Sesquiterpenoids with anti-epileptic activity as well as preparation method and application of sesquiterpenoids

Abstract

The invention discloses a sesquiterpenoids compound with anti-epileptic activity and a preparation method and application thereof, the sesquiterpenoids compound is characterized in that the structural formula of the sesquiterpenoids compound is shown as I, and the preparation method comprises the following steps: firstly, obtaining a fermentation product of marine fungi through fermentation culture; soaking the fermented product with ethyl acetate, and extracting to obtain a crude extract; according to the present invention, the crude extract is subjected to normal-phase silica gel column chromatography, reverse medium-pressure column chromatography and reversed-phase semi-preparative high performance liquid chromatography to separate and purify the crude extract so as to finally obtain the target sesquiterpenoids, and the target sesquiterpenoids have advantages of significant anti-epileptic effect and good development prospect, and can be used for research and development of drugs for prevention and treatment of epilepsy.

Description

Technical Field

[0001] The invention relates to a sesquiterpenoid compound, in particular to a compound with anti-epileptic activity extracted from secondary metabolites of marine fungi, and a preparation method and application thereof. Background Art

[0002] Epilepsy is a serious chronic neurological disease characterized by spontaneity, uncontrollability and recurrent attacks. The disease not only significantly increases the risk of patients suffering from various physiological complications, but also greatly increases the chance of sudden death. The pathological mechanisms of epilepsy are complex and diverse, usually involving abnormal discharges of neurons and pathological synchronization of brain electrical activity. They are also closely related to ion channel dysfunction, neurotransmitter imbalance, neural network abnormalities and genetic factors. Current anti-epileptic drugs (abbreviated as AEDs), such as phenytoin sodium, carbamazepine and valproate, mainly control epileptic seizures by regulating the above pathological mechanisms. However, these drugs have limited efficacy in certain patient groups and are often accompanied by serious adverse reactions such as hepatotoxicity, anemia, endocrine disorders and cognitive decline. In addition, about 30% of epilepsy patients show resistance to existing drugs, leading to the emergence of refractory epilepsy. An important mechanism of drug-resistant epilepsy is the overexpression of multidrug resistance (MDR) proteins in the blood-brain barrier, especially the overexpression of P-glycoprotein, which limits the penetration of antiepileptic drugs into the central nervous system, reduces the bioavailability of drugs, and thus weakens the control effect on epileptic seizures.

[0003] In view of the limitations of current anti-epileptic drugs in the treatment of drug-resistant epilepsy, there is an urgent need to develop new, more effective and less side-effect treatment strategies. In recent years, seaweed and its symbiotic fungi have become a hot topic of scientific research due to their complex and sophisticated symbiotic relationship. This symbiotic system can produce a variety of secondary metabolites with complex structures and diverse functions through chemical signaling, nutrient exchange and synergistic defense mechanisms. These metabolites not only have unique structural diversity, but also exhibit a wide range of biological activities, including antibacterial, anticancer, antiviral and neuromodulatory functions, especially in the field of anti-epileptic drug development, showing great application potential. At present, there are no relevant research reports on the extraction of compounds with anti-epileptic activity from the secondary metabolites of seaweed and its symbiotic fungi at home and abroad. Summary of the invention

[0004] The technical problem to be solved by the present invention is to provide a sesquiterpenoid compound with anti-epileptic activity and a preparation method and use thereof.

