A sesquiterpene compound, a preparation method and a use as acetylcholinesterase inhibitor

Abstract

The application discloses a sesquiterpene compound, a preparation method and an application as an acetylcholinesterase inhibitor. A novel compound physochlatone A is separated, molecular docking is performed on the compound and acetylcholinesterase, the binding activity of the two is simulated, the binding energy is-9.6 kcal / mol, the blood-brain barrier permeability prediction result shows that the compound can pass through the blood-brain barrier, and the acetylcholinesterase inhibition activity evaluation result shows that the compound has obvious acetylcholinesterase inhibition activity. It is known to those skilled in the art that a significant decrease in acetylcholine level is one of important causes of various neurodegenerative diseases, and inhibiting acetylcholinesterase activity can inhibit the decrease in acetylcholine level, thereby playing an anti-neurodegenerative disease role. Therefore, the physochlatone A has a prospect of being developed into a drug for resisting neurodegenerative diseases (such as Alzheimer's disease).

Description

Technical Field

[0001] This invention belongs to the field of medicinal chemistry and relates to the preparation and use of new compounds, specifically to a sesquiterpene compound, its preparation method, and its use as an acetylcholinesterase inhibitor. Background Technology

[0002] Acetylcholinesterase (AChE) is a key hydrolytic enzyme found in the synaptic cleft of cholinergic nerves. Its core function is to catalyze the hydrolysis of the neurotransmitter acetylcholine (ACh), thereby terminating nerve signal transmission. A significant decrease in acetylcholine levels is one of the causes of various neurodegenerative diseases, especially Alzheimer's disease (AD). Based on this "cholinergic hypothesis," inhibiting AChE activity, reducing ACh hydrolysis, and thus increasing the ACh concentration in the synaptic cleft has become one of the important treatment strategies for improving AD. However, existing clinically used AChE inhibitors are often accompanied by some side effects. For example, galantamine has serious adverse reactions such as bradycardia, heart failure, esophageal perforation, gastrointestinal bleeding, and thrombocytopenia [Liang Zhi, Fan Linlin, Li Hongliang, et al. Current status and new progress of drug treatment for Alzheimer's disease [J]. Southwest National Defense Medicine, 2018, 28(01):85-87]. Therefore, finding novel, highly effective, and low-toxicity AChE inhibitors, especially by mining molecules with unique structures and mechanisms of action from natural products, is an important direction and urgent need in the field of drug development.

[0003] Huashan ginseng is the dried root of *Physochlaina infundibularis* Kuang, a plant belonging to the genus *Physochlaina* in the Solanaceae family. It was first recorded in *Compendium of Materia Medica Supplement* and included in the *Chinese Pharmacopoeia* in 2010. Huashan ginseng possesses the effects of warming the lungs and resolving phlegm, relieving asthma and cough, and calming the nerves and relieving convulsions. Modern pharmacological studies have found that it has analgesic, sedative, and anti-inflammatory pharmacological activities. Phytochemical studies have shown that Huashan ginseng contains various alkaloids, flavonoids, coumarins, and other active ingredients.

[0004] This invention isolates a novel compound from Panax ginseng, a compound that has been discovered for the first time. This invention also reveals that this compound possesses significant biological activity, hence this invention. Summary of the Invention

[0005] The first objective of this invention is to provide a novel sesquiterpene compound, the second objective is to provide a method for preparing the compound, and the third objective is to provide the use of the compound.

[0006] The above-mentioned objective of this invention is achieved through the following technical solution:

[0007] A sesquiterpene compound or its pharmaceutically acceptable salt or solvate with the following structural formula:

[0008]

[0009] A method for preparing the above-mentioned sesquiterpene compound includes the following steps:

[0010] S1. Pulverize the Chinese ginseng, extract it with an ethanol aqueous solution, filter it, and concentrate the filtrate under reduced pressure to obtain a crude extract. Resuspend the crude extract in water to make a suspension, extract it with an equal volume of ethyl acetate, concentrate the organic phase under reduced pressure to obtain the ethyl acetate extract, and extract the remaining aqueous phase with an equal volume of n-butanol. Concentrate the organic phase under reduced pressure to obtain the n-butanol extract.

