A woodfordioid triterpene CNA, preparation method and application thereof

Abstract

The application discloses a woodfordioid toxic alkane type sesquiterpene CNA, a preparation method and application thereof, and belongs to the field of traditional Chinese medicine pharmacy. The woodfordioid toxic alkane type sesquiterpene CNA compound of formula I is obtained by cold soaking, silica gel and gel column chromatography from Artocarpus styracifolius. The CNA compound has a new cis-hydrogenated indene core structure, contains nine continuous chiral centers and a highly crowded polycyclic skeleton feature, and has good biological activity. Cell experiments show that the woodfordioid toxic alkane type sesquiterpene CNA has signal transmission to endogenous and exogenous system GABA B receptor and can be used for preparing drugs for treating or preventing nervous system related diseases, and opens up a new direction for developing a new GABA B receptor antagonist or modulator and exploring a GABA receptor related treatment target and developing a nervous system disease drug.

Description

Technical Field

[0001] This invention relates to a tetrandrine-type sesquiterpene CNA, its preparation method, and its application, belonging to the field of traditional Chinese medicine pharmaceuticals. Background Technology

[0002] Neurological disorders are diseases affecting the central or peripheral nervous system, including the brain and spinal cord. These diseases can be caused by genetics, degenerative processes, trauma, infection, or other factors. Neurological disorders are a leading cause of disability and death worldwide, encompassing a wide variety of conditions, including common ones like depression and anxiety, epilepsy, schizophrenia, Alzheimer's disease, Parkinson's disease, stroke, and multiple sclerosis (MS), as well as less common ones like amyotrophic lateral sclerosis (ALS) and Huntington's disease. With an aging population, particularly in developed countries and some developing countries, the incidence of neurodegenerative diseases (such as Alzheimer's and Parkinson's) is rising significantly. The development of drugs for neurological diseases is an important direction in the global biomedical field, but it also faces significant challenges. Due to the complexity of neurological diseases, individual differences, and the presence of the blood-brain barrier (BBB), drug development has a high failure rate. Current treatments for major neurodegenerative diseases, such as Alzheimer's and Parkinson's, primarily include acetylcholinesterase inhibitors, such as donepezil and galantamine, which enhance cholinergic transmission and improve cognitive function. NDA receptor antagonists, such as memantine, can reduce glutamate toxicity. Treatments for Parkinson's disease (PD) mainly include levodopa (L-DOPA), dopamine agonists such as pramipexole and ropinirole, and monoamine oxidase B inhibitors (MAO-B inhibitors) such as selegiline. Treatments for epilepsy include those that regulate sodium channels, calcium channels, and gamma-aminobutyric acid (GABA). Drugs for treating depression mainly consist of selective serotonin reuptake inhibitors (SSRIs) such as sertraline. Benzodiazepines and 5-HT1A receptor agonists are commonly used to treat anxiety disorders. Treatment for schizophrenia primarily involves dopamine D2 receptor antagonists, such as clozapine; other new drugs targeting glutamate and brain network function are under development.

[0003] GABA B The receptor (γ-aminobutyric acid type B receptor) is a metabolic G protein-coupled receptor (GPCR) responsible for mediating inhibitory neurotransmission. As an important member of the GABA receptor family, its main ligand is γ-aminobutyric acid (GABA), the primary inhibitory neurotransmitter in the brain. B Their widespread distribution in the brain and involvement in multiple signaling pathways suggest a close association with various mental and neurological disorders, which often involve GABA. B Alterations in receptor function, including overactivation or functional decline. GABA BGABA receptors are widely distributed in the central nervous system and play a key role in regulating neural activity. B Receptors have become an important drug target in the development of drugs related to nervous system diseases. (This is in contrast to GABA.) B Neurological disorders related to receptors include epilepsy, depression and anxiety, chronic pain, addiction and dependence, hypotension, muscle spasms, and Alzheimer's disease. Therefore, the development of novel GABA receptors is crucial. B Receptor antagonists or modulators have opened up new avenues for exploring therapeutic targets related to GABA receptors and developing drugs for neurological diseases. Summary of the Invention

[0004] In order to more effectively treat neurological diseases, the purpose of this invention is to provide a drug and its preparation method that can target GABA. B Receptor signal transduction has an inhibitory effect; development of novel GABA receptors is needed. B Receptor antagonists or modulators. Among them, the tetrandrine-type sesquiterpene CNA provided by this invention has the structure shown in Formula I:

[0005]

[0006] A method for preparing a tetrandrine-type sesquiterpene CNA includes the following steps:

[0007] (1) Take dried horse chestnut flowers, crush them, and extract them by cold soaking in a mixture of ethanol and water to obtain an extract. Then, distill the extract under reduced pressure and concentrate it to obtain an extract paste.

