and d C 47.0 (C-6) correlation, proving that the unsaturated lactone is linked at the C-6 position; d H 1.77 (C H 3-15) and d C The correlations of 68.5 (C-2), 127.9 (C-4), and 137.5 (C-3) prove that CH3-15 is attached at the C-3 position. In the NOESY spectrum, d H 4.75 (H-14) and d H 2.60 (H-8a) and 2.12 (H-1a) are related. d H 2.52 (H-8b) and d H 2.94 (H-10) and 2.96 (H-6) are relevant. d H 1.74 (H-1b) and d H 2.31 (H-5) correlation proves that H-5, H-6, and H-10 have the same orientation. The relative configurations at positions H-2 and H-7 cannot be determined based on the NOESY spectrum. The relative configurations of this compound were ultimately determined by calculating the carbon NMR spectrum, and the two relative configurations were compared. S , 5 S , 6 R , 7 S , 10 R (13a) and 2 S , 5 S , 6 R , 7 R , 10 R (13b) Perform DP4+ carbon spectrum calculations. Measured data and 2 S , 5 S , 6 R , 7 R , 10 R (13b) Linear correlation coefficient R of the calculated data of the configuration 2 The value is 0.9989, and the DP4+ probability is 100.00%, thus determining the relative configuration of compound 13 to be 2. S , 5 S , 6 R , 7 R , 10 R The NOESY spectral analysis results were verified. The absolute configuration was determined by comparing the calculated ECD with the measured spectrum, confirming that the absolute configuration of compound 13 was 2. S , 5 S , 6 R , 7 R , 10 R In summary, based on Sci-finder search results, this compound is a novel compound that has not been previously reported, and it is named eupacadinolide S.

[0048] Table 5. Attribution of 1H NMR and 1C NMR data for compounds 11-13

[0049] The structural identification data of compound 14 are as follows: Colorless oily substance (methanol). -54.0 ( c 0.10, MeOH). HR-ESI-MS yielded quasi-molecular ion peaks. m / z 411.1783 [M+Na] + (calcd. 411.1784 for C 22 H 28 O6Na). The molecular formula of the compound is presumed to be C. 22 H 28 O6 has an unsaturation degree of 9. 1 The H-NMR (600 MHz, CDCl3) spectrum gives four olefin hydrogen signals: d H 5.07 (1H, t, J =7.6Hz, H-1), d H 5.21 (1H, dt, J =10.7, 1.3 Hz, H-5), d H 6.35 (1H, d, J =2.3 Hz, H-13a), d H 5.75 (1H, d, J =2.3 Hz, H-13b); 3 oxygen-hydrogen signals: d H 5.60 (1H, dd, J =11.7, 5.2 Hz, H-3), d H 5.26 (1H, dd, J =10.7, 2.2 Hz, H-6), d H 5.24 (1H, t, J =3.1 Hz, H-8); 2 sets of methylene hydrogen signals: d H 2.75 (1H, ddd, J =13.0, 8.5, 5.4 Hz, H-2a), d H 2.07 (1H, m, H-2b), d H 2.70(1H, dd, J =14.1, 3.4 Hz, H-9a), d H 2.37 (1H, dd, J =14.1, 3.1 Hz, H-9b); 1 methylene hydrogen signal: d H2.95 (1H, dt, J =3.1, 1.5 Hz, H-5) and 2 sets of methyl hydrogen signals: d H 1.89 (3H, s, C H 3-14) d H 1.80 (3H, m, C H 3-15) indicates that compound 14 is a gemmaeane-type sesquiterpene lactone. Additionally, a set of acetyl hydrogen signals is given: d H 2.07 (3H, s, C H 3-2'') and a set of acyl hydrogen signals: d H 6.83 (1H,tdt, J =7.2, 5.5, 1.9 Hz, H-3'), d H 1.80 (3H, m, C H 3-4'), d H 1.80 (3H, m, C H 3-5'). 13 The C-NMR (150 MHz, CDCl3) spectrum yielded 22 carbon signals, including 15 core carbon signals from gemmaane-type sesquiterpene lactones: d C 124.3 (C-1), 30.7 (C-2), 70.9 (C-3), 135.8 (C-4), 125.7 (C-5), 74.5 (C-6), 48.9 (C-7), 79.0 (C-8), 43.5 (C-9), 136.3 (C-10), 137.5 (C-11), 169.5 (C-12), 124.7 (C-13), 18.6 (C-14), 18.2 (C-15). In addition, one set of tegacyl carbon signals is also given: d C 167.0 (C-1'), 128.0 (C-2'), 139.2 (C-3'), 14.8 (C-4'), 12.2 (C-5') and one set of acetyl carbon signals: d C 170.4 (C-1''), 21.3 (C-2''). In summary, this suggests that compound 14 is a gemimaline-type sesquiterpene lactone substituted with teicoyl and acetyl groups. Its data were assigned according to HSQC. In the HMBC spectrum, it can be observed... d H 6.35 (H-13a) and d C The correlation between 48.9 (C-7) and 169.5 (C-12) proves the presence of an unsaturated lactone ring and its coupling at positions 6 and 7. d H 5.24 (H-8) and d C The correlation of 167.0 (C-1') proves that the tegacyl group is linked at the C-8 position; d H 1.89(C H 3-14) and d C The relevant proofs for 43.5 (C-9), 124.3 (C-1), and 136.3 (C-10) show that CH3-14 is connected at position C-10; d H 1.80 (C H 3-15) and d C The correlation of 70.9 (C-3), 125.7 (C-5), and 135.8 (C-4) proves that CH3-15 is connected at position C-4; d H 5.60 (H-3) and d C The correlation between 170.4 (C-1'') and the acetyl group indicates that the acetyl group is attached at the C-3 position. This can be observed in the NOESY spectrum. d H 5.60 (H-3) and d H 5.26 (H-6) related d H 5.26 (H-6) and d H 6.83 (H-3') correlation, thus proving that H-3, H-6 and the acetyl side chain have the same orientation, i.e., opposite to H-8; according to d H Coupling constant of 6.35 (H-13a) J =2.3 Hz is less than 3, indicating that the lactone ring is trans-fused, that is, H-6 and H-7 are reverse-coupled. Therefore, the relative configuration of the compound is inferred to be 3. R , 6 S , 7 R , 8 R The absolute configuration of compound 14 was determined by comparing calculated ECD with measured spectra, and was found to be 3. R , 6 S , 7 R , 8 R A Sci-finder search revealed that this compound was a previously unreported novel compound, and it was named 3. α -acetoxy-8 β -tigloyloxyhelianolide.