[0005] The technical solution adopted by the present invention to solve the above technical problems is:

[0006] 1. A sesquiterpenoid compound having anti-epileptic activity, wherein the structural formula of the sesquiterpenoid compound is as shown in (I):

[0007]

[0008] 2. The method for preparing the above-mentioned sesquiterpenoid compound having anti-epileptic activity comprises the following steps:

[0009] (1) Fermentation production

[0010] The Aspergillus pyrophorus with a deposit number of CCTCC NO: M2014086 was inoculated on a plate containing a potato dextrose agar solid medium (abbreviated as PDA solid medium), and after inverted culture in an incubator at 28° C. for 3 days for activation, the activated single colony was transferred to a potato dextrose liquid medium (abbreviated as PDB liquid medium) sterilized and cooled by high-pressure steam, and continued to shake culture at 28° C. and 150 rpm for 4 days to obtain a seed solution; 5 to 7 mL of the seed solution was inoculated into a rice culture medium, and incubated at 28° C. for 30 days to obtain a fermentation product;

[0011] (2) Extraction

[0012] Add ethyl acetate to the fermentation product obtained in step (1), extract repeatedly for 3 to 4 times, and evaporate the ethyl acetate extract to dryness to obtain a crude extract;

[0013] (3) Isolation, purification and preparation of compounds

[0014] First, the crude extract obtained in step (2) is fully dissolved in a mixed solvent of dichloromethane and methanol in a volume ratio of 1:1, and then 200-300 mesh silica gel is added to mix the sample, and normal phase medium pressure column chromatography is performed, and a mixed solution composed of petroleum ether and ethyl acetate in a volume ratio of 2:3 is used as an eluent for elution, and the eluate is collected and added to 100-200 mesh reversed phase silica gel for mixing, and then reversed phase medium pressure column chromatography is performed, and a methanol-water eluent with a methanol volume percentage of 20-100% is used for linear gradient elution, and the elution time is 120min. The eluted fractions are collected and arranged in descending order according to the polarity of the fractions, and 7 components are obtained by merging; the obtained 7th component is separated and purified by semi-preparative reversed phase high performance liquid chromatography using a mixed solution composed of acetonitrile and water in a volume ratio of 50:50 as a mobile phase to obtain compound I, whose structure is shown in (I):

[0015]

[0016] Furthermore, the preparation method of the PDA solid culture medium described in step (1) is as follows: 200 g of peeled and diced potatoes are boiled for 30 minutes and then filtered, the filtrate is retained, 20 g of glucose and 20 g of agar are added to the filtrate, heated until completely dissolved, cooled to below 50° C., the volume is adjusted to 1000 mL with distilled water, and after high-pressure sterilization at 121° C. for 20 minutes, cooled to 45-50° C., poured into a sterile culture dish, and waited for it to solidify before use.

[0017] Furthermore, the preparation method of the PDB liquid culture medium described in step (1) is as follows: take 200g of peeled and diced potatoes, add 1000mL of distilled water and boil for about 30min, filter and retain the filtrate, add 20g of glucose to the filtrate, stir until completely dissolved, adjust the volume to 1000mL with distilled water, sterilize 15-20mL at 121°C under high pressure, and then cool it for use in culture.

[0018] Furthermore, the preparation method of the rice culture medium described in step (1) is as follows: take 100g of rice, add 110mL of distilled water and 3g of sea salt, put the mixture into a culture bottle, and then sterilize it with high-pressure steam at 121°C for 20min and then cool it for inoculation and fermentation.

[0019] Furthermore, the flow rate of the mobile phase of the semi-preparative reverse-phase HPLC described in step (3) is 2 mL / min.

[0020] The present invention also provides use of the sesquiterpenoid compound in preparing anti-epileptic drugs.

[0021] Compared with the prior art, the advantages of the present invention are: the present invention provides a sesquiterpenoid compound with anti-epileptic activity, a preparation method and use thereof, obtains a fermentation product by microbial fermentation culture, then extracts the fermentation product by immersion in ethyl acetate to obtain a crude extract, and then separates and purifies the crude extract by medium-pressure normal-phase column chromatography, medium-pressure reverse-phase column chromatography, and semi-preparative high-performance liquid chromatography, and the compound has significant anti-epileptic activity and can be used for the development of drugs for inhibiting related diseases caused by epilepsy.

