Novel amide alkaloid compound in purslane as well as extraction and separation method and application thereof

Abstract

The invention belongs to the technical field of traditional Chinese medicine extraction and separation, and particularly relates to a novel amide alkaloid compound in purslane as well as an extraction and separation method and application thereof. The molecular formula of the novel amide compound is C9H13NO2, and the novel amide compound is named as 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide, and the molecular formula of the novel amide compound is as shown in the specification. The invention also provides an extraction and separation method of the novel compound, which sequentially adopts water extraction, macroporous resin adsorption and elution, ODS medium-pressure column preparation, Sephadex LH-20 and high performance liquid chromatograph preparation, and successfully extracts and separates the novel amide alkaloid compound. The structure is determined by mass spectrum, carbon spectrum, hydrogen spectrum and two-dimensional nuclear magnetic spectrum analysis methods. The novel compound has anti-inflammatory and anti-cholinesterase effects, and the novel compound and salts or derivatives thereof can be used as synthetic primers of other compounds and raw materials for new drug development and pharmacological activity research, and can be used for preparing anti-inflammatory and anti-cholinesterase drugs or health care products.

Description

Technical Field

[0001] The invention belongs to the technical field of traditional Chinese medicine extraction and separation, and in particular relates to a new amide alkaloid compound in purslane and an extraction and separation method and application thereof. Background Art

[0002] Portulaca oleracea L., also known as horse amaranth, ant vegetable, and five-element grass, is an annual herbaceous plant of the Portulacaceae family. Portulaca oleracea is widely distributed and rich in resources. It is one of the 78 wild plants with medicinal and edible properties specified by the former Ministry of Health of my country. Portulaca oleracea is included in the 2020 edition of the Pharmacopoeia of the People's Republic of China. It has the effects of clearing away heat and detoxifying, cooling blood and stopping bleeding, and stopping dysentery. It is used for heat-toxic bloody dysentery, carbuncle, furuncle, eczema, erysipelas, snake and insect bites, blood in stool, hemorrhoids, metrorrhagia, etc.

[0003] Modern pharmacological studies on purslane have shown that it has the effects of lowering blood pressure, lowering blood lipids, anti-oxidation, anti-inflammatory, anti-viral, anti-cancer, neuroprotective, and regulating immune function. The main chemical components of purslane include flavonoids, coumarins, volatile oils, terpenes, steroids, alkaloids, amino acids, polysaccharides, various pigments and mineral substances, which provide the material basis for its diverse pharmacological effects. Among them, alkaloid compounds are a major class of chemical components in purslane. The currently reported alkaloid components include 1-methanesulfonic acid-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, 5-hydroxy-2-carboxypyridine, tropaeoline, 3-isobutyl-6-methylpiperazine-2,5-dione, oleraciamide E, oleraciamide F, oleraciamide G, and oleracrylimide AF.

[0004] Currently, most of the chemical components isolated from Purslane are known and have low structural novelty. Therefore, the development and separation of new compounds in Purslane are urgently needed. Summary of the invention

[0005] In view of the above problems, the present invention provides a new amide alkaloid compound extracted and separated from Portulaca oleracea. Studies have found that the new compound of the present invention has anti-inflammatory and anticholinesterase effects. At the same time, a simple, rapid, environmentally friendly and high-purity extraction and separation method for the new compound of the present invention is provided.

[0006] In order to achieve the above-mentioned object of the present invention, the present invention provides a new amide alkaloid compound, the molecular formula of which is C 9 H 13 NO 2 , named 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide according to the structural formula, the chemical structure is:

[0007]

[0008] In order to achieve the above-mentioned object of the present invention, the present invention also provides a method for extracting and separating a new amide alkaloid compound in Portulaca oleracea, which specifically comprises the following steps:

[0009] Step 1, take dried purslane medicinal material, extract it with water, filter the water extract, combine the filtrate, concentrate it under reduced pressure, cool it to room temperature, and obtain the medicinal solution for use;

[0010] Step 2, evaporating the Chinese medicine solution in step 1 to dryness, adsorbing it with a macroporous resin, eluting it with a water-ethanol gradient, and concentrating the hot water portion under reduced pressure to obtain an extract, thereby obtaining a hot water extract;