[0011] S2. The ethyl acetate extract was loaded onto a macroporous adsorption resin column for chromatography. A gradient elution with different proportions of methanol-water mixed solvent was used to obtain five fractions, Fr.A to Fr.E. Fraction Fr.D was used for subsequent separation. The gradient elution ratios, volumes, and order of the macroporous adsorption resin column chromatography were as follows: 0:100 elution for 1 column volume → 30:70 elution for 3 column volumes → 50:50 elution for 3 column volumes → 80:20 elution for 4 column volumes → 100:0 elution for 3 column volumes.

[0012] S3. The fraction Fr.D was loaded onto a normal-phase silica gel column for chromatography, eluted with a gradient of petroleum ether-ethyl acetate mixed solvent, and detected and combined by TLC to obtain 7 fractions Fr.D.1 to D.7. Fraction Fr.D.2 was used for subsequent separation. The gradient elution ratio, volume and order of the normal-phase silica gel column chromatography were as follows: 10:1 elution for 2 column volumes → 8:1 elution for 3 column volumes → 5:1 elution for 3 column volumes → 3:1 elution for 4 column volumes → 2:1 elution for 4 column volumes → 1:1 elution for 3 column volumes → methanol elution for 2 column volumes.

[0013] S4. Split Fr.D.2 was loaded onto a normal-phase silica gel column for chromatography, eluted with a gradient of petroleum ether-ethyl acetate mixed solvent, and detected and combined by TLC to obtain 6 fractions Fr.D.2.1 to D.2.6. Split Fr.D.2.1 was used for subsequent separation. The gradient elution ratio, volume and order of the normal-phase silica gel column chromatography were as follows: 10:1 elution for 2 column volumes → 7:1 elution for 3 column volumes → 5:1 elution for 4 column volumes → 3:1 elution for 4 column volumes → 1:1 elution for 3 column volumes → methanol elution for 2 column volumes.

[0014] S5. Load Fr.D.2.1 onto an ODS reversed-phase column and elute with a gradient of methanol-water mixed solvents in different proportions to obtain 7 fractions Fr.D.2.1.1 to Fr.D.2.1.7. Fraction Fr.D2.1.2 is used for subsequent separation. The gradient elution ratio, volume, and order of the ODS reversed-phase column chromatography are as follows: 0:100 elution for 1 column volume → 30:70 elution for 3 column volumes → 40:60 elution for 3 column volumes → 50:50 elution for 3 column volumes → 60:40 elution for 3 column volumes → 80:20 elution for 3 column volumes → 100:0 elution for 2 column volumes.

[0015] S6. Load Fr.D2.1.2 onto a Sephadex LH-20 column, elute isocratically with methanol, and detect and combine by TLC to obtain three fractions Fr.D2.1.2.1 to Fr.D2.1.2.3, of which the fraction Fr.D2.1.2.2 is used for subsequent separation;

[0016] S7. The fraction of Fr.D2.1.2.2 was subjected to preparative HPLC chromatography with methanol-water isocratic elution at a volume ratio of 51:49. The fraction at the peak of Huashan Panax lactone A was collected, concentrated and dried to obtain the final product.

[0017] Preferably, the ethanol aqueous solution in step S1 is an ethanol aqueous solution with an ethanol volume fraction of 80%.

[0018] Preferably, the extraction temperature in step S1 is 90°C.

[0019] Preferably, the macroporous adsorption resin is HP-20.

[0020] The above-mentioned sesquiterpene compounds or their pharmaceutically acceptable salts or solvates are used in the preparation of acetylcholinesterase inhibitor drugs.

[0021] The above-mentioned sesquiterpene compounds or their pharmaceutically acceptable salts or solvates are used in the preparation of medicaments for the treatment of neurodegenerative diseases.

[0022] Preferably, the neurodegenerative disease is Alzheimer's disease.

[0023] Preferably, the drug uses the sesquiterpene compound or its pharmaceutically acceptable salt or solvate as the active ingredient and is formulated into a pharmaceutically acceptable dosage form with pharmaceutically acceptable excipients.

[0024] More preferably, the excipient is a solid, liquid, or semi-solid, and the dosage form is a tablet, capsule, injection, or pill.