[0008] (2) Distribute the extract into water, first extract with petroleum ether, and then extract the water-soluble part with ethyl acetate to obtain ethyl acetate phase extract.

[0009] (3) The ethyl acetate phase extract was separated by normal silica gel chromatography. First, the fractions F1, F2, F3, F4 and F5 were obtained by gradient elution with petroleum ether-acetone as the mobile phase. Then, the fractions F6, F7 and F8 were obtained by gradient elution with dichloromethane-methanol as the mobile phase. All fractions were concentrated by rotation and weighed in sequence to obtain a total of 8 polar fractions F1 to F8.

[0010] (4) F5 was subjected to normal silica gel column chromatography and gradient elution was performed using dichloromethane-methanol as the mobile phase to obtain 9 fractions F5-1 to F5-9.

[0011] (5) Pass the F5-5 segment through normal silica gel column chromatography and use chloroform-methanol as the mobile phase for gradient elution to obtain 4 fractions F5-5-1 to F5-5-4;

[0012] (6) The F5-5-2 segment was separated by a gel column, eluted with chloroform-methanol to remove impurities, and recrystallized in acetone to obtain the sesquiterpene CNA substance of the tetrandrine type.

[0013] Preferably, in step (1), the material-to-liquid ratio of *Corydalis yanhusuo* flowers is 0.44 kg / L, the volume ratio of ethanol to water is 95:5, the extraction time is 24 h, and the cold soaking extraction is performed 3 times.

[0014] Preferably, in step (2), petroleum ether extraction is used to remove impurities with low polarity.

[0015] Preferably, in step (3), the volume ratio of petroleum ether to acetone is 50:1 to 1:1, and the volume ratio of dichloromethane to methanol is 10:1 to 1:1, wherein F5 is obtained by elution with petroleum ether to acetone at a ratio of 6:1.

[0016] Preferably, the volume ratio of dichloromethane to methanol in step (4) is 70:1 to 3:1, wherein F5-5 is obtained by elution with dichloromethane to methanol at a ratio of 8:1.

[0017] Preferably, the volume ratio of chloroform to methanol in step (5) is 8:1 to 2:1, wherein F5-5-2 is obtained by elution with chloroform-methanol at a ratio of 8:1.

[0018] Preferably, the volume ratio of chloroform to methanol in step (6) is 10:1 to 3:1.

[0019] In an embodiment of the present invention, the tetrandrine-type sesquiterpene CNA is extracted and separated from the flowers of *Corydalis yanhusuo*.

[0020] The drugs described in this invention include tetrandrine-type sesquiterpene CNA compounds.

[0021] Preferably, the drug includes tablets, capsules, pills, injections, sustained-release formulations, and controlled-release formulations, but is not limited thereto.

[0022] Preferably, the drug is a combination drug formed by combining a tetrandrine-type sesquiterpene CNA with at least one pharmaceutically acceptable carrier.

[0023] Preferably, the carrier includes alumina, polylactic acid-polyvinyl alcohol copolymer, polyvinyl alcohol, chitosan, liposomes, sodium alginate, etc.

[0024] Preferably, the combination drug is a drug composed of a tetrandrine-type sesquiterpene CNA and a pharmaceutically acceptable salt.

[0025] Preferred, pharmaceutically acceptable salts include: citric acid, maleic acid, fumaric acid, hydrochloric acid, sulfuric acid, phosphoric acid, lithium, sodium, potassium, calcium, magnesium, and lysine.

[0026] The tetrandrine-type sesquiterpene CNA compound described in this invention exhibits systemic inhibition of GABA by both endogenous and exogenous sources. B Receptor signal transduction.

[0027] The present invention relates to the application of the tetrandrine-type sesquiterpene CNA compounds and tetrandrine-type sesquiterpene CNA drugs in the preparation of drugs for the treatment or prevention of nervous system diseases, including epilepsy, depression, anxiety, schizophrenia, and neurodegenerative diseases.

[0028] The beneficial effects of this invention are:

[0029] (1) This invention provides a novel sesquiterpene CNA of the tetrandrine type, which has a unique cis-hydroindene core structure containing nine consecutive chiral centers and has a highly crowded polycyclic skeleton feature.