[0050] Table 6. Attribution of 1H NMR and 1C NMR data for compound 14

[0051] Example 2 (1) 1 kg of the whole herb of Wild Horse Chase was extracted three times by heating and refluxing with 95% ethanol (dosage: 10 L), and the crude extract was obtained by vacuum recovery. (2) The ethanol extract obtained in step (1) above is dissolved in an organic solvent (methanol), then mixed with macroporous adsorption resin and dried. It is then separated by D101 macroporous adsorption resin and eluted sequentially with mixed solvents of pure water-ethanol volume ratios of 100:0, 85:15, 70:30, 50:50, 30:70, and 5:95. (3) The pure water-ethanol fraction with a volume ratio of 85:15 to 30:70 obtained in step (2) above was separated by ODS chromatography and eluted by gradient with mixed solvents of methanol-water volume ratios of 2:8, 4:6, 6:4, 8:2, and 9:1. (4) The methanol-water fraction with a volume ratio of 4:6 to 8:2 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 28:72, to obtain compound 1 (t R = 25 min (yield 0.0027%); (5) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 30:70, to obtain compound 2 (t R = 35 min (yield 0.0006%); (6) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 23:78, to obtain compound 3 (t R = 35 min (yield 0.0012%); (7) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 25:75, to obtain compound 4 (t R = 34 min (yield 0.0015%); (8) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 33:67, to obtain compound 5 (t R = 19 min (yield 0.0011%); (9) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 26:74, to obtain compound 6 (t R = 43 min (yield 0.0015%); (10) The methanol-water fraction with a volume ratio of 4:6 ~ 8:2 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 27:73, to obtain compound 7 (t R = 32 min (yield 0.0021%); (11) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 40:60, to obtain compound 8 (t R = 36 min (yield 0.0003%); (12) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 26:74, to obtain compound 9 (t R = 20 min (yield 0.0004%); (13) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 25:75, to obtain compound 10 (t R = 29 min (yield 0.0008%); (14) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 23:77, to obtain compound 11 (t R = 22 min (yield 0.0015%); (15) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 34:66, to obtain compound 12 (t R = 32 min (yield 0.0011%); (16) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 25:75, to obtain compound 13 (t R = 24 min (yield 0.0015%); (17) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 37:63, to obtain compound 14 (t R = 33 min (yield 0.0002%).

[0052] The structural identification methods for new compounds 1-14 are described in Example 1.