[0011] Step 3, separating the hot water extract in step 2 through an ODS (octadecylsilane bonded silica gel filler) column, using a methanol-water gradient elution to obtain several elution fractions, detecting them through thin layer chromatography, developing colors, combining the elution fractions with developed colors, and concentrating the combined elution fractions to dryness under reduced pressure for later use;

[0012] Step 4, separating the color-developed part in step 3 through an ODS (octadecylsilane bonded silica gel filler) column, using a methanol-water gradient elution to obtain several elution parts, detecting by thin layer chromatography, developing color, combining the color-developed elution parts, and concentrating the combined elution parts to dryness under reduced pressure for standby use;

[0013] Step 5, the obtained product in step 4 is further separated by chromatography on a pretreated Sephadex LH-20 column, isocratically eluted with 20% methanol to obtain several elution fractions, which are detected by thin layer chromatography, developed, and the developed elution fractions are combined, and the combined elution fractions are concentrated to dryness under reduced pressure for later use;

[0014] Step 6: The concentrate obtained in step 5 is separated and prepared by HPLC (high performance liquid chromatography), and methanol: 0.1% formic acid is used as a mobile phase for isocratic elution to prepare the new amide alkaloid compound of the present invention.

[0015] Furthermore, in step 1, water reflux extraction is performed twice, each time for 2 hours, and the amount of water used is 10 times that of the medicinal material.

[0016] Furthermore, the mobile phase elution procedure used in step 2 is a gradient elution of cold water, hot water, 30% ethanol, 50% ethanol, and 70% ethanol.

[0017] Furthermore, in step 3, the volume ratio of water to methanol is 100:0, 95:5, 90:10, 80:20 and 100:0 for gradient elution.

[0018] Furthermore, in step 4, the volume ratio of methanol to water is 100:0, 95:5, 90:10, 80:20 and 100:0 for gradient elution.

[0019] Furthermore, the pretreatment process of the ODS and dextran gel is: soaking in methanol for 24 hours, loading on the column, washing with methanol until there is no turbidity when dripped into water, and then balancing with the initial mobile phase.

[0020] Furthermore, the methanol elution procedure in step 5 is 20% methanol isocratic elution.

[0021] Furthermore, in step 6, the volume ratio of methanol to water in the isocratic elution of methanol:0.1% formic acid is 45:55.

[0022] The novel amide alkaloid compound of the invention can be used for preparing anti-inflammatory and anti-cholinesterase drugs or health products.

[0023] Compared with the prior art, the invention has the beneficial effects.

[0024] The separation and pharmacological activity research of the new amide alkaloid compound from purslane described in the present invention has not been reported in existing paper journals; the present invention provides a new amide alkaloid compound derived from purslane and an extraction and separation method for the new compound of the present invention, which sequentially adopts water extraction, macroporous resin adsorption elution, ODS medium-pressure column, Sephadex LH-20 and high-performance liquid chromatography for separation, purification and preparation, and successfully extracts and separates the new compound. The method has only five operating steps, and the operation method is simple and fast. The extraction and separation process mainly adopts water extraction and methanol elution, the process method is environmentally friendly, and the purity of the compound separated by the method is greater than 90%. In addition, studies have shown that the compound has anti-inflammatory and anti-cholinesterase effects. Therefore, the new compound of the present invention and its salts and derivatives can be used as synthetic leads of other compounds, as well as raw materials for new drug development and pharmacological activity research, and can also be used to prepare anti-inflammatory and anti-cholinesterase drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 It is a high-resolution mass spectrum of the new amide alkaloid compound 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide of the present invention.

[0026] Figure 2 The novel amide alkaloid compound 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide of the present invention 1 H-NMR spectrum.

[0027] Figure 3The novel amide alkaloid compound 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide of the present invention 13 C-NMR spectrum.

[0028] Figure 4 The DEPT spectrum of the new amide alkaloid compound 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide of the present invention is shown in FIG.

[0029] Figure 5 This is the HMBC spectrum of the new amide alkaloid compound 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide of the present invention.

[0030] Figure 6 The novel amide alkaloid compound 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide of the present invention 1 H- 1 H COSY spectrum.