[0025] Beneficial effects:

[0026] This invention isolated a novel compound, physochlatone A. Molecular docking was performed between this compound and acetylcholinesterase (AChE, PDB: 1EVE) to simulate their binding activity; the binding energy was -9.6 kcal / mol. Blood-brain barrier permeability prediction results showed that the compound could cross the blood-brain barrier. Acetylcholinesterase inhibitory activity evaluation results indicated that the compound possessed significant acetylcholinesterase inhibitory activity. Those skilled in the art know that a significant decrease in acetylcholine levels is one of the important causes of various neurodegenerative diseases. Inhibiting acetylcholinesterase activity can inhibit the decrease in acetylcholine levels, thereby exerting an anti-neurodegenerative effect. Therefore, the compound physochlatone A discovered in this invention has the potential to be developed into a drug for treating neurodegenerative diseases (such as Alzheimer's disease). Attached Figure Description

[0027] Figure 1 The HR-ESI-MS spectrum of compound physochlatone A;

[0028] Figure 2 For the compound physochlatone A 1 1H NMR spectrum (500MHz, CD3OD);

[0029] Figure 3 For the compound physochlatone A 13 C NMR spectrum (500MHz, CD3OD);

[0030] Figure 4 DEPT135 of compound physochlatone A and 13 C NMR comparison (500MHz, CD3OD);

[0031] Figure 5 The HSQC spectrum (500 MHz, CD3OD) of compound physochlatone A is shown.

[0032] Figure 6 The HMBC spectrum (500 MHz, CD3OD) of compound physochlatone A is shown.

[0033] Figure 7 For the compound physochlatone A 1 H- 1 H COSY spectrum (500MHz, CD3OD);

[0034] Figure 8The NOESY spectrum (500 MHz, CD3OD) of compound physochlatone A;

[0035] Figure 9 HMBC of compound physochlatone A and 1 H- 1 H COSY related graphs;

[0036] Figure 10 ECD image of compound physochlatone A;

[0037] Figure 11 DP4+ calculation analysis for compound physochlatone A;

[0038] Figure 12 This is a schematic diagram of the predicted interaction between compound physochlatone A and 1EVE (the yellow dashed lines represent hydrogen bonds);

[0039] Figure 13 A schematic diagram of the predicted BBB permeability of compound physochlatone A (arrows indicate the BBB region). Detailed Implementation

[0040] The following describes the substantive content of the present invention in detail with reference to embodiments, but this does not limit the scope of protection of the present invention.

[0041] Huashan ginseng was collected in Pingdingshan City, Henan Province (September 2021), and was later identified by Professor Li Jun of the School of Traditional Chinese Medicine, Henan University of Traditional Chinese Medicine as the dried root of Physochlaina infundibularis Kuang, a plant of the Solanaceae family.

[0042] Example 1: Isolation and identification of sesquiterpene compounds (named physochlatone A, Huashan ginseng lactone A)

[0043] 1. Extraction and separation

[0044] After pulverizing 60 kg of Huashan ginseng, it was extracted twice with an 80% ethanol aqueous solution at 90°C for 2 hours each time. The extracts were combined, filtered, and the filtrate was concentrated under reduced pressure to obtain a crude extract. The crude extract was added to distilled water to form a suspension and extracted three times with an equal volume of ethyl acetate. The organic phase was concentrated under reduced pressure at 40°C using a rotary evaporator to obtain the ethyl acetate extract. The remaining aqueous phase was extracted three times with an equal volume of n-butanol. The organic phase was concentrated under reduced pressure at 40°C using a rotary evaporator to obtain the n-butanol extract.

[0045] The ethyl acetate extract was loaded onto a macroporous adsorption resin column (HP-20). A gradient elution with different ratios of methanol-water mixed solvents was used to obtain five fractions, Fr.A to Fr.E. Fraction Fr.D was used for subsequent separation. The gradient elution ratios, volumes, and order of the macroporous adsorption resin column chromatography were as follows: 0:100 elution for 1 column volume → 30:70 elution for 3 column volumes → 50:50 elution for 3 column volumes → 80:20 elution for 4 column volumes → 100:0 elution for 3 column volumes.