[0053] Example 3 (1) 1.5 kg of the whole herb of Wild Horse Chase was extracted three times by heating and refluxing with 80% ethanol (dosage: 15 L), and the crude extract was obtained by vacuum recovery. (2) The ethanol extract obtained in step (1) above was dissolved in an organic solvent (methanol), then mixed with macroporous adsorption resin and dried. It was then separated by D101 macroporous adsorption resin and eluted sequentially with a mixed solvent of pure water and ethanol in volume ratios of 100:0, 80:20, 60:40, 40:60, 20:80, and 5:95. (3) The pure water-ethanol fraction with a volume ratio of 80:20 to 20:80 obtained in step (2) above was subjected to ODS chromatography and eluted with a gradient of mixed solvents of acetonitrile-water at ratios of 1:9, 3:7, 5:5, 7:3, and 8:2. (4) The methanol-water fraction with a volume ratio of 3:7 to 7:3 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 26:74, to obtain compound 1 (t R = 26 min (yield 0.0026%); (5) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 33:67, to obtain compound 2 (t R = 33 min (yield 0.0006%); (6) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 22:78, to obtain compound 3 (t R = 37 min (yield 0.0010%); (7) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 24:76, to obtain compound 4 (t R = 35 min (yield 0.0016%); (8) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 35:65, to obtain compound 5 (t R = 20 min (yield 0.0012%); (9) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 24:76, to obtain compound 6 (t R = 45 min (yield 0.0013%); (10) The methanol-water fraction with a volume ratio of 3:7 to 7:3 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 26:74, to obtain compound 7 (t R = 31 min (yield 0.0020%); (11) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 42:58, to obtain compound 8 (t R = 34 min (yield 0.0005%); (12) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 28:72, to obtain compound 9 (t R = 19 min (yield 0.0004%); (13) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 22:78, to obtain compound 10 (t R = 32 min (yield 0.0009%); (14) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 25:75, to obtain compound 11 (t R = 20 min (yield 0.0013%); (15) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 33:67, to obtain compound 12 (t R = 34 min (yield 0.0013%); (16) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 27:73, to obtain compound 13 (t R = 27 min (yield 0.0016%); (17) The methanol-water fraction with a volume ratio of 1:9 to 3:7 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 37:63, to obtain compound 14 (t R = 33 min (yield 0.0002%).

[0054] The structural identification methods for new compounds 1-14 are described in Example 1.

[0055] Example 4 (1) 1 kg of wild horse herb was extracted three times by heating and refluxing with 95% methanol (dosage: 10 L), and the crude extract was obtained by vacuum recovery of the extract. (2) The methanol extract obtained in step (1) above was dissolved in an organic solvent (methanol), then mixed with macroporous adsorption resin and dried. It was separated by D101 macroporous adsorption resin and eluted sequentially with mixed solvents of pure water-ethanol volume ratios of 100:0, 70:30, 50:50, 30:70, and 5:95. (3) The pure water-ethanol fraction with a volume ratio of 100:0 to 30:70 obtained in step (2) above was subjected to ODS chromatography and eluted with a gradient of mixed solvents with a volume ratio of acetonitrile-water of 1:9, 2:8, 4:6, 6:4, and 8:2. (4) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 25:75, to obtain compound 1 (t R = 29 min (yield 0.0024%); (5) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 34:66, to obtain compound 2 (t R = 30 min (yield 0.0005%).

[0056] (6) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 21:79, to obtain compound 3 (t R = 37 min (yield 0.0010%); (7) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 22:78, to obtain compound 4 (t R = 37 min (yield 0.0016%); (8) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 36:64, to obtain compound 5 (t R= 16 min (yield 0.0014%); (9) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 26:74, to obtain compound 6 (t R = 42 min (yield 0.0015%); (10) The methanol-water fraction with a volume ratio of 2:8 to 6:4 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 23:77, to obtain compound 7 (t R = 36 min (yield 0.0021%); (11) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 43:57, to obtain compound 8 (t R = 33 min (yield 0.0003%); (12) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 26:74, to obtain compound 9 (t R = 20 min (yield 0.0004%); (13) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 22:78, to obtain compound 10 (t R = 28 min (yield 0.0008%); (14) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 21:79, to obtain compound 11 (t R = 26 min (yield 0.0013%); (15) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 33:67, to obtain compound 12 (t R = 30 min (yield 0.0010%); (16) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 25:75, to obtain compound 13 (t R = 25 min (yield 0.0016%); (17) The methanol-water fraction with a volume ratio of 1:9 to 4:6 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 35:65, to obtain compound 14 (t R = 33 min (yield 0.0002%).

[0057] The structural identification methods for new compounds 1-14 are described in Example 1.

