A dihydroisoquinoline alkaloid compound from Portulaca oleracea and its extraction and separation method and use

Abstract

The present invention belongs to the field of traditional Chinese medicine extraction and separation technology, and in particular relates to a first-class dihydroquinoline alkaloid compound extracted, separated and identified from Portulaca oleracea and its extraction and separation method. The first-class dihydroquinoline alkaloid compound has a molecular formula of C 10 H 11 NO3, named 6,7‑dihydroxy‑3,4‑dihydroisoquinoline‑2(1 H )-carbaldehyde. A method for extracting and isolating the above-mentioned first-class compound is also provided, which is sequentially separated, purified and prepared by water decoction extraction, macroporous resin column chromatography, ODS column chromatography and high performance liquid chromatography. Its structure is determined to be 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1 H The first compound has anti-inflammatory activity, anticholinesterase activity and antioxidant activity. The first compound of the present invention and its salt or derivative can be used as raw materials for drug development and pharmacological activity research.

Description

Technical Field

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

[0002] Purslane is derived from the dried aerial parts of Portulaca oleracea L., a member of the Portulacaceae family. Used as a traditional Chinese medicine for thousands of years, it is widely distributed, abundantly available, and possesses exceptional adaptability and tenacious vitality. The 2020 edition of the Pharmacopoeia of the People's Republic of China lists the dried aerial parts of Portulaca oleracea as a medicinal herb. It has a sour and cold flavor and enters the liver and large intestine meridians. It has the effects of clearing heat and detoxifying, cooling blood and stopping bleeding, and stopping dysentery. It is used to treat heat-toxic bloody dysentery, carbuncles, furuncles, eczema, erysipelas, snake and insect bites, blood in stool, hemorrhoids, and metrorrhagia.

[0003] Portulaca oleracea contains a variety of chemical components, including alkaloids, flavonoids, organic acids, terpenes, coumarins, polysaccharides, amino acids, and minerals. Among these, purslane contains a particularly large variety of alkaloid compounds, including uracil, adenine, norepinephrine, dopamine, adenosine, N,N-dicyclohexylurea, purslanamide, allantoin, N-trans-feruloyltyramine, p-hydroxyphenylethylamine, oleracein AE, oleraindole AG, oleraisoindole, and portulaceramide A. The chemical composition of purslane is closely related to its diverse pharmacological effects. Modern pharmacological research has shown that purslane exhibits antioxidant, neuroprotective, anti-inflammatory, antibacterial, anti-tumor, hypoglycemic, hepatoprotective, and immune-enhancing properties. There are many kinds of chemical components in Purslane and their pharmacological activities are diverse. The development and separation of compounds in Purslane can provide a basis for in-depth research on Purslane. Summary of the Invention

[0004] To address the above problems, the present invention provides a dihydroisoquinoline alkaloid compound extracted from Portulaca oleracea. Research has found that the dihydroisoquinoline alkaloid compound of the present invention has anti-inflammatory activity, anticholinesterase activity and antioxidant activity. At the same time, a simple, rapid, environmentally friendly and high-purity extraction and separation method for the compound of the present invention is provided.

[0005] To achieve the above object, the present invention provides a dihydroisoquinoline alkaloid compound having the molecular formula C 10 H 11 NO3, named 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde according to its structure, has the following chemical formula:

[0006]

[0007] The present invention also provides a method for extracting and separating the dihydroisoquinoline alkaloid compound 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde, comprising the following steps:

[0008] Step 1: Take dried purslane, decoct in water and extract, concentrate the extract, cool to room temperature, and set aside the medicinal solution.

[0009] Step 2: The concentrated solution in step 1 is subjected to macroporous resin, and eluted with water and ethanol of different concentrations. The 30% ethanol portion is recovered under reduced pressure to obtain an extract, and the concentrate is set aside.

[0010] Step 3: Separate the concentrate in step 2 through an ODS column (Octadecylsilyl, octadecylsilane bonded silica gel filler) using a methanol-water gradient elution, detect by thin layer chromatography, develop color, combine the 30% methanol elution fractions and evaporate to dryness to obtain a concentrate for use.

[0011] Step 4: The concentrate obtained in step 3 is further separated by ODS column chromatography, using a methanol-water gradient elution, detected by thin layer chromatography, and color developed. The 50% methanol elution portion is combined and evaporated to dryness to obtain a concentrate for use.

[0012] Step 5: The concentrate obtained in step 4 is separated and prepared by HPLC (high performance liquid chromatography) using acetonitrile-0.1% formic acid (volume percentage) as the mobile phase for isocratic elution to finally obtain the dihydroisoquinoline alkaloid compound 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention.