[0031] Figure 7 This is the HSQC spectrum of the new amide alkaloid compound 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide of the present invention.

[0032] Figure 8 The figure is the ROESY spectrum of the new amide alkaloid compound 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide of the present invention. DETAILED DESCRIPTION

[0033] The present invention provides a new compound having the molecular formula C 9 H 13 NO 2 , named 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide, the chemical formula is:

[0034]

[0035] Table 1 shows the NMR data of the new amide alkaloid compound: 1 H-NMR and 13 C-NMR in DMSO.

[0036] Table 1 NMR data of the new amide alkaloid compound 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide.

[0037]

[0038] The structural identification of the compounds of the present invention is shown in Figure 1-Figure 8 .

[0039] 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide: light yellow powder, easily soluble in water, slightly soluble in methanol. UHPLC-ESI-Q-TOF-MS gives m / z: 168.1019 [M+H] + The quasi-molecular ion peak of is 168.1019. 1 H-NMR, 13 C-NMR and DEPT data suggest that the possible molecular formula of the compound is C 9 H 13 NO 2 , the unsaturation is 4. 13 The C-NMR spectrum and DEPT spectrum showed 9 carbon signals, including 2 methyl carbons (δ C :27.14;27.20)、1 methylene carbon (δ C :46.69)、3 methyl carbons (δ C :44.32;128.32;128.61)、2 carbonyl carbons (δ C :168.93;200.04)、1 quaternary carbon (δ C :49.70).

[0040] 1 The H-NMR spectrum shows 2 CH 3 The signal is δ H 1.04 (3H, s), δ H 1.05 (3H, s); 1 methylene signal is δ H 2.57, 2.60 (2H, dd, J = 7.74, 17.64 Hz), and the three methine signals are δ H 2.46 (1H, d, J=17.64, 7.74Hz), δ H 6.34 (2H, d, J = 10.80 Hz, overlap). 13 The C-NMR spectrum shows a typical acetyl signal, C-1′(δ C :200.04), C-2′(δ C :27.20); HMBC spectrum shows H-2′(δH 1.05, 3H, s) and C-5 (δ C :49.70) correlation, indicating that C-1′ is connected to C-5; according to H-1″ (δ H 1.04) and C-1′, H-4 (δ H 2.57, 2.60) are related to C-1′, indicating that C-1″ and C-4 are both connected to C-5. 1 H-2(δ H 6.34) and H-3(δ H 6.34) overlap, and it is inferred that C-2 is directly connected to C-3; 1 H- 1 The H COSY spectrum shows that H-4 is related to H-3, H-2 is related to H-1 (δ H 2.46), indicating that C-1, C-2, C-3, and C-4 are connected in sequence. The HMBC spectrum shows that H-2 is correlated with C-1″′, indicating that C-1″′ is connected to C-2 through C-1; H-1 is correlated with C-1″, indicating that C-1 is connected to C-1″ through C-5, that is, there is a substituted unsaturated five-membered ring. 13 The C-NMR spectrum shows that C-1″′ is located in the high field, indicating that the amino group is directly connected to C-1″′. Therefore, based on the above information, it can be determined that this new compound has the above structure.

[0041] The present invention also provides a method for extracting and separating the above-mentioned amide alkaloid compounds, which specifically comprises the following steps:

[0042] Step 1: Weigh 250 kg of dried purslane medicinal material, extract it with water, the amount of water is 10 times the amount of the medicinal material, extract it twice, each time for 2 hours, filter the water extract, combine the filtrate, concentrate it under reduced pressure to 250 L, cool it to room temperature, and obtain the medicinal solution for use.

[0043] Step 2: evaporate the drug solution obtained in step 1 to dryness, adsorb and elute with AB-8 resin, and isocratically elute with hot water. Recover the drug solution to an extract under reduced pressure at below 80°C to obtain a hot water extract.

[0044] Step 3: The hot water extract in step 2 is separated by medium- and low-pressure ODS columns, wherein the filler particle size is 40-70 μm, and gradient elution is performed using methanol-water (0 / 100, 5 / 95, 10 / 90, 20 / 80, 50 / 50, 100 / 0, v / v) to obtain 35 fractions in total (i.e., 35 bottles in total, each with 100 mL), which are detected by thin layer chromatography and color developed. The colored 6-15 eluted fractions are combined, and the combined 6-15 fractions are concentrated to dryness under reduced pressure below 75° C. for later use.

