CN110294733B - Peroxide bond-containing compound Oleracone I in purslane, and extraction separation method and application thereof - Google Patents

Abstract

The invention relates to the field of extraction and separation of traditional Chinese medicines, in particular to a novel compound extracted, separated and identified from a purslane medicinal material, an extraction and separation method and application thereof, and specifically relates to a compound Oleracone I containing a peroxide bond in purslane, and an extraction and separation method and application thereof. The extraction and separation method of the new compound adopts water decoction extraction, silica gel column chromatography, polyamide column chromatography, ODS medium-pressure column and Sephadex LH-20 separation in sequence, and a new compound containing a peroxide bond is successfully extracted and separated. The structure of the compound is identified as a new compound containing a peroxide bond by a method of mass spectrum, carbon spectrum, hydrogen spectrum and two-dimensional nuclear magnetic spectrum analysis. The new compound has the functions of resisting tumor and inflammation, and the new compound and the salt or the derivative thereof can be used as a lead for synthesizing other compounds and raw materials for developing new drugs and researching pharmacological activity and can be used for preparing anti-tumor and anti-inflammation drugs.

Description

Peroxide bond-containing compound Oleracone I in purslane, and extraction separation method and application thereof

Technical Field

The invention relates to the field of extraction and separation of traditional Chinese medicines, in particular to a novel compound extracted, separated and identified from a purslane medicinal material and an extraction and separation method thereof, and specifically relates to a compound Oleracone I containing a peroxide bond in purslane, and an extraction and separation method and application thereof.

Background

Purslane (purslane)Portulaca oleraceaL.), herba Portulacae, herba Peperomiae Longipedunculatae, and Formica Fusca, which are annual herbaceous plants of Portulacaceae. Purslane is widely distributed and rich in resources, and is one of 78 wild plants which are homologous in medicine and food and specified by the Ministry of health of China. Purslane is recorded in 2015 edition of pharmacopoeia of the people's republic of China, has the effects of clearing heat and removing toxicity, cooling blood and stopping bleeding, stopping dysentery and the like, and is used for treating heat-toxin bloody dysentery, carbuncle, furuncle, eczema, erysipelas, snake and insect bite, hematochezia, hemorrhoidal bleeding, metrorrhagia and metrostaxis and the like.

Modern pharmacological research of purslane shows that the purslane has the effects of resisting inflammation, bacteria and viruses, reducing blood pressure, reducing blood fat, resisting oxidation, resisting cancer and tumors, relaxing skeletal muscles and smooth muscles, regulating immune function and the like. The main chemical components of purslane comprise flavonoids, coumarins, terpenoids, steroids, alkaloids, amino acids, lignans, volatile oil, polysaccharides, various pigments, minerals and the like, which provide a material basis for the various pharmacological actions of the purslane. Wherein alkaloids are a main chemical component in purslane, and the alkaloid components reported at present comprise norepinephrine, dopamine, a small amount of dopa, adenosine, uracil, adenine, N-dicyclohexylurea, allantoin and N-trans-feruloyltyramine; cyclic dipeptide alkaloids and alkaloids are also present: oleracein A-I, K, L, N-S.

Most of the chemical components separated from purslane are known and have low structural novelty, so the development and separation of new compounds in purslane are urgently needed.

Disclosure of Invention

Aiming at the problems, the invention provides a compound Oleracone I containing a peroxide bond in purslane and an extraction and separation method thereof, in particular to a new compound extracted from purslane.

In order to achieve the above object, the present invention provides the following technical solutions.

The purslane contains a peroxy bond compound Oleracone I with a molecular formula of C₁₈H₁₈O₆Named as oleracene I, the chemical structural formula is as follows:

。

a method for extracting and separating a compound Oleracone I containing a peroxide bond in purslane comprises the following specific steps.

Step 1, taking a purslane dried medicinal material, extracting twice with water (the amount of water is 8-16 times of the medicinal material), filtering the water extract, combining the filtrates, directly heating and concentrating, and cooling to room temperature to obtain a liquid medicine for later use.

And 2, evaporating the liquid medicine obtained in the step 1 to dryness, putting the liquid medicine on a silica gel column, eluting the liquid medicine by using ethyl acetate, and recovering the ethyl acetate under reduced pressure to obtain an extract so as to obtain an ethyl acetate extract.

And 3, separating the ethyl acetate extract obtained in the step 2 by using a polyamide column, performing gradient elution by using ethanol-water, evaporating 50% ethanol to dryness, then putting the ethyl acetate extract on a silica gel column, sequentially performing gradient elution by using ethyl acetate-methanol to obtain a plurality of elution parts, detecting by using a thin-layer chromatography, developing, combining the developed elution parts, and concentrating the combined elution parts under reduced pressure until the combined elution parts are dry for later use.

And 4, carrying out chromatographic separation on the product obtained in the step 3 by using a pretreated ODS (octadecylsilane chemically bonded silica) column, carrying out gradient elution by using methanol-water to obtain a plurality of elution parts, detecting by using a thin-layer chromatography, developing, and concentrating each developed elution part under reduced pressure until the elution parts are dried to obtain a concentrate for later use.

And 5, carrying out chromatographic separation on the product obtained in the step 4 by using a pretreated Sephadex LH-20 column, eluting by using methanol to obtain a plurality of elution parts, detecting by using a thin-layer chromatography, developing, combining the developed elution parts, and concentrating the combined elution parts under reduced pressure until the elution parts are dried for later use.

And 6, separating and preparing the new compound obtained in the step 5 by HPLC (high performance liquid chromatography), and carrying out isocratic elution by using acetonitrile-0.1% formic acid as a mobile phase to finally obtain a compound oleracene I.

The pretreatment process of the ODS and the sephadex comprises the steps of soaking in methanol for 24 hours, loading on a column, washing with methanol until no turbidity exists in dropping water, and then balancing with an initial mobile phase.

Compared with the prior art, the invention has the beneficial effects.

The separation and pharmacological activity research of the novel compound containing the peroxide bond in the purslane is not reported in the journal of the prior paper; the invention provides a novel compound containing peroxide bond from purslane and an extraction and separation method aiming at the novel compound, which sequentially adopts water decoction extraction, silica gel column chromatography, polyamide column, ODS medium-pressure column, Sephadex LH-20 and high performance liquid chromatograph for separation, purification and preparation to successfully extract and separate out the novel compound, the method has the advantages of six steps, simple and rapid operation method, environment-friendly process method, and mainly adopts water extraction and ethyl acetate elution in the extraction and separation process, the purity of the compound obtained by the separation by the method is higher than 90 percent, in addition, the research shows that the compound has the functions of anti-tumor and anti-inflammatory, therefore, the novel compound and the salt and the derivative thereof can be used as a precursor for synthesizing other compounds, and raw materials for new drug development and pharmacological activity research, and can also be used for preparing anti-tumor and anti-inflammatory drugs.

Drawings

FIG. 1 is a UV spectrum of the novel compound, oleracene I.

FIG. 2 is an infrared spectrum of the novel compound oleracene I.

FIG. 3 is a high resolution mass spectrum of the novel compound, oleracene I.

FIG. 4 shows the novel compound oleracene I¹H-NMR spectrum chart.

FIG. 5 shows the novel compound oleracene I¹³C-NMR spectrum chart.

FIG. 6 is a nuclear magnetic resonance carbon spectrum (DEPT) spectrum of the novel compound oleracene I.

FIG. 7 shows the NMR of the novel compound, oleracene I¹H-¹H COSY spectrogram.

FIG. 8 is a nuclear magnetic resonance HMBC spectrum of the novel compound oleracene I.

FIG. 9 is a diagram of the nuclear magnetic resonance HSQC spectrum of the novel compound oleracee I.

FIG. 10 is a NOESY spectrum of the novel compound, oleracene I.

Detailed Description

Examples are given.

The present invention provides novel compounds of formula C₁₈H₁₈O₆Named as oleracene I, the chemical formula is:

。

the new compound is named oleracene I according to the structure, and table 1 is the nuclear magnetic data of the new compound:¹H-NMR of¹³C-NMR in deuterated DMSO.

TABLE 1 NMR data for novel Compounds: (¹H-NMR of¹³C-NMR in deuterated DMSO).

Structural characterization of the compounds of the invention is illustrated in FIGS. 1-10.

Oleracone I: yellowish brown powder, readily soluble in methanol. UV (MeOH) lambda_max: 284, 214 nm。IR (KBr) v _max 2920, 2850, 2360, 1670, 1610, 1460, 1215, 1160, 831, 762 cm^-1. HRESI (+) TOFMS gave M/z 331.1141 [ M + H]⁺Has an excimer ion peak of 331.1137 molecular weight. Bonding of¹H-NMR，¹³C-NMR and DEPT data, presuming that the possible molecular formula of the compound is C₁₈H₁₈O₆The unsaturation degree was 10.¹³The C-NMR spectrum and the DEPT spectrum showed 18 carbon signals, respectively 2 OCH₃（δ: 55.7; 55.9), 3 CH₂（δ: 33.2; 72.1; 72.5), 6 alkenylcarbons: (δ: 92.9; 93.3; 115.0; 118.6; 127.5; 132.4), 7 quaternary carbons (5 double-bonded carbons to O,δ: 189.9, respectively; 165.4 of the total weight of the mixture; 163.9, respectively; 162.3; 155.7; 2 of the carbon atoms of the double bond,δ：122.1；103.5）。

¹H-NMR spectrum showed 2 CH₃Signals are respectivelyδ3.78（3H，s）；δ3.80 (3H, s); 3 methylene signals, δ 2.89(2H, dd, J =13.8,20.5), respectively; δ