[0058] Example 5 (1) 1.5 kg of the whole herb of Wild Horse Chase was extracted three times by heating and refluxing with 80% methanol (dosage: 15 L), and the crude extract was obtained by vacuum recovery of the extract. (2) The methanol extract obtained in step (1) above was dissolved in an organic solvent (methanol), then mixed with macroporous adsorption resin and dried. It was then separated by D101 macroporous adsorption resin and eluted sequentially with mixed solvents of pure water-ethanol volume ratios of 95:5, 75:25, 50:50, 25:75, and 5:95. (3) The pure water-ethanol fraction with a volume ratio of 95:5 to 25:75 obtained in step (2) above was subjected to ODS chromatography and eluted with a gradient of mixed solvents with volume ratios of methanol-water of 3:7, 5:5, 7:3, 8:2, and 9:1. (4) The methanol-water fraction with a volume ratio of 3:7~8:2 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 28:72, to obtain compound 1 (t R = 25 min (yield 0.0027%); (5) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 33:67, to obtain compound 2 (t R = 33 min (yield 0.0005%); (6) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 22:78, to obtain compound 3 (t R = 35 min (yield 0.0011%); (7) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 23:77, to obtain compound 4 (t R = 35 min (yield 0.0013%); (8) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 33:67, to obtain compound 5 (t R = 19 min (yield 0.0011%); (9) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 25:75, to obtain compound 6 (t R = 41 min (yield 0.0015%); (10) The methanol-water fraction with a volume ratio of 3:7~8:2 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 28:72, to obtain compound 7 (t R = 29 min (yield 0.0020%); (11) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water volume ratio of 44:56, to obtain compound 8 (t R = 32 min (yield 0.0005%); (12) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 26:74, to obtain compound 9 (t R = 20 min (yield 0.0005%); (13) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 22:78, to obtain compound 10 (t R = 33 min (yield 0.0008%); (14) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 25:75, to obtain compound 11 (t R = 20 min (yield 0.0015%); (15) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 32:68, to obtain compound 12 (t R = 35 min (yield 0.0010%); (16) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 26:74, to obtain compound 13 (t R = 22 min (yield 0.0013%); (17) The methanol-water fraction with a volume ratio of 3:7 to 5:5 obtained in step (3) above was separated by HPLC-UV chromatography, detected at 210 nm, with a flow rate of 3 mL / min and a mobile phase of acetonitrile:water with a volume ratio of 35:65, to obtain compound 14 (t R = 35 min (yield 0.0003%).

[0059] The structural identification methods for new compounds 1-14 are shown in Example 1.

[0060] Example 6 Lung protective activity tests of the new compounds 1-14 prepared in Examples 1-5.

[0061] (1) Experimental principle: Lung tissue inflammation and damage are closely related to abnormal activation of macrophages and release of inflammatory factors. The functional status of normal alveolar type II epithelial cells in MLE-12 mice is a key indicator of lung health. This experiment evaluated the protective effect of the Yemazhui series of compounds on alveolar epithelial cells by culturing MLE-12 cells in vitro and detecting cell viability using the MTT assay, and further explored its lung protective effect.

[0062] (2) Experimental method: ① Cell Culture G-F12 cell culture medium was prepared using FBS medium as a base. Cells were cultured at approximately 2.5 × 10⁶ cells / day. 5 The cells were cultured at a density of 5% CO2 at 37 ℃. By the third day, the adherent cells occupied about 80%-90% of the bottom area of ​​the culture flask. The adherent cells were digested with trypsin and passaged to another culture flask.

[0063] ② MTT assay for the effect of compounds on cell viability MLE-12 cells cultured in the logarithmic growth phase were taken and their density adjusted to 2.5 × 10⁶ cells / year using G-F12 medium. 5 Cells were seeded at 100 μL / well in 96-well plates and cultured at 37 °C in a 5% CO2 incubator. After 12 h of adherent culture, the medium was replaced with fresh medium, and the drug was added simultaneously. Different doses of the compound were provided, along with a blank control. After drug addition, cells were cultured for another 48 h, then MTT solution (50 μL / well) was added to the cell culture medium. Cells were co-incubated with 2 mg / mL MTT at 37 °C for 4 h. The culture medium was then removed, and 100 μL of DMSO solution was added. The optical density (OD) value was measured. The average OD value of three wells for each sample was calculated, and the cell viability (CV%) was calculated using the average value according to the following formula.