[0045] Step 4: The product obtained in step 3 is further separated by pretreated ODS medium-pressure column chromatography, wherein the filler particle size is 20-40 μm, and gradient elution is performed with methanol-water (0 / 100, 5 / 95, 10 / 90, 20 / 80, 50 / 50, 100 / 0, v / v) (pressurized to a flow rate of 1 mL / min and room temperature) to obtain 14 fractions (i.e., 14 bottles are obtained by gradient elution, each with 120 mL), which are detected by thin layer chromatography and color developed. The colored 4-8 fractions are retained and concentrated to dryness under reduced pressure below 50° C. for later use.

[0046] Step 5: The obtained in step 4 is separated by pre-treated dextran gel column chromatography (Sephadex LH-20), eluted with methanol, and 30 elution fractions are obtained (i.e., 30 bottles are obtained, each bottle is 50 mL), detected by thin layer chromatography, color is developed, and the 16-22 colored fractions are retained, and concentrated to dryness under reduced pressure below 50°C for standby use. The pre-treatment process of the ODS and dextran gel is to soak in methanol for 24 hours, load the column, wash with methanol until there is no turbidity when dripped into water, and then balance with the initial mobile phase.

[0047] Step 6: The product obtained in step 5 was separated and prepared by HPLC, with methanol: 0.1% formic acid (45:55, v / v) as the mobile phase for isocratic elution, and the detection wavelengths were 210 nm and 254 nm, to separate and prepare the new amide alkaloid compounds of the present invention, and the purity determined by the normalization method was 90-99%.

[0048] The anti-inflammatory effect of the novel amide alkaloid compound of the present invention.

[0049] 1Main materials.

[0050] 1.1 Drugs and reagents: The new amide alkaloid compounds used in the experiment were prepared by the above method with a purity of 90-99%. They were accurately weighed and diluted with DMSO to the required solutions for each dose group as follows: DMEM high-glucose medium, fetal bovine serum (Hyclone, USA); penicillin and streptomycin (Hangzhou Sijiqing Company); LPS (Sigma, USA); ELISA kits for IL-1β and TNF-α (Cayman, USA); cell lysate, Griess reagent (Biyuntian Biotechnology Co., Ltd.).

[0051] 1.2 Cell line: RAW264.7 macrophages (ATCC cell bank, USA).

[0052] 1.3 Grouping: Divided into control group, LPS group and experimental group, one group each.

[0053] 2 Experimental methods.

[0054] 2.1 Cell culture: DMEM high-glucose medium, with 10% fetal bovine serum, 1% antibiotics (100 U / mL penicillin and 100 μg / mL streptomycin), was placed at 37.5°C, CO 2 Culture in an incubator.

[0055] 2.2 CCK-8 assay for cell viability: The three groups of RAW264.7 macrophages in logarithmic growth phase were inoculated in 96-well culture plates at a cell density of 1×10 4 pcs / mL, 100 μL per well, 37°C, 5% CO 2 After culturing overnight under the conditions, different concentrations of the compound of the present invention, 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide (5μM~50μM) were added to the experimental group. After incubation for 1 hour, LPS with a concentration of 1μg / mL was added to the LPS group and the experimental group, and a zero adjustment group (culture solution containing DMSO solvent) was set up. Each group had 3 replicate wells to investigate the effect of the addition of drugs on the cells. After the cells in the above groups were cultured for 24 hours, 10μL of CCK-8 was added to the cells in each well at 37℃ and 5% CO 2 After incubation for 2 h under the same conditions, the absorbance of each well was measured at a wavelength of 450 nm using an ELISA reader.