[0022] The Aspergillus ustus mentioned above is the DJ003 strain, which is classified and named Aspergillus ustus, with a preservation number of CCTCCNO: M2014086. It was deposited in the China Center for Type Culture Collection on March 14, 2014, and the preservation address is Wuhan University, Wuhan, China. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 is the high resolution mass spectrum (HR-ESI-MS) of compound I of the present invention;

[0024] Figure 2 is the ultraviolet spectrum (UV) of compound I of the present invention;

[0025] Figure 3 is the infrared spectrum (IR) of compound I of the present invention;

[0026] Figure 4 is the hydrogen nuclear magnetic resonance spectrum of compound I of the present invention ( 1 HNMR);

[0027] Figure 5 is the carbon NMR spectrum of compound I of the present invention ( 13 CNMR);

[0028] Figure 6 is the nuclear magnetic resonance DEPT-135 spectrum of compound I of the present invention;

[0029] Figure 7 is the nuclear magnetic resonance COSY spectrum of compound I of the present invention;

[0030] Figure 8 is the nuclear magnetic resonance HSQC spectrum of compound I of the present invention;

[0031] Fig. 9 is the nuclear magnetic resonance HMBC spectrum of compound I of the present invention;

[0032] Fig.10 is the nuclear magnetic resonance NOESY spectrum of compound I of the present invention;

[0033] Fig.11 The key to compound I of the present invention 1 H- 1 H COSY (blue thick line) and HMBC (red arrow) correlation;

[0034] Fig.12 Key NOE correlations of Compound I of the present invention;

[0035] Fig.13 Experimental and calculated values ​​of the ECD spectrum of compound I of the present invention;

[0036] Fig.14 Results of the anti-epileptic activity of compound I of the present invention in a zebrafish model, wherein (A) Typical movement trajectory of zebrafish at 6 dpf. (BE) Total distance, average speed, high movement frequency and curvature. (F) AChE activity. (GI) Epilepsy-related mRNA levels, p<0.05 compared with the control group. # <0.05 (significant), compared with the model group, p * <0.05 (significant). DETAILED DESCRIPTION

[0037] The present invention is further described in detail below with reference to the accompanying drawings:

[0038] Example 1: A sesquiterpenoid compound with anti-epileptic activity derived from marine fungi, the structural formula of the compound is shown in (I):

[0039]

[0040] The preparation method of the sesquiterpenoid compound shown in Example 2 and Example 1 is as follows:

[0041] Step 1: Fermentation production

[0042] The Aspergillus pyrophorus with the accession number of CCTCC NO: M2014086 was streaked on PDA solid medium (formula: 200 g peeled and diced potato, 20 g glucose, 15-20 g agar, 1000 mL distilled water), and after activation at 28°C for 3 days, the mycelium was cut into small pieces with a sterile inoculating loop, and the mycelium was transferred to 10 bottles of 1L round-bottom conical bottles sterilized at 121°C, each bottle containing 300 mL PDB liquid medium (formula: 200 g peeled and diced potato, 20 g glucose, 1000 mL distilled water). After inoculation, the culture bottle was placed on a rotary oscillator at 28°C and 150rpm for 4 days to obtain seed liquid; then 400 bottles of 1L round-bottom conical bottles were prepared, each bottle was equipped with rice culture medium (formula: 100g rice, 110mL distilled water, 3.0g sea salt), and after high-pressure sterilization at 121°C and cooling, 5-7mL of seed liquid was inoculated into each rice culture medium. The inoculated culture bottles were incubated under static conditions at 28°C for 30 days to obtain fermentation products. The entire inoculation and fermentation process were carried out in a sterile environment to ensure contamination-free.