[0064] Cell viability % = (Average OD value of sample group - Average OD value of blank group) / (Average OD value of blank control group - Average OD value of blank group) × 100% ③ Statistical analysis Experimental results included at least three data points expressed as mean ± SEM. Statistical analysis of the experimental data was performed using the GraphPadPrism 8.2.1 biostatistical software system. Statistical inference was conducted using the Independent Samples t-test and Single-factor ANOVA to assess the significance of differences between groups. A statistically significant difference between groups was considered to exist when the calculated probability value (P) met the condition of P < 0.05.

[0065] (3) Experimental results: The cytotoxicity of some compounds at different concentration gradients (5, 10, and 20 μM) was tested using the MTT assay, and the results are shown in Table 7. Subsequent cell damage protection experiments were conducted within the concentration range where the monomeric compounds showed no cytotoxic activity. Compounds 2-7 and 12-13 did not show cytotoxicity at test concentrations of 5-50 μM.

[0066] Table 7. Effects of single addition of the Wild Horse Chase series compounds on the survival rate (%) of MLE-12 cells (Mean±SEM)

[0067] The protective activity of some monomeric compounds isolated from *Pterocarya stenoptera* against lung epithelial cells was evaluated using an NNK-induced MLE-12 cell injury model. The results showed that compounds 2-3, 5-6, and 13 exhibited significant lung epithelial cell protective activity at test concentrations of 5-10 μM, and compound 12 at 10 μM. Among these, compounds 2 and 5-6 showed stronger reversal effects against NNK-induced lung epithelial cell injury at a test concentration of 1 μM than the same dose of the positive control drug, glycyrrhizin chalcone A (LicoA). The experimental results are shown in Table 8.

[0068] Table 8. Effects of Wild Horse Chase series compounds on NNK-induced MLE-12 cell survival (%) (Mean±SEM)

[0069] Note: P <0.05, P <0.01, P <0.001 compared to the NNK-induced group; ### P <0.001 compared to the control group.

Claims

1. A class of compounds having the following chemical structural formula or pharmaceutically acceptable salts thereof, characterized in that: 。 2. The method for preparing the compound according to claim 1, characterized in that: Includes the following steps: (1) Wild horse chasing ( Eupatorium lindleyanum DC. is extracted by heating and reflux with pure water, ethanol solution or methanol solution, and the extract is recovered to obtain crude extract; (2) The crude extract obtained in step (1) is separated by D101 macroporous adsorption resin and eluted with a water-ethanol mixed solvent in a gradient. The fractions with a water-ethanol volume ratio of 100:0 to 20:80 are collected to obtain the target fraction A. (3) The target fraction A obtained in step (2) above is separated by ODS column chromatography, and gradient elution is performed using methanol-water or acetonitrile-water mixed solvent as mobile phase. The fractions with a volume ratio of methanol-water or acetonitrile-water of 1:9 to 8:2 are collected to obtain the target fraction B. (4) The target fraction B obtained in step (3) above is further separated and prepared by preparative high performance liquid chromatography. Gradient elution is performed using methanol-water mixed solvent or acetonitrile-water mixed solvent as the mobile phase to obtain compounds 1-14.

3. The preparation method according to claim 2, characterized in that: In step (1), the extraction method is to heat and reflux for 2 to 5 times; the material-to-liquid ratio is 1:8 to 1:20 g / mL; the volume concentration of the ethanol solution is 70% to 95%; and the volume concentration of the methanol solution is 80% to 95%.

4. The preparation method according to claim 2, characterized in that: In step (2), the volume ratio of water to ethanol in the water-ethanol mixed solvent is 100:0~5:

95.

5. The preparation method according to claim 2, characterized in that: In step (3), the volume ratio of methanol to water in the methanol-water mixed solvent is 1:9~9:1; the volume ratio of acetonitrile to water in the acetonitrile-water mixed solvent is 1:9~8:

2.

6. The preparation method according to claim 2, characterized in that: In step (4), the volume ratio of methanol to water in the methanol-water mixed solvent is 2:8~6:4; the volume ratio of acetonitrile to water in the acetonitrile-water mixed solvent is 1:9~6:

4.

7. The preparation method according to claim 2, characterized in that: In step (4), the detection wavelength is 210 nm and the flow rate is 3 mL / min.

8. The preparation method according to claim 2, characterized in that: In step (2), the crude extract is dissolved in an organic solvent, mixed with macroporous adsorption resin and dried, and then separated by D101 macroporous adsorption resin.

9. A pharmaceutical composition, characterized in that: The pharmaceutical composition comprises the compound of claim 1 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier thereof.

10. The use of the compound of claim 1 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition of claim 9 in the preparation of a medicament for the prevention or treatment of lung diseases.

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