[0046] The fraction Fr.D was loaded onto a normal-phase silica gel column and eluted with a gradient of petroleum ether-ethyl acetate. The fractions were then analyzed and combined by TLC to obtain seven fractions, Fr.D.1 to D.7. Fraction Fr.D.2 was used for subsequent separation. The gradient elution ratios, volumes, and order of the normal-phase silica gel column chromatography were as follows: 10:1 for 2 column volumes → 8:1 for 3 column volumes → 5:1 for 3 column volumes → 3:1 for 4 column volumes → 2:1 for 4 column volumes → 1:1 for 3 column volumes → methanol for 2 column volumes.

[0047] Fraction Fr.D.2 was loaded onto a normal-phase silica gel column and eluted with a gradient of petroleum ether-ethyl acetate. The fractions were then analyzed and combined by TLC to obtain six fractions, Fr.D.2.1 to D.2.6. Fraction Fr.D.2.1 was used for subsequent separation. The gradient elution ratio, volume, and order of the normal-phase silica gel column chromatography were as follows: 10:1 elution for 2 column volumes → 7:1 elution for 3 column volumes → 5:1 elution for 4 column volumes → 3:1 elution for 4 column volumes → 1:1 elution for 3 column volumes → methanol elution for 2 column volumes.

[0048] Fr.D.2.1 was loaded onto an ODS reversed-phase column and eluted with a gradient of methanol-water mixed solvents in different proportions to obtain seven fractions, Fr.D.2.1.1 to Fr.D.2.1.7. Fraction Fr.D2.1.2 was used for subsequent separation. The gradient elution ratios, volumes, and order of the ODS reversed-phase column chromatography were as follows: 0:100 elution for 1 column volume → 30:70 elution for 3 column volumes → 40:60 elution for 3 column volumes → 50:50 elution for 3 column volumes → 60:40 elution for 3 column volumes → 80:20 elution for 3 column volumes → 100:0 elution for 2 column volumes.

[0049] Fr.D2.1.2 was loaded onto a Sephadex LH-20 column and eluted isocratically with methanol. The fractions were then analyzed by TLC and combined to obtain three fractions: Fr.D2.1.2.1 to Fr.D2.1.2.3. The fraction Fr.D2.1.2.2 was used for subsequent separation.

[0050] The fraction of Fr.D2.1.2.2 was subjected to preparative HPLC chromatography with isocratic elution of methanol-water at a volume ratio of 51:49. The fraction at the peak of Huashan ginsenoside A was collected, concentrated and dried to obtain Huashan ginsenoside A.

[0051] 2. Structural identification of Physochlatone A

[0052] Colorless oily substance, HR-ESI-MSm / z = 249.1121 [M+H] + (The calculated value is 249.1121) (e.g.) Figure 1 As shown), the molecular formula is C 14 H 16 O4 (combined) 1 H and 13 According to C-NMR data, the degree of unsaturation of this compound is 7. 1 H-NMR spectrum ( Figure 2 ), δ H 5.16 (1H, d, J = 4.5 Hz, H⁻³) is the proton signal on the hydroxymethyl group; δ H 4.84(2H,s,H2-12) represents the signal of two protons on a set of terminal double bonds; δ H 3.22 (1H,dd,J=18.0,4.5Hz,H-4a), 2.93 (1H,ddt,J=19.3,8.9,1.8Hz,H-9a), 2.79 (1H,d,J=18.0Hz,H-4b), 2.49 (1H,ddd,J=19.3,10.4,2.0Hz,H-9b), 1.89 (1H,d,J=14.0Hz,H-7b), and 2.22 (1H,ddt,J=13.3,5.6,1.2Hz,H-7a) represent the proton signals of three groups of methylene groups; one methine proton signal δ H 3.08 (1H, m, H-8); 2 sets of methyl proton signals δ H 1.91 (3H, s, H3-13) and 1.80 (3H, s, H3-14). 13 C-NMR spectrum ( Figure 3 ) displays 14 carbon signals, combined with DEPT ( Figure 4 ) and HSQC ( Figure 5 The spectrum assigned 14 carbon signals to 2 carbonyl carbon signals δ. C 175.3 (C-5) and 164.9 (C-2); δ signals for 4 olefin carbons C 152.8 (C-10), 146.6 (C-11), 125.8 (C-1) and 110.8 (C-12); one oxygen-linked quaternary carbon signal δ C 87.5 (C-6); δC 80.9 (C-3) is the methylene carbon signal bonded to an oxygen atom; the three methylene carbon signals δ C 42.1 (C-7), 38.1 (C-4) and 34.7 (C-9); 1 methylene carbon signal δ C 44.3 (C-8); 2 methyl carbon signals δ C 21.1(C-13) and 13.4(C-14).