[0013] Furthermore, in step 1, the extraction is performed twice with water decoction, each time for 2 hours, and the amount of water used is 8 to 16 times the amount of the medicinal material.

[0014] Furthermore, in step 2, the macroporous resin is AB-8 macroporous resin, and the volume ratio of ethanol to water is 30:70, 50:50, 70:30 and 100:0 in the elution gradient.

[0015] Furthermore, in step 3, the volume ratio of methanol to water is 10:0, 30:70, 50:50, 70:30 and 100:0 elution gradient; and the ODS particle size is 40-70 μm.

[0016] Furthermore, in step 4, the volume ratio of methanol to water is 10:0, 30:70, 50:50, 70:30 and 100:0 elution gradient; and the ODS particle size is 40-70 μm.

[0017] Furthermore, the ODS pretreatment process in step 3 and step 4 is: soaking in methanol for 24 hours, after loading on the column, washing with methanol until there is no turbidity when dripping into water, and then balancing with the initial mobile phase.

[0018] Furthermore, in step 5, the volume ratio of acetonitrile-0.1% formic acid is 15:85, and the retention time of the compound is 3.979 min.

[0019] The present invention also provides a use of the 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde isolated from the purslane medicinal material in the preparation of anti-inflammatory drugs, anticholinesterase drugs and antioxidant drugs.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] The isolation and pharmacological activity research of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde from purslane in the present invention have not been reported in purslane. The present invention provides a dihydroisoquinoline alkaloid compound derived from purslane and a method for extracting and separating the compound of the present invention. The method sequentially uses water decoction extraction, macroporous resin column chromatography, ODS column chromatography and high performance liquid chromatography for separation, purification and preparation to successfully extract and separate a dihydroisoquinoline alkaloid compound. The method has only five steps and is simple and rapid. The extraction is carried out with water, which is environmentally friendly. The compound separated by the method has a high purity of greater than 90%. In addition, studies have shown that the above compound has anti-inflammatory activity, anticholinesterase activity and antioxidant activity. Therefore, the isolated compound 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde and its salts and derivatives can be used as raw materials for drug development and pharmacological activity research. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention 1 H-NMR spectrum.

[0023] Figure 2 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention 13 C-NMR spectrum.

[0024] Figure 3 This is the DEPT spectrum of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention.

[0025] Figure 4 This is the HSQC spectrum of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention.

[0026] Figure 5 This is the HMBC spectrum of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention.

[0027] Figure 6 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention 1 H- 1 H COSY spectrum.

[0028] Figure 7 This is the ROESY spectrum of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention.

[0029] Figure 8 This is a high-resolution mass spectrum of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention. DETAILED DESCRIPTION

[0030] The following examples will help to understand the present invention, but these examples are only for illustration of the present invention and the present invention is not limited to these contents. The operating methods in the examples are all conventional operating methods in the art.

[0031] Example 1 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde.

[0032] The present invention provides a dihydroisoquinoline alkaloid compound, the molecular formula of which is C 10 H 11NO3, named 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde, has the chemical formula:

[0033]

[0034] Table 1 shows the NMR data of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde. The solvent used for NMR was CD3OD.

[0035] Table 1 NMR data of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde.

[0036]

[0037] 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde: brown powder, easily soluble in methanol, slightly soluble in water. After being spotted on a silica gel thin layer plate, the spots will appear orange when sprayed with dilute potassium bismuth iodide test solution, indicating that the compound is an alkaloid component. 1 H-NMR, 13 C-NMR and HR-ESI-TOF-MS signals suggest that the possible molecular formula of the compound is C 10 H 11 NO3, unsaturation degree is 6. HR-ESI-TOF-MS gives m / z 194.0817 [M+H] + The quasi-molecular ion peak of is 194.0817. 1 In H-NMR, δ H 8.16 (1H, s, H-9) suggests that there may be aldehyde hydrogen in the structure, δ H 6.57 (1H, m, H-4) and δ H 6.57 (1H, m, H-7) suggests the presence of two aromatic hydrogens in the structure, δ H 4.50(2H,s,H-8),δ H 3.67(2H,t,J=6Hz,12Hz,H-2),δ H 2.76 (2H, t, J = 6 Hz, 12 Hz, H-3) suggests the presence of three methylene groups in the structure. HMBC shows that H-4 and C-6 (δ C 143.97), C-7a(δ C 122.09), H-7 and C-5 (δ C144.06), C-3a(δ C 124.57), indicating the presence of a tetrasubstituted benzene ring structure in the structure. Since C-5 and C-6 are located in the low field region, it is speculated that the C-5 and C-6 positions are substituted with hydroxyl groups; H-2 is closely related to C-3a, C-8 (δ C 31.40), H-3 is related to C-7a, H-8 is related to C-2 (δ C 43.54) and C-3a. Since C-2 and C-8 are located in the low-field region, it is speculated that C-2 and C-8 are connected to heteroatoms, suggesting the presence of a piperidine ring structure, which shares C-3a and C-7a with the benzene ring. Furthermore, H-3 is related to C-4, and H-8 is related to C-7, further indicating that the piperidine ring is connected to the benzene ring, indicating the presence of dihydroquinoline in the structure. H-9 is related to C-2 and C-8, indicating that the aldehyde group is connected to the nitrogen of the piperidine ring. Combined with the molecular weight of the HR-ESI-TOF-MS spectrum, this first-isolated compound can be confirmed to have the above structure. The spectrum shows double signals due to isomerization caused by rotation of the C-N bond of formamide.