[0056] 2.3ELISA method for determination of inflammatory factors IL-1β and TNF-α: RAW264.7 macrophages in the logarithmic growth phase were inoculated in 24-well culture plates at a cell density of 1×10 5 pcs / mL, 1 mL per well, temperature 37°C, 5% CO 2 The cells were cultured overnight under the conditions of 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide (1-20 μM), the new amide alkaloid compound of the present invention was added to the experimental group, and after 1 hour of incubation, LPS (final concentration of 1 μg / mL) was added to each well, and the cells were incubated for 24 hours. Each group was treated with 3 wells. The ELISA method was used to determine the content of IL-1β and TNF-α secreted by RAW264.7 macrophages after being treated with the new alkaloid compound derived from Purslane.

[0057] 3 Experimental results.

[0058] The experimental results show that the new amide alkaloid compound of the present invention has no effect on the proliferation of LPS-induced macrophages RAW264.7 at 25 μM, is safe and non-toxic; and can effectively inhibit the excessive inflammatory cytokines IL-1β and TNF-α produced by LPS-induced macrophages RAW264.7 in a concentration-dependent manner.

[0059] The relative survival rate experimental results of the new amide alkaloid compounds of the present invention are shown in Table 2.

[0060] Table 2 Effects of the novel amide alkaloid compounds of the present invention on the relative survival rate of RAW264.7 macrophages.

[0061]

[0062] Note: * P<0.05 compared with the control group. # P<0.05 compared with LPS group.

[0063] The results of ELISA determination of inflammatory factors IL-1β and TNF-α are shown in Table 3.

[0064] Table 3 Effects of the novel amide alkaloid compounds of the present invention on the levels of IL-1β and TNF-α secreted by RAW264.7 cells induced by LPS (mean ± standard deviation, n = 3).

[0065]

[0066]

[0067] Note: * P<0.05 compared with the control group. # P<0.05 compared with LPS group.

[0068] The anticholinesterase effect of the novel amide alkaloid compounds of the present invention.

[0069] 1Main materials.

[0070] 1.1 Drugs and reagents: The new compounds used in the experiment were prepared by the above method with a purity of 96%. Physostigmine (Shanghai Hanxiang Biotechnology Co., Ltd.), acetylcholine iodide (ATCI) and acetylcholinesterase (AChE) (Dalian Meilun Biotechnology Co., Ltd.), dithiodinitroformic acid (DTNB) (Shanghai Jinshui Biotechnology Co., Ltd.), disodium hydrogen phosphate and sodium dihydrogen phosphate (Shanghai Sinopharm Reagent Co., Ltd.).

[0071] 1.2 Experimental instruments and equipment: HBS-1096A96-well microplate reader (Nanjing Detie Experimental Equipment Co., Ltd.), 1 / 100,000 balance (METTLER, Switzerland), HH-4 digital display constant temperature water bath (Jiangsu Jintan Ronghua Instrument Manufacturing Co., Ltd.).

[0072] 2 Experimental methods.

[0073] This experiment was conducted to determine the anticholinesterase activity of new amine alkaloid compounds according to the modified Ellman method. 5-ac etyl-5-methylcyclopent-2-ene-1-carboxamide and physostigmine were accurately weighed and prepared into five series of sample solutions with concentrations of 3.125 μM, 6.25 μM, 12.5 μM, 25.0 μM and 50.0 μM using methanol. The specific operation is as follows: add 140 μL of phosphate buffered saline (0.1M, pH=8.0, containing 0.1 mol / L disodium hydrogen phosphate and sodium dihydrogen phosphate), 20 μL of sample solution, 15 μL of AChE (0.2U / mL) to a 96-well ELISA plate, incubate at 37°C for 10 minutes, add 10 μL of ATCI (4mmol / L) and 10 μL of DTNB (15mmol / L), incubate at 37°C for 20 minutes, place the 96-well plate in an ELISA instrument, and measure the absorbance of each group of samples at a wavelength of 405nm. Among them, methanol was used instead of the sample solution as the blank group, and physostigmine was used instead of the sample solution as the positive control group. The cholinesterase inhibition rate of each compound was calculated according to the following formula (A represents absorbance):

[0074] Inhibition rate (%) = (A 空白 -A 样品 ) / A 空白 ×100%

[0075] 3 Experimental results.

[0076] The experimental results show that the new compound of the present invention exhibits certain anticholinesterase activity, and the inhibitory effect on cholinesterase increases with the increase of the compound concentration, showing a dose-dependent trend.

[0077] The anticholinesterase activities of the novel amine alkaloid compounds of the present invention are shown in Table 4.