[0053] 1 H- 1 H COSY( Figure 7 The spectrum showed a correlation between H2-7 / H-8 / H2-9; and HMBC ( Figure 6 In the spectrum, H2-9 is correlated with C-10 and C-6, and H2-7 is correlated with C-10 and C-6, confirming the presence of a five-membered ring (A ring) composed of C-6—C-7—C-8—C-9—C-10 in the compound. H3-13 is correlated with C-12, C-11, and C-8, and H2-12 is correlated with C-8, indicating the presence of an isopropenyl group in the compound, attached at the C-8 position. 1 H- 1 The 1H COSY spectrum showed a correlation between H-3 and H2-4, and the HMBC spectrum showed correlations between H-3 / C-5, C-6, and H2-4 / C-5, confirming the presence of a five-membered lactone ring (C ring). Further correlation of H-3 / C-7 confirmed that rings A and C are connected via C-6. The HMBC spectrum showed correlations between H3-14 and C-1, C-2, and C-10, and between H2-4 and C-2. Combined with the compound's unsaturation level, this confirmed the presence of an unsaturated lactone ring (B ring), with rings A and B connected via C-6 and C-10, and rings B and C connected via C-3 and C-6. This confirmed the planar structure of the compound. Figure 9 As shown.

[0054] The absolute configuration of the compound was determined by comparing the measured and calculated ECD curves. The ECD curve of the compound was calculated at the b3lyp / 6-31g+(d,p) level using a pcm solvation model with methanol as the dielectric constant. Figure 10 As shown, the calculated ECD curves of (3R,6R,8R)-1 and (3R,6R,8S)-1 fit well with the measured ECD curves of compound physochlatone A. Therefore, the absolute configurations at the C-3 and C-6 positions of compound physochlatone A are determined to be 3R,6R. To determine the absolute configuration at the C-8 position, the calculated ECD curves of (3R,6R,8R)-1 and (3R,6R,8S)-1 were calculated. 13CNMR chemical shifts show (3R, 6R, 8R)-1 ( 13 C data: R 2 =0.9989) has a better match with the experimental data than (3R,6R,8S)-1( 13 C data: R 2 =0.9986). Furthermore, according to DP4+ probability analysis, the probability of (3R,6R,8R)-1 is higher than that of (3R,6R,8S)-1 (99.62 vs 0.38%). Figure 11 In summary, the absolute configuration of compound physochlatone A is determined to be 3R,6R,8R.

[0055] The C-H signal assignments of compound physochlatone A are shown in Table 1.

[0056] Table 1 Compounds 1 H (500MHz) and 13 C(125MHz) NMR data (CD3OD)

[0057]

[0058] Example 2: Molecular docking study of Physochlatone A and acetylcholinesterase

[0059] Ligands: The target compound was downloaded from the PubChem database (https: / / pubchem.ncbi.nlm.nih.gov / ). The molecular structure was optimized for energy minimization (MM2 force field in Chemdraw 3D). After optimization, the file was saved in mol2 format and then converted to pdbqt format using AutoDockTools as the ligand file for molecular docking. Acceptor: The crystal structure file of the target protein acetylcholinesterase (PDB ID: 1EVE) was downloaded from the RCSB protein database (https: / / www.rcsb.org / ). Small molecules and water molecules were removed using UCSF Chimera, and after hydrogenation, it was saved in pdb format and converted to pdbqt format using AutoDockTools as the acceptor file for molecular docking. Docking Grid Box Setup: The center coordinates and size of the docking grid box were set using AutoDockTools software to ensure that the grid covers the protein's active site. The center coordinates (x, y, z) and size (length, width, height) of the grid box were recorded for subsequent docking calculations. Molecular docking calculations: Molecular docking calculations were performed using AutoDockVina. The acceptor (pdbqt format) and ligand (pdbqt format) files were imported into Vina, along with the recorded Grid Box information. The docking parameters were correctly set to run the docking calculations. After docking, the results were visualized using PyMOL to analyze the interaction modes and binding free energies between the ligand and acceptor.