[0038] The present invention also provides a method for extracting and separating 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde, which specifically comprises the following steps:

[0039] Step 1: Weigh 250 kg of dried purslane medicinal materials, use water decoction to extract, the amount of which is 10 times that of the medicinal materials, extract twice, each time for 2 hours, combine the extracts, heat and concentrate, cool to room temperature, and obtain the medicinal solution for use.

[0040] Step 2: The medicinal solution obtained in step 1 is evaporated to dryness and separated by chromatography on an AB-8 macroporous resin column using an ethanol-water (30:70, 50:50, 70:30 and 100:0, v:v) gradient elution, collecting the 30% ethanol portion, and recovering it under reduced pressure to obtain an extract to obtain a concentrate for later use.

[0041] Step 3: The concentrate in step 2 was separated by a pretreated ODS column with a filler particle size of 40 to 70 μm, and gradient elution was performed using methanol-water (10:0, 30:70, 50:50, 70:30 and 100:0, v:v). The mixture was detected by thin layer chromatography and color was developed. The 30% methanol elution fractions were combined, concentrated to dryness under reduced pressure, and set aside.

[0042] Step 4: The product obtained in step 3 is further separated by pre-treated ODS column chromatography using a filler with a particle size of 40-70 μm. The product is eluted with a gradient of methanol-water (10:0, 30:70, 50:50, 70:30, and 100:0, v / v). Thin layer chromatography is performed for color development. The 50% methanol elution fractions are combined and concentrated to dryness under reduced pressure for later use. The ODS pre-treatment process is as follows: soaking in methanol for 24 hours. After loading the column, the column is washed with methanol until there is no turbidity when dripped into water, and then equilibrated with the initial mobile phase.

[0043] Step 5: The fraction obtained in step 4 was separated and prepared by HPLC using acetonitrile and 0.1% formic acid in a volume ratio of 15:85 as the mobile phase and detection wavelengths of 210 and 254 nm to obtain the new natural product 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention; the purity was determined to be 95% by normalization method.

[0044] Example 2 Anti-inflammatory effect of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde.

[0045] 1 Main materials.

[0046] 1.1 Drugs and Reagents: The new natural product used in this experiment was prepared by the above-mentioned method and had a purity of 95%. DMEM high-glucose medium and fetal bovine serum were obtained from Hyclone (USA); penicillin and streptomycin were obtained from Hangzhou Sijiqing Company; LPS was obtained from Sigma (USA); an IL-1β ELISA kit was obtained from Cayman (USA); and cell lysate was used.

[0047] 1.2 Cell line: RAW264.7 macrophages (ATCC cell bank, USA).

[0048] 1.3 Grouping: Divided into control group, LPS group and experimental group.

[0049] 2 Experimental methods.

[0050] 2.1 Cell culture: Add 10% fetal bovine serum and 1% antibiotics (100 U / mL penicillin and 100 μg / mL streptomycin) to DMEM high-glucose medium and store at 4°C.

[0051] 2.2 CCK-8 assay for cell viability: The three groups of RAW264.7 macrophages in logarithmic growth phase were inoculated into 96-well culture plates at a cell density of 1×10 4 / mL, 100μL per well, 37 ℃, 5% CO2 conditions after overnight incubation, the experimental group added different concentrations of the compound of the present invention 6,7-dihydroxy-3,4-dihydroisoquinoline-2 (1H) -carbaldehyde (5μM ~ 50μM), incubation for 1 hour, LPS at a concentration of 1μg / mL was added to the LPS group and experimental group, respectively, and a zero adjustment group (culture medium containing DMSO solvent) was set up. Each group had 3 replicates to investigate the effect of drug addition on the cells. After 24 hours of cell culture in each group, 10μL of CCK-8 was added to each well of cells, and incubation continued for 4 hours at 37 ℃, 5% CO2 conditions. The absorbance of each well was measured at a wavelength of 450nm using a microplate reader.