[0043] Step 2: Extraction of extract

[0044] Add sufficient amount of ethyl acetate to the fermentation product obtained in step 1, extract repeatedly for 3-4 times, and then vacuum evaporate the ethyl acetate extract using a rotary evaporator to obtain a crude extract;

[0045] Step 3: Isolation and preparation of compounds

[0046] The crude extract obtained in step 2 was first fully dissolved in a mixed solvent of dichloromethane and methanol in a volume ratio of 1:1, and then 200-300 mesh silica gel was added to mix the sample and separated by normal phase medium pressure column chromatography (the separation column used was Silica Flash Column 330g), using a mixed solution of petroleum ether and ethyl acetate in a volume ratio of 2:3 as an eluent for elution, collecting the eluate and adding 100-200 mesh reversed-phase silica gel to mix the sample, and then performing reversed-phase medium-pressure column chromatography separation, using methanol-water with a methanol volume percentage of 20-100% as an eluent for linear gradient elution, the elution time is 120min, collecting the eluted fractions, arranging them from large to small according to the polarity of the fractions, and merging to obtain 7 components; the obtained 7th component is separated and purified by semi-preparative reversed-phase high performance liquid chromatography (HPLC) using a mixed solution of acetonitrile and water in a volume ratio of 1:1 as a mobile phase at a flow rate of the mobile phase of 2mL / min to obtain compound I, whose structural formula is shown in (I) in Example 1.

[0047] Example 3: Structural identification of the sesquiterpenoid compounds prepared by the method of Example 2.

[0048] The sesquiterpene compound I prepared in Example 2 above is a white powder. Figure 1 is the HR-ESI-MS spectrum of the compound of the present invention, which shows the quasi-molecular ion peak m / z 275.2376 [M+H] + (The calculated molecular weight is 275.275.2369, C 19 H 31 O), and its molecular formula is determined to be C 19 H 30 O, the unsaturation is 5. Figure 4 As shown, compound I 1 The H NMR spectral data (Table 1) show the presence of three distinct olefin protons, located at δ H 5.07(1H,br s,H-4),δ H 7.15 (1H, d, J = 16.6 Hz, H-11), and δ H 5.97 (1H, d, J = 16.6 Hz, H-12); In addition, three groups of methyl doublet peak signals were observed, located at δ H 0.94(3H,d,J=7.0Hz,H-2'), δ H 0.80 (3H, d, J = 6.8 Hz, H-7') and δ H 0.85 (3H, d, J = 6.5 Hz, H-9'); and three methyl singlets, located at δ H 1.01(3H,s,H-1'),δ H1.61(3H,s,H-3') and δ H 2.21 (3H, s, H-14). In addition, a comprehensive analysis of the one-dimensional carbon spectrum of the compound 13 C NMR (such as Figure 5 )、DEPT(such as Figure 6 ) and HSQC spectra (such as Figure 8 ), compound I showed 19 carbon signals, including two methylene carbons, located at δ C 46.9(C-8) and δ C 42.1 (C-10); five methyl carbon signals, located at δ C 49.2(C-2),δ C 39.6(C-5),δ C 47.3(C-6),δ C 34.8(C-7) and δ C 32.5 (C-9); a saturated quaternary carbon signal is located at δ C 41.3 (C-1); an unsaturated quaternary carbon signal is located at δ C 135.3 (C-3); and a carbonyl carbon signal in the low-field region, located at δ C 198.0 (C-13). These data indicate that compound I contains two conjugated C=C double bonds and a carbonyl group, and these functional groups contribute three degrees of unsaturation, with the other two degrees of unsaturation being attributed to two fully saturated rings. Figure 2 As shown, the UV spectrum has a maximum absorption at 211 nm, indicating that compound I has a conjugated π system. Figure 3 As shown, 1700cm in the infrared spectrum -1 Up to 1725cm -1 The characteristic absorption band between the two further confirmed the existence of carbonyl group.

[0049] Table 1. Compound I 1 H and 13 C NMR data (600 MHz, 150 MHz, DMSO-d6)

[0050]

[0051] Note 1: s—singlet, d—doublet, q—quartet, m—multiplet, overlapped—overlapped, br s—broad singlet.

[0052] Note 2: 1 H was obtained at 600 MHz NMR; 13 C was obtained at 150 MHz NMR.