[0078] Table 4 Anticholinesterase activity of the novel amine alkaloid compounds of the present invention.

[0079]

[0080] In summary, the present invention provides a new amide alkaloid compound and an extraction and separation method thereof, which uses water extraction, macroporous resin adsorption elution, ODS medium pressure column, Sephadex LH-20 and high performance liquid chromatography for separation, purification and preparation, and successfully separates and obtains the new compound. The method is simple, rapid and environmentally friendly, and the compound separated by the method has a high purity. Since the obtained new amide alkaloid compound has a unique chemical structure and is extracted from the commonly used traditional Chinese medicine Portulaca oleracea, it has anti-inflammatory and anticholinesterase effects. Therefore, the new amide alkaloid compound and its salts and derivatives of the present invention can be used as natural products to develop new traditional Chinese medicines, and have broad prospects.

Claims

1. A new amide alkaloid compound isolated from Portulaca oleracea, characterized in that: The molecular formula is: C9H 13 NO2, and named 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide according to the structure, its chemical structure is as follows: 。 2. A method for extracting and separating a new amide alkaloid compound from Portulaca oleracea, characterized in that: The specific steps include: Step 1, take dried purslane medicinal material, extract it with water, filter the water extract, combine the filtrate, concentrate it under reduced pressure, cool it to room temperature, and obtain a medicinal solution for use; Step 2, evaporating the Chinese medicine solution in step 1 to dryness, adsorbing it with a macroporous resin, eluting it with a water-ethanol gradient, and concentrating the hot water portion under reduced pressure to obtain an extract, thereby obtaining a hot water extract; Step 3, separating the hot water extract in step 2 through an ODS column, using a methanol-water gradient elution to obtain several elution fractions, detecting them through thin layer chromatography, developing colors, combining the elution fractions with developed colors, and concentrating the combined elution fractions to dryness under reduced pressure for later use; Step 4, separating the color-developing part in step 3 through an ODS column, using a methanol-water gradient elution to obtain several elution parts, detecting by thin layer chromatography, developing color, combining the color-developing elution parts, and concentrating the combined elution parts to dryness under reduced pressure for standby use; Step 5, the obtained product in step 4 is further separated by pre-treated dextran gel column chromatography, isocratically eluted with 20% methanol to obtain several elution fractions, detected by thin layer chromatography, color developed, combined the color developed elution fractions, and concentrated the combined elution fractions to dryness under reduced pressure for standby use; Step 6: The concentrate obtained in step 5 is separated and prepared by HPLC, and methanol: 0.1% formic acid (v / v) is used as the mobile phase for isocratic elution to prepare the new amide alkaloid compound of the present invention.

3. The extraction and separation method according to claim 2, characterized in that: In step 1, water reflux extraction is performed twice, each time for 2 hours, and the amount of water used is 10 times that of the medicinal material.

4. The extraction and separation method according to claim 2, characterized in that: The mobile phase elution program used in step 2 is cold water, hot water, 30% ethanol, 50% ethanol, 70% ethanol gradient elution.

5. The extraction and separation method according to claim 2, characterized in that: In step 3, the volume ratio of water to methanol is 100:0, 95:5, 90:10, 80:20 and 100:0 for gradient elution.

6. The extraction and separation method according to claim 2, characterized in that: In step 4, the volume ratio of methanol to water is 100:0, 95:5, 90:10, 80:20 and 100:0 for gradient elution.

7. The extraction and separation method according to claim 2, characterized in that: The methanol elution procedure in step 5 is 20% methanol isocratic elution.

8. The extraction and separation method according to claim 2, characterized in that: The pretreatment process of ODS and dextran gel is: soaking in methanol for 24 hours, loading on the column, washing with methanol until there is no turbidity when dripping into water, and then balancing with the initial mobile phase.

9. The extraction and separation method according to claim 2, characterized in that: The volume ratio of methanol to water in the methanol:0.1% formic acid isocratic elution used in step 6 is 45:

55.

10. A use of the new amide alkaloid compound as claimed in claim 1, characterized in that: The amide alkaloid compound can be used to prepare anti-inflammatory and anti-cholinesterase drugs or health products.

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