[0052] 2.3 ELISA assay for inflammatory factor IL-1β: RAW264.7 macrophages in the logarithmic growth phase were seeded in 24-well culture plates at a cell density of 1×10 5 Cells were incubated overnight at 37°C in 5% CO2 with 1 mL per well. The experimental group was treated with the compound of this invention, 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde (1 μM to 20 μM). After incubation for 1 hour, LPS (final concentration of 1 μg / mL) was added to each well and incubated for 24 hours. Each treatment was repeated in triplicate. IL-1β levels were measured by ELISA.

[0053] 3 Experimental results.

[0054] The experimental results show that the compound 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention has no effect on the proliferation of LPS-induced macrophages RAW264.7; it can effectively inhibit the secretion of excessive inflammatory cytokine IL-1β produced by LPS-induced macrophages RAW264.7 in a concentration-dependent manner.

[0055] The results of the relative cell survival rate experiment are shown in Table 2.

[0056] Table 2 Effect of compound 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde on the relative survival rate of RAW264.7 macrophages.

[0057]

[0058] The results of ELISA determination of inflammatory factor IL-1β are shown in Table 3.

[0059] Table 3 Effects of compound 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde on the IL-1β content secreted by RAW264.7 cells induced by LPS (mean ± standard deviation, n = 3).

[0060]

[0061] Note: * P<0.05 compared with the control group, # P<0.05 compared with the LPS group.

[0062] Example 3 Anticholinesterase Effect of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde.

[0063] 1 Main materials.

[0064] 1.1 Drugs and Reagents: The new natural products used in this study were prepared by the above-mentioned method with a purity of 95%. Physostigmine was obtained from Shanghai Hanxiang Biotechnology Co., Ltd., acetylcholine iodide (ATCI) and acetylcholinesterase (AChE) were obtained from Dalian Meilun Biotechnology Co., Ltd., dithiodinitroformic acid (DTNB) was obtained from Shanghai Jinshui Biotechnology Co., Ltd., and sodium dihydrogen phosphate and sodium dihydrogen phosphate were obtained from Shanghai Sinopharm Reagent Co., Ltd.

[0065] 1.2 Experimental instruments and equipment: HBS-1096A 96-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.).

[0066] 2 Experimental methods.

[0067] The anticholinesterase activity of each compound was determined using a modified Ellman method. Accurately weigh 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde and physostigmine and prepare a series of five sample solutions in methanol at concentrations of 31.25 μM, 62.5 μM, 125 μM, 250 μM, and 500 μM. The specific operation is as follows: 140 μL of phosphate buffered saline (0.1 M, pH = 8.0, containing 0.1 mol / L sodium dihydrogen phosphate and sodium dihydrogen phosphate) was added to a 96-well microplate, 20 μL of sample solution, 15 μL of AChE (0.2 U / mL), and incubated at 37 ° C for 10 minutes. Then, 10 μL of ATCI (4 mmol / L) and 10 μL of DTNB (15 mmol / L) were added. After incubation at 37 ° C for 20 minutes, the 96-well plate was placed in a microplate reader and the absorbance of each group of samples was measured at a wavelength of 405 nm. 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):

[0068] Inhibition rate (%) = (A 空白 -A 样品 ) / A 空白 ×100%

[0069] 3 Experimental results.

[0070] The experimental results show that the first compound of the present invention, 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde, exhibits certain anticholinesterase activity, and the inhibitory effect on cholinesterase increases with the increase of the compound concentration, showing a dose-dependent trend.

[0071] The anticholinesterase activity of the first compound of the present invention, 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde, is shown in Table 4.

[0072] Table 4 Anticholinesterase activity of compound 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde.

[0073]

[0074] Example 4 Antioxidant Effect of 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde.

[0075] 1 Main materials.

[0076] 1.1 Drugs and Reagents: The new natural products used in the experiment were prepared by the above method with a purity of 95%. 1,1-Diphenyl-2-picrylhydrazyl free radical (DPPH) was obtained from Sigma-Aldrich, USA; butylated hydroxyanisole (BHA) was obtained from Shanghai Xiangrui Biological Co., Ltd.; and methanol (chromatographically pure, Tianjin Kaixin Chemical Industry Co., Ltd.) was used.