[0060] The binding mode of compound Physochlatone A to 1EVE protein is as follows: Figure 12 As shown, the two compounds form a suitable spatial complementarity. The carbonyl oxygen atom of compound Physochlatone A forms hydrogen bonds with the hydrogen atoms of GLY-118 and GLY-119, respectively; the oxygen atom on the ester bond forms a hydrogen bond with the hydroxyl hydrogen atom of TYR-121. The binding energy of Physochlatone A to 1 EVE is -9.6 kcal / mol.

[0061] Example 3: Predictive Study of Blood-Brain Barrier Permeability Using Physochlatone A

[0062] By inputting the compound's structural formula into the Swiss Target Prediction tool (http: / / swisstargetprediction.ch / ), one can predict the target sites that the compound may bind to. By combining this with known information on targets related to the blood-brain barrier, one can infer whether the compound may cross the blood-brain barrier.

[0063] like Figure 13 As shown in the figure, compound physochlatone A is located in the BBB region, which is permeable to the blood-brain barrier, indicating that compound physochlatone A has good blood-brain barrier permeability.

[0064] Example 4: Acetylcholinesterase inhibitory activity of compound physochlatone A

[0065] Ellman Method: 140 μL of PBS buffer (0.1 M pH = 8.0), 20 μL of sample solutions (1 μM, 10 μM, and 100 μM), and 15 μL of AChE solution (0.28 U / ml, pH = 8.0, dissolved and diluted in PBS) were added sequentially to a 96-well plate and incubated (4℃) for 20 min. Then, 10 μL each of DTNB (0.075 mol / L) and ATCI (0.01 mol / L) solutions were added to the 96-well plate. After incubation (37℃) for 20 min, the absorbance was measured using a microplate reader (405 nm). The inhibition rate was calculated using the following formula.

[0066] Inhibition rate (%) = [(blank group - positive group) - (experimental group - baseline reaction group)] / (blank group - positive group) × 100%.

[0067] In the blank group, 20 μL of sample solution was replaced with 20 μL of PBS (pH=8.0); in the positive group, 20 μL of sample solution was replaced with 20 μL of galantamine (0.125 mg / ml); and in the reaction background group, 15 μL of L-ChE (0.28 U / ml, dissolved and diluted in PBS at pH=8.0) was replaced with 15 μL of PBS (pH=8.0).

[0068] The inhibition rate results are shown in Table 2.

[0069] Table 2 Results of anti-acetylcholinesterase ( n=3)

[0070]

[0071] Experimental results show that compound physochlatoneA has acetylcholinesterase inhibitory activity and has the potential to be developed into an acetylcholinesterase inhibitor.

[0072] In summary, this invention isolated a novel compound, physochlatone A. Molecular docking of this compound with acetylcholinesterase (AChE, PDB: 1EVE) simulated their binding activity, revealing a binding energy of -9.6 kcal / mol. Blood-brain barrier permeability prediction showed that the compound could cross the blood-brain barrier. Acetylcholinesterase inhibitory activity evaluation indicated that the compound exhibited significant acetylcholinesterase inhibitory activity. Those skilled in the art know that a significant decrease in acetylcholine levels is a major cause of various neurodegenerative diseases. Inhibiting acetylcholinesterase activity can suppress the decrease in acetylcholine levels, thereby exerting an anti-neurodegenerative effect. Therefore, the compound physochlatone A discovered in this invention shows promise for development into a drug for neurodegenerative diseases (such as Alzheimer's disease).

[0073] The purpose of the above embodiments is to specifically illustrate the substantive content of the present invention, but those skilled in the art should know that the scope of protection of the present invention should not be limited to the specific embodiments.