[0030] (2) The tetrandrine-type sesquiterpene CAN provided by this invention is the first naturally discovered natural GABA. B Receptor antagonists can be used to prepare drugs for the treatment and prevention of nervous system diseases, and have potential therapeutic significance for epilepsy, depression, anxiety, as well as schizophrenia and neurodegenerative diseases.

[0031] (3) The extraction process of this invention is simple, and the confirmation and cell experiment verification of the CNA structure of the tetrandrine-type sesquiterpene CNA provide a basis for the development of novel GABA. B Receptor antagonists or modulators have opened new avenues for exploring therapeutic targets related to GABA receptors and developing drugs for neurological diseases. Attached Figure Description

[0032] Figure 1 This is a flowchart illustrating the preparation method of the tetrandrine-type sesquiterpene CNA of the present invention.

[0033] Figure 2 The sesquiterpene CNA of the tetrandrine type of the present invention 1 HNMR image.

[0034] Figure 3 The sesquiterpene CNA of the tetrandrine type of the present invention 13 C NMR spectrum.

[0035] Figure 4 This is a single-crystal X-ray diffraction pattern of the sesquiterpene CNA of the present invention.

[0036] Figure 5 The diagram shows the structure of the sesquiterpene CNA of the tetrandrine type of the present invention.

[0037] Figure 6 For the compound CNA to target endogenous GABA BThe graph shows the inhibitory activity of receptor activation, with the vertical axis representing GABA in the endogenous system. B Receptor signaling Ca 2+ Conduction, with the horizontal axis representing the concentration of compound CNA in μM.

[0038] Figure 7 The compound CNA reacts with exogenous GABA upon GABA stimulation. B Inhibitory activity plot of receptor activation, with the vertical axis representing GABA in the endogenous system. B Receptor signaling Ca 2+ Conduction, with the horizontal axis representing the concentration of compound CNA in μM, where the concentration of the GABA agonist is 5 μM.

[0039] Figure 8 The compound CNA reacts with exogenous GABA under Baclofen stimulation. B Inhibitory activity plot of receptor activation, with the vertical axis representing GABA in the endogenous system. B Receptor signaling Ca 2+ Conduction, with the horizontal axis representing the dose of compound CNA in μM, where the concentration of Baclofen agonist is 1 μM. Detailed Implementation

[0040] The technical solution of the present invention will be further described below with reference to the accompanying drawings and specific embodiments. However, the following embodiments are merely simple examples of the present invention and do not represent or limit the scope of protection of the present invention. The scope of protection of the present invention is determined by the claims.

[0041] Example 1

[0042] A method for preparing a tetrandrine-type sesquiterpene CNA, such as... Figure 1 As shown, the steps include the following:

[0043] (1) Take 11 kg of dried horse chestnut flowers, crush them, and dissolve them in a mixed solvent of ethanol:water volume ratio of 95:5 at a material-to-liquid ratio of 0.44 kg / L. Extract the mixture three times at room temperature, combine the extracts, and then distill and concentrate them under reduced pressure to remove the organic solvent to obtain the extract.

[0044] (2) Distribute the extract into an aqueous solution, first extract it three times with petroleum ether to remove small polar components and impurities, and then extract the water-soluble part after extraction with ethyl acetate three times to obtain a total ethyl acetate phase extract of 500.0g.

[0045] (3) The ethyl acetate extract was separated by normal silica gel column chromatography. First, it was eluted with petroleum ether-acetone at a volume ratio of 50:1 to 1:1. Specifically, elution with petroleum ether-acetone at a ratio of 50:1 yielded F1, elution with petroleum ether-acetone at a ratio of 30:1 yielded F2, elution with petroleum ether-acetone at a ratio of 15:1 yielded F3, elution with petroleum ether-ethyl acetate at a ratio of 10:1 yielded F4, and elution with petroleum ether-acetone at a ratio of 6:1 yielded F5. Then, it was eluted with dichloromethane-methanol at a ratio of 10:1 yielded F6, elution with dichloromethane-methanol at a ratio of 5:1 yielded F7, and elution with dichloromethane-methanol at a ratio of 1:1 yielded F8. All the eluted samples were collected, and then concentrated by rotation and weighed in sequence to obtain a total of 8 polar fractions.