[0077] 1.2 Experimental instruments and equipment: Hitachi UV-3010 ultraviolet-visible spectrophotometer (Hitachi, Japan), 1 / 100,000 balance (METTLER, Switzerland).

[0078] 2 Experimental methods.

[0079] This experiment uses the DPPH free radical scavenging method to determine the antioxidant activity of each compound. Accurately weigh 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde and BHA, and use methanol to prepare five series of sample solutions with concentrations of 12.5μM, 25μM, 50μM, 100μM and 200μM. In addition, the DPPH solution is prepared as needed, and an appropriate amount of DPPH is accurately weighed and used to prepare a solution with a concentration of 80μM with methanol. The whole process is protected from light. The specific operation is as follows: 1mL of sample solution is fully mixed with 1mL of DPPH solution, and placed at room temperature in a light-proof environment for 10 minutes. After setting the detection wavelength of the ultraviolet spectrophotometer to 517nm, the absorbance value of the sample is measured. Among them, methanol is used instead of the sample solution as the blank group, BHA is used instead of the sample solution as the positive control group, and a mixed solution of 1mL methanol and 1mL sample solution is used as the control group. The DPPH scavenging rate of each compound is calculated according to the following formula (A represents absorbance):

[0080] DPPH clearance rate (%) = (1-(A 样品 -A 对照 ) / A 空白 )×100%

[0081] 3 Experimental results.

[0082] The experimental results show that the first compound 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde of the present invention exhibits certain antioxidant activity, and the antioxidant effect increases with the increase of the compound concentration, showing a dose-dependent trend.

[0083] The antioxidant activities of the first compounds of the present invention are shown in Table 5.

[0084] Table 5 Antioxidant activity of compound 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde.

[0085]

[0086] In summary, the present invention provides a compound, 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde, and a method for its extraction and isolation. Separation, purification, and preparation are performed sequentially using water decoction extraction, macroporous resin column chromatography, ODS column chromatography, and high-performance liquid chromatography. A dihydroquinoline alkaloid compound is successfully extracted and isolated. This method is simple, rapid, and environmentally friendly, and the compound isolated by this method is of high purity. Because the resulting compound is extracted from the commonly used traditional Chinese medicine Portulaca oleracea, it exhibits anti-inflammatory, anticholinesterase, and antioxidant activities. Therefore, the new natural product 6,7-dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbaldehyde, its salts, and derivatives, can provide insights into drug development and have broad development prospects.

Claims

1. A method for extracting and separating dihydroquinoline alkaloid compounds from Portulaca oleracea, characterized in that: The specific steps include: Step 1: Take dried purslane, decoct in water, concentrate the extract, and cool to room temperature to obtain a medicinal solution for later use; Step 2: The concentrated solution in step 1 is subjected to a macroporous resin, and eluted with water and ethanol of different concentrations. The 30% ethanol portion is recovered under reduced pressure to obtain an extract, and the concentrate is prepared for use; the macroporous resin is AB-8 macroporous resin, and the volume ratio of ethanol to water is 30:70, 50:50, 70:30 and 100:0 for gradient elution; Step 3: The concentrate in step 2 is separated by ODS column, eluted with methanol-water gradient, detected by thin layer chromatography, and color developed. The 30% methanol elution portion is combined and evaporated to dryness to obtain a concentrate for use; the volume ratio of methanol to water is 10:0, 30:70, 50:50, 70:30 and 100:0 for gradient elution; the ODS particle size is 40-70 μm; Step 4: The concentrate obtained in step 3 is further separated by ODS column chromatography, eluted with a methanol-water gradient, detected by thin layer chromatography, and developed. The 50% methanol elution portion is combined and evaporated to dryness to obtain a concentrate for use; the volume ratio of methanol to water is 10:0, 30:70, 50:50, 70:30 and 100:0 for gradient elution; the ODS particle size is 40-70 μm; Step 5: The concentrate obtained in step 4 was separated and prepared by HPLC, using acetonitrile-0.1% formic acid as the mobile phase for isocratic elution, wherein the volume ratio of acetonitrile-0.1% formic acid was 15:85, and the compound retention time was 3.979 min; finally, the dihydroquinoline alkaloid compound was obtained; The chemical structural formula of the dihydroquinoline alkaloid compound is .

2. The extraction and separation method according to claim 1, wherein In step 1, the extraction is performed by decocting in water twice, each time for 2 hours, and the amount of water used is 8 to 16 times the amount of the medicinal material.

3. A method for separating dihydroquinoline alkaloid compounds from Portulaca oleracea according to claim 1, characterized in that: The dihydroquinoline alkaloid compound is used for preparing anticholinesterase drugs and antioxidant drugs.

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