Claims

1. A sesquiterpene compound with the following structural formula or a pharmaceutically acceptable salt thereof: 。 2. A method for preparing the sesquiterpene compound according to claim 1, characterized in that, Includes the following steps: S1. Pulverize the Chinese ginseng, extract it with an ethanol aqueous solution, filter it, and concentrate the filtrate under reduced pressure to obtain a crude extract. Resuspend the crude extract in water to make a suspension, extract it with an equal volume of ethyl acetate, concentrate the organic phase under reduced pressure to obtain the ethyl acetate extract, and extract the remaining aqueous phase with an equal volume of n-butanol. Concentrate the organic phase under reduced pressure to obtain the n-butanol extract. S2. The ethyl acetate extract was loaded onto a macroporous adsorption resin column for chromatography. A gradient elution with different proportions of methanol-water mixed solvent was used to obtain five fractions, Fr. A to Fr. E. Fraction Fr. D was used for subsequent separation. The gradient elution ratios, volumes, and order of the macroporous adsorption resin column chromatography were as follows: 0:100 elution for 1 column volume → 30:70 elution for 3 column volumes → 50:50 elution for 3 column volumes → 80:20 elution for 4 column volumes → 100:0 elution for 3 column volumes. S3. The fraction Fr. D was loaded onto a normal-phase silica gel column for chromatography, eluted with a gradient of petroleum ether-ethyl acetate mixed solvent, and detected and combined by TLC to obtain 7 fractions Fr. D.1 to D.

7. Fraction Fr. D.2 was used for subsequent separation. The gradient elution ratio, volume and order of the normal-phase silica gel column chromatography were as follows: 10:1 elution for 2 column volumes → 8:1 elution for 3 column volumes → 5:1 elution for 3 column volumes → 3:1 elution for 4 column volumes → 2:1 elution for 4 column volumes → 1:1 elution for 3 column volumes → methanol elution for 2 column volumes. S4. Split Fr. D.2 was loaded onto a normal-phase silica gel column for chromatography, eluted with a gradient of petroleum ether-ethyl acetate mixed solvent, and detected and combined by TLC to obtain 6 fractions Fr. D.2.1 to D.2.

6. Split Fr. D.2.1 was used for subsequent separation. The gradient elution ratio, volume and order of the normal-phase silica gel column chromatography were as follows: 10:1 elution for 2 column volumes → 7:1 elution for 3 column volumes → 5:1 elution for 4 column volumes → 3:1 elution for 4 column volumes → 1:1 elution for 3 column volumes → methanol elution for 2 column volumes. S5. Load Fr. D.2.1 onto an ODS reversed-phase column for elution with a gradient of methanol-water mixed solvents in different proportions to obtain 7 fractions Fr. D.2.1.1 to Fr. D.2.1.

7. Fraction Fr. D2.1.2 is used for subsequent separation. The gradient elution ratio, volume, and order of the ODS reversed-phase column chromatography are as follows: 0:100 elution for 1 column volume → 30:70 elution for 3 column volumes → 40:60 elution for 3 column volumes → 50:50 elution for 3 column volumes → 60:40 elution for 3 column volumes → 80:20 elution for 3 column volumes → 100:0 elution for 2 column volumes. S6. Load Fr. D2.1.2 onto a Sephadex LH-20 column, elute isocratically with methanol, and detect and combine by TLC to obtain three fractions Fr. D2.1.2.1 to Fr. D2.1.2.3, of which the fraction Fr. D2.1.2.2 is used for subsequent separation; S7. The fraction of Fr. D2.1.2.2 was subjected to preparative HPLC chromatography with isocratic elution of methanol-water at a volume ratio of 51:

49. The fractions corresponding to the elution peaks were collected, concentrated and dried to obtain the final product.

3. The preparation method according to claim 2, characterized in that: In step S1, the ethanol-water solution is an ethanol-water solution with an ethanol volume fraction of 80%.

4. The preparation method according to claim 2, characterized in that: The extraction temperature in step S1 is 90℃.

5. The preparation method according to claim 2, characterized in that: The macroporous adsorption resin is model HP-20.

6. Use of the sesquiterpene compound of claim 1 or a pharmaceutically acceptable salt thereof in the preparation of an acetylcholinesterase inhibitor medicament.

7. Use of the sesquiterpene compound of claim 1 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating neurodegenerative diseases.

8. The use according to claim 7, characterized in that: The neurodegenerative disease mentioned is Alzheimer's disease.

9. The use according to claim 7 or 8, characterized in that: The drug uses the sesquiterpene compound or its pharmaceutically acceptable salt as the active ingredient and is formulated into a pharmaceutically acceptable dosage form with pharmaceutically acceptable excipients.

10. The use according to claim 9, characterized in that: The excipients are solid, liquid, or semi-solid, and the dosage form is tablet, capsule, injection, or pill.

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