[0046] (4) The 52.8g fraction was passed through normal silica gel column chromatography and eluted with dichloromethane-methanol at a volume ratio of 70:1 to 3:1. Specifically, dichloromethane-methanol at a ratio of 60:1 yielded F5-1, dichloromethane-methanol at a ratio of 35:1 yielded F51-2, dichloromethane-methanol at a ratio of 20:1 yielded F5-3, dichloromethane-methanol at a ratio of 15:1 yielded F5-4, dichloromethane-methanol at a ratio of 10:1 yielded F5-5, dichloromethane-methanol at a ratio of 7:1 yielded F5-6, dichloromethane-methanol at a ratio of 5:1 yielded F5-7, dichloromethane-methanol at a ratio of 4:1 yielded F5-8, and dichloromethane-methanol at a ratio of 3:1 yielded F5-9, resulting in a total of 9 polar fractions.

[0047] (5) The 3.0 g fraction F5-5 was passed through normal silica gel column chromatography and eluted with chloroform-methanol at a volume ratio of 8:1 to 2:1. The chloroform-methanol elution at 8:1 yielded F5-5-1, the chloroform-methanol elution at 6:1 yielded F5-5-2, the chloroform-methanol elution at 4:1 yielded F5-5-3, and the chloroform-methanol elution at 2:1 yielded F5-5-4, resulting in four mixtures.

[0048] (6) The 1.2g fraction was separated into F5-5-2 by gel column separation, and then separated and purified by chloroform-methanol at a volume ratio of 10:1 to 3:1. The fraction was recrystallized in acetone and dried to obtain 7.0mg of the tetrandrine-type sesquiterpene CAN compound, which was in the form of colorless needle crystals.

[0049] Example 2

[0050] Nuclear magnetic resonance analysis was performed on the tetrandrine-type sesquiterpene CNA compounds. 1 H NMR and 13 C NMR such as Figure 2 and Figure 3 As shown, where, 1 H NMR (600MHz in methanol-d4) and 13The data and signal assignments for C NMR (150MHz in methanol-d4) are shown in Table 1 below.

[0051] Table 1. NMR data and signal assignments of tetrandrine-type sesquiterpene CNAs

[0052]

[0053]

[0054] The tetrandrine-type sesquiterpene CNA compounds isolated and extracted in Example 1 were analyzed by mass spectrometry (MS) and high-resolution mass spectrometry (HR-ESI-MS). The molecular ion peak [M+Na] of the tetrandrine-type sesquiterpene CNA compounds was determined. + m / z: 335.1111, calculated value 335.1101. Based on the analysis of NMR spectral data, the molecular formula of the sesquiterpene CNA compound of the tetrandrine type was determined to be C2. 15 H 20 O7. X-ray single-crystal diffraction analysis was performed on the tetrandrine-type sesquiterpene CNA compounds to determine their stereoconfiguration, such as... Figure 4 As shown.

[0055] Based on comprehensive structural analysis including NMR spectroscopy, mass spectrometry (MS), high-resolution mass spectrometry (HR-ESI-MS), and X-ray single-crystal diffraction analysis, the crystal structure of the tetrandrine-type sesquiterpene CNA isolated and extracted in Example 1 was further determined. Figure 5 As shown.

[0056] Example of effect

[0057] sesquiterpenoids, specifically tetrandrine-type CAN, inhibit GABA. B Receptor activity verification includes the following specific steps:

[0058] 1. Primary mouse cerebellar cell (CGNs) culture

[0059] One-week-old Balb / c mice were provided by Kunming Medical University. After euthanasia, their cerebellar tissue was dissected, and the tissue was gently dissected using a fire-polished Pasteur pipette. After being filtered through a cell filter, the tissue was seeded into culture dishes pre-coated with poly-L-ornithine (MAKLIN). Cell culture was carried out in DMEM medium supplemented with 10% Hepes buffer and 10% penicillin-dextrin antibiotics, followed by the addition of 10% heat-inactivated fetal bovine serum. After 5 days of culture, the cell density reached 90%.

[0060] 2. HEK293T cell culture

[0061] (1) Cell screening model: HepG2.2.15 cells were purchased from Guangzhou Saiku Biotechnology, passaged and cultured in our laboratory, and stored in liquid nitrogen and -80℃ freezer.

[0062] (2) Culture medium: Add 10% antibiotics and 10% Hepes buffer to 500 mL of DMEM medium.

[0063] (3) Cell resuscitation: Before the experiment, prepare complete culture medium in a clean bench and store it at 4°C for later use. The complete culture medium consists of 90% culture medium + 10% FBS. Shake it up and down to mix it thoroughly and heat it in a 37°C water bath. Take 5 mL of the thawed complete culture medium and store it in a T25 culture flask for later use. At this time, take out one frozen HEK293T cell from the liquid nitrogen tank and quickly place it in a 37°C water bath. Shake it gently and continuously until it is completely thawed. Then, seed the cell suspension into a T25 culture flask containing 5 mL of complete culture medium. The cells will adhere to the wall and grow to full size in about 48 hours.

[0064] (4) Cell passage: When the cell growth density reaches 90%, remove the cells from the incubator, discard the original culture medium, add 2 mL of pre-warmed PBS each time, wash 3 times, then add 1 mL of trypsin, gently shake to ensure all cells are in full contact, aspirate the trypsin, place the cells in the incubator for 3 min of digestion, and examine under a microscope. When significant changes in cell morphology are observed, add 2 mL of complete culture medium to stop digestion, pipette the cells, transfer them to 5 mL of ep tube, centrifuge at 1000 rpm for 3 min, discard the supernatant, add 1 mL of culture medium and pipette evenly, inoculate each culture dish with 300 μL of single cell suspension, add 5 mL of culture medium, pipette evenly, and place in the incubator.

[0065] (5) Cell cryopreservation: Remove cells in good growth condition (F3 / F4) from the incubator, discard the original culture medium, add 2 mL of fetal bovine serum PBS, wash 3 times, then add 1 mL of trypsin to digest the cells, place in an incubator for 3 min of digestion, examine under a microscope, add 2 mL of complete culture medium to stop digestion, pipette the cells, transfer to an Eppendorf tube, centrifuge at 1000 rpm for 3 min, discard the supernatant, add 1 mL of serum-free cryopreservation solution, and label the cryopreservation tube with the time, cell passage number, cryopreservation solution, cell name, and operator. Then perform gradient cooling cryopreservation: 4℃ for 30 min → -20℃ for 2 h → -80℃ overnight, and place in liquid nitrogen the next day.

[0066] 3. Plasmid transfection

[0067] HEK293T cells were cultured in DMEM medium containing 10% fetal bovine serum (FBS) and seeded in 6-well plates. When the cell density reached 70%-90%, the medium was replaced with a serum-free medium without antibiotics. Plasmids GABBR1 and GABBR2 were co-transfected into the cells using Lip3000 transfection reagent. After 6 hours, the medium was replaced with complete medium and cultured for another 24 hours.

[0068] 4. Fluo-4AM fluorescent probe for detecting GABA in endogenous and exogenous cells B Changes in downstream calcium flow signals of receptors

[0069] (1) Drug preparation: The tetrandrine-type sesquiterpene CAN compound was dissolved in DMSO to prepare a 50 mM solution. When administering the drug, it was serially diluted to 50, 25, 12.5, and 6.25 μM, with three replicates for each concentration.

[0070] (2) Cell seeding: Select CGNs cells in good growth condition, with a cell density of 1×10⁶. 5 Inoculate 100 μL of each probe per well into a 96-well plate and incubate for 24 h. Discard the original culture medium and incubate with the prepared 2 μM Fluo-4AM fluorescent probe for 1 h.

[0071] (3) Detection of calcium signal using Fluo-4AM fluorescent probe: After culturing for 24 h, the original culture medium was discarded, and the cells were incubated with the prepared 2 μM Fluo-4AM fluorescent probe for 1 h. The dye was removed, and the cells were washed twice with PBS. The pre-prepared CAN drug solution was added to each well and incubated for 1 h. Exogenous cells were stimulated with the agonists GABA (5 μM) and Baclofen (1 μM) for 1 h, respectively, and then the fluorescence intensity data were measured using an inverted fluorescence microscope.

[0072] Fluorescence intensity data were obtained using ImageJ quantization analysis, and graph plotting and statistical analysis were performed using GraphPad Prism software version 5.0 from GraphPad Software Inc. IC 50 The value is calculated using nonlinear regression and a three-parameter logistic equation.

[0073] The results of the endogenous primary mouse cerebellum cell experiment were as follows: Figure 6 The results show that the compound CNA can inhibit GABA in the endogenous system. B It involves receptor signal transduction and exhibits a certain dose-dependent effect; in exogenous HEK293T cell experiments, the compound CNA can inhibit GABA in the exogenous system. B Receptor signal transduction, such as Figure 7As shown, in exogenous HEK293T cells, the compound CNA exhibits a certain dose-dependent effect upon stimulation by GABA agonists; for example... Figure 8 As shown, in exogenous HEK293T cells, compound CNA exhibits a dose-dependent effect upon stimulation with baclofen agonists, and the IC50 values ​​of compound CNA under stimulation with GABA and baclofen agonists were calculated. 50 The values ​​are shown in Table 2 below.