[0053] The planar structure of compound I was further confirmed by detailed analysis of one-dimensional and two-dimensional NMR correlations. Figure 7 As shown, 1 H- 1 In the H COSY spectrum, δ H 0.76 and δ H The cross signal between 1.80 indicated that the compound had a saturated closed ring structure CH(5)-CH(6)-CH(7)-CH2(8)-CH(9)-CH2(10)-CH(5), confirming the connection between H-7 and H-7', H-9 and H-9'. Fig. 9 As shown in the HMBC spectrum, the correlation between H-7' and C-6 / C-7 / C-8, the correlation between H-9' and C-8 / C-9 / C-10, and the correlation between H-4 and C-5 / C-6 / C-10 support the assignment of two methyl groups (C-7' and C-9') and one olefin carbon (C-4). H 0.76, overlapped) and H-10b (δ H 1.72,m) and C-6 / C-8, H-2(δ H 1.45,q,J=7.2,6.6Hz) and C-1 / C-2' / C-3' / C-3 / C-4 / C-6, further confirming the presence of two six-membered rings (ring A and ring B) in the compound and clarifying the positions of the two methyl groups (C-2' and C-3'). Combined with the analysis results of unsaturation, it was confirmed that compound I contains two ring structures. In addition, the HMBC correlation between H-14 and C-12 / C-13, the correlation between H-11 and C-1 / C-1' / C-6 / C-13, and the correlation between H-11 (δ H 7.15, d, J = 16.6 Hz) and H-12 (δ H 5.97, d, J = 16.6 Hz) These two hydrogen atoms 1 H- 1 The correlation in the H COSY spectrum further confirmed a methyl group (C-1′) and a methyl acrylate structural unit, which was assigned to the C-1-attached unit (see the schematic diagram of COSY and HMBC signals of compound I for details). Fig.11 shown).

[0054] The relative configuration of compound I was deduced by analysis of coupling constants and NOESY spectra. The coupling constants of H-11 (J = 16.6 Hz) and H-12 (J = 16.6 Hz) indicated that the double bond was in trans configuration, with two hydrogen atoms located on opposite sides of the double bond, corresponding to the E configuration. Fig.10As shown, the cross peaks between H-11 and H-6 / H-7' / H-9', H-12 and H-1' / H-2 / H-5 / H-7 in the NOESY spectrum, as well as the coupling constant (J = 7.00 Hz) between H-2 and H-2', indicate that ring A and ring B are in trans fusion, and the β orientation of Me-2', Me-7', Me-9' and H-6, and the α orientation of Me-1', H-2, H-5, H-7 and H-9 are established (as shown in Figure 2). Fig.12 shown).

[0055] The electronic excited state of compound I was predicted by quantum chemical methods based on TD-DFT, and its rotational strength and transition energy were calculated. Two possible absolute configurations were proposed, namely (1S, 2R, 5S, 6R, 7S, 9R) and its enantiomer (1R, 2S, 5R, 6S, 7R, 9S). Subsequently, quantum chemical TD-DFT calculations were performed on these two structures to predict their ECD spectra. Fig.13 As shown, the calculated ECD curve of the (1R, 2S, 5R, 6S, 7R, 9S) configuration is consistent with the experimental results. Therefore, the structure of compound I was constructed.

[0056] Example 4. Antiepileptic activity of sesquiterpenoid compound I prepared by the method of Example 2.

[0057] 1. Preparation of experimental sample solution: Pentylenetetrazol (abbreviated as PTZ) was used as an epilepsy inducing agent to establish a zebrafish inflammation model. The test sample was the pure compound I separated and purified in Example 2 above. An appropriate amount of sample was accurately weighed and prepared with DMSO into a solution of the required concentration for testing anti-epileptic activity (5 mg / L). The zebrafish used in this experiment were raised in a flow-through water tank, the water temperature was maintained at 28±0.5°C, and the light cycle was 14 hours of light: 10 hours of darkness. Zebrafish were fed with newly hatched Artemia.