[0074] Table 2 IC50 of compound CNA under stimulation by two agonists 50 value

[0075]

[0076] According to Table 2, the IC50 values ​​of the positive control drug CGP54626 under stimulation by two agonists are... 50 Compared to the values, compound CNA showed greater susceptibility to GABA under stimulation by both agonists. B The inhibitory activity of the receptor is lower than that of the positive control drug, but from IC50... 50 In terms of values, compound CAN significantly inhibited exogenous GABA under stimulation by both agonists. B Receptor activation.

[0077] In summary, the above experiments demonstrate that the tetrandrine-type sesquiterpene CNA prepared in this invention effectively inhibits GABA in both endogenous and exogenous systems. B The activation of receptors has a significant inhibitory effect, affecting GABA. B Receptor signal transduction, therefore, can be used to prepare drugs for the prevention or treatment of nervous system diseases.

[0078] Finally, it should be noted that the above preferred embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail through the above preferred embodiments, those skilled in the art should understand that various changes can be made to it in form and detail without departing from the scope defined by the claims of the present invention.

Claims

1. A CNA of the lndicine-type sesquiterpene, characterized in that, The structure of the woodfordioid triterpene CNA is shown as formula I:

2. The lignanocembranoid sesquiterpene CNA according to claim 1, characterized by, The woodfordioid triterpene CNA is extracted and separated from the leaves of Coriaria sinica.

3. The process for the preparation of the timber CNA of the toxane sesquiterpene according to claim 1 or 2, characterized by, The method comprises the following steps: (1) The dried leaves of Coriaria sinica are crushed and extracted by cold soaking with an ethanol aqueous solution, and the extracted solution is concentrated by distillation under reduced pressure to obtain an extract; (2) The extract is distributed in an aqueous solution, and is extracted with petroleum ether first, and then with ethyl acetate to obtain an ethyl acetate phase extract; (3) The ethyl acetate phase extract is separated by normal silica gel column chromatography, and is gradient eluted with petroleum ether-acetone as the mobile phase, and 5 fractions F1, F2, F3, F4 and F5 are collected; then the fractions are gradient eluted with dichloromethane-methanol as the mobile phase, and 3 fractions F6, F7 and F8 are collected, and all the fractions are successively concentrated and weighed, and 8 polar segment fractions are obtained; (4) The fraction F5 is gradient eluted with dichloromethane-methanol as the mobile phase by normal silica gel column chromatography, and 9 fractions F5-1 to F5-9 are collected; (5) The fraction F5-5 is gradient eluted with chloroform-methanol as the mobile phase by normal silica gel column chromatography, and 4 fractions F5-5-1 to F5-5-4 are collected; (6) The fraction F5-5-2 is gradient eluted with chloroform-methanol by gel column chromatography, impurities are removed, and the woodfordioid triterpene CNA compound is obtained by recrystallization in acetone.

4. The process for the preparation of the timber CNA of the toxane sesquiterpene according to claim 3, characterized by the fact that, In step (1), the solid-liquid ratio of the leaves of Coriaria sinica is 0.44 kg / L; and the volume ratio of the ethanol and water is 95:

5.

5. The process for the preparation of the timber CNA of the wood of the poison type sesquiterpene CNA according to claim 3, characterized by, In step (3), the gradient ratio of petroleum ether-acetone is 50:1 to 1:1; and the gradient ratio of dichloromethane-methanol is 10:1 to 1:

1.

6. The process for the preparation of the timber CNA of the wood of the poison type sesquiterpene CNA according to claim 3, characterized by, In step (4), the gradient ratio of dichloromethane-methanol is 70:1 to 3:

1.

7. The process for the preparation of the timber CNA of the wood of the poison type sesquiterpene CNA according to claim 3, characterized by, In step (5), the gradient ratio of chloroform-methanol is 8:1 to 2:

1.

8. The process for the preparation of the timber CNA of the wood of the poison type sesquiterpene CNA according to claim 3, characterized by, In step (6), the gradient ratio of chloroform-methanol is 10:1 to 3:

1.

9. The woodfordioid triterpene CNA of claim 1 is used for preparing a medicine for treating or preventing nervous system diseases, and the nervous system diseases include epilepsy, depression, anxiety, schizophrenia and neurodegenerative diseases.

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