[0058] 2. Experimental methods: Zebrafish larvae 6 days after fertilization were randomly assigned to experimental groups, each containing 100 larvae. The experimental groups included a control group, a PTZ-treated group, a compound I-treated group (5 mg / L), and a positive control group (50 μM CBZ) treated with carbamazepine (CBZ). Except for the control group, all experimental groups were pre-treated with 10 mM PTZ for 30 minutes, and then the corresponding compound I and positive drug CBZ were added to the compound-treated group and the positive drug CBZ-treated group. The zebrafish larvae were placed in a dark environment to adapt for 10 minutes, and then the behavior was recorded for 15 minutes. The behavioral data were analyzed using EthoVision XT 10.0 software, and the main parameters of interest included travel distance and epileptic-like behavior.

[0059] For gene expression analysis, 30 zebrafish larvae were randomly collected from each group and homogenized. Total RNA was extracted using a spin column-based animal total RNA purification kit. Subsequently, 1.0 μg of RNA was reverse transcribed into cDNA using a cDNA synthesis premix. Quantitative real-time PCR (qPCR) was performed on a qTOWER 3G instrument using a fast SYBR mix to measure relative mRNA levels, with elongation factor 1α (ef1α) as an internal reference gene. The cycling conditions for qPCR were: initial denaturation at 94°C for 180 seconds, followed by 42 cycles, each cycle consisting of denaturation at 94°C for 5 seconds and annealing at 60°C for 30 seconds. Melting curve analysis was performed in a temperature range of 60°C to 95°C with an increment of 1°C. The primer sequences of the target genes are shown in Table 1.

[0060] Table 1. Primer sequences of epilepsy-related target genes used in the study

[0061]

[0062] Acetylcholinesterase (AChE) activity was detected according to the kit instructions. Total protein was extracted from 60 zebrafish embryos in each group, and protein quantification was performed using Coomassie Brilliant Blue G-250. The absorbance was measured on a PerkinElmer multifunctional microplate reader, and each sample was repeated three times. All data are expressed as mean ± standard error (SEM), and statistical analysis was performed using GraphPad Prism 8.0 software. Statistical significance was assessed by unpaired t-test, and the significance level was set at p < 0.05.

[0063] 3. Experimental results: The anti-epileptic activity of compound I of the present invention in the zebrafish model is as follows Fig.14 As shown, the 1 on the horizontal axis in the figure is the compound.

[0064] (1) Behavioral improvement: Fig.14 As shown in Figures A, B, C, D, and E, compound I effectively alleviated PTZ-induced hyperactivity behavior in zebrafish, including reductions in parameters such as total travel distance, average speed, height movement frequency, and tortuosity. The motor performance was almost restored to a level comparable to or even better than that of the CBZ-positive control group, indicating that epileptic seizure-related hyperactivity was significantly reduced.

[0065] (2) Increase the activity of acetylcholinesterase (AChE): Fig.14 As shown in Figure F, compared with the blank control group (control group), compound I significantly increased the AChE activity in zebrafish, indicating that it may exert anti-epileptic effects by regulating the cholinergic pathway. The increase in AChE activity was significantly different from that in the positive control group (p<0.05). # <0.05 (significant), compared with the model group, p* <0.05 (significant).

[0066] (3) Regulate the expression of related genes: Fig.14 As shown in G, H, and I, in the PTZ-induced epilepsy model, compound I can effectively reverse the upregulation of c-fos and gria1b genes and the downregulation of gat1 gene, restoring the expression levels of these genes to close to the control group. This indicates that compound I helps to rebalance excitatory glutamate and inhibitory GABAergic neurotransmission, maintain normal neuronal excitability, and prevent epileptic seizures. Compared with the control group, p # <0.05 (significant), compared with the model group, p * <0.05 (significant).

[0067] The above description is not intended to limit the present invention, and the present invention is not limited to the above examples. Any changes, modifications, additions or substitutions made by a person skilled in the art within the spirit and scope of the present invention shall also fall within the protection scope of the present invention.

Claims

1. A sesquiterpenoid compound having anti-epileptic activity, characterized in that The compound is a sesquiterpenoid compound derived from marine fungi, and its structural formula is shown in (I).

2. The method for preparing a sesquiterpenoid compound having anti-epileptic activity according to claim 1, characterized in that The steps include: (1) Fermentation production The Aspergillus pyrophorus with a deposit number of CCTCC NO: M2014086 was inoculated on a plate containing a PDB solid culture medium, and after inverted culture in an incubator at 28° C. for 3 days for activation, the activated single colony was transferred to a PDB liquid culture medium that had been sterilized by high-pressure steam and cooled, and continued to be shaken and cultured at 28° C. and 150 rpm for 4 days to obtain a seed solution, 5 to 7 mL of the seed solution was inoculated into a rice culture medium, and the culture was statically incubated at 28° C. for 30 days to obtain a fermentation product; (2) Extraction Add ethyl acetate to the fermentation product obtained in step (1), extract repeatedly for 3 to 4 times, and evaporate the ethyl acetate extract to dryness to obtain a crude extract; (3) Isolation, purification and preparation of compounds First, the crude extract obtained in step (2) is fully dissolved in a mixed solvent of dichloromethane and methanol in a volume ratio of 1:1, and then 200-300 mesh silica gel is added to mix the sample, and normal phase medium pressure column chromatography is performed, and a mixed solution composed of petroleum ether and ethyl acetate in a volume ratio of 2:3 is used as an eluent for elution, and the eluate is collected and added to 100-200 mesh reversed phase silica gel for mixing, and then reversed phase medium pressure column chromatography is performed, and a methanol-water eluent with a methanol volume percentage of 20-100% is used for linear gradient elution, and the elution time is 120min. The eluted fractions are collected and arranged in descending order according to the polarity of the fractions, and 7 components are obtained by merging; the obtained 7th component is separated and purified by semi-preparative reversed phase high performance liquid chromatography using a mixed solution composed of acetonitrile and water in a volume ratio of 50:50 as a mobile phase to obtain compound I, whose structure is shown in (I):

3. The method for preparing a sesquiterpenoid compound having anti-epileptic activity according to claim 2, characterized in that The preparation method of the PDA solid culture medium described in step (1) is as follows: 200 g of peeled and diced potatoes are boiled for 30 minutes and then filtered, the filtrate is retained, 20 g of glucose and 20 g of agar are added to the filtrate, heated until completely dissolved, cooled to below 50° C., the volume is adjusted to 1000 mL with distilled water, and after high-pressure sterilization at 121° C. for 20 minutes, cooled to 45-50° C., poured into a sterile culture dish, and waited for it to solidify before use.

4. The method for preparing a sesquiterpenoid compound having anti-epileptic activity according to claim 2, characterized in that The preparation method of the PDB liquid culture medium described in step (1) is as follows: take 200g of peeled and diced potatoes, add 1000mL of distilled water and boil for about 30min, filter and retain the filtrate, add 20g of glucose to the filtrate, stir until completely dissolved, adjust the volume to 1000mL with distilled water, sterilize 15-20mL at 121°C under high pressure, and then cool.

5. The method for preparing a sesquiterpenoid compound having anti-epileptic activity according to claim 2, characterized in that The preparation method of the rice culture medium described in step (1) is as follows: take 100g of rice, add 110mL of distilled water and 3g of sea salt, put the mixture into a culture bottle, and then sterilize it with high pressure steam at 121°C for 20min and then cool it.

6. The method for preparing a sesquiterpenoid compound with anti-epileptic activity according to claim 2, characterized in that: The flow rate of the semi-preparative reversed-phase HPLC compound separation preparation described in step (3) is 2.0 mL / min.

7. Use of the sesquiterpenoid compound according to claim 1 in the preparation of anti-epileptic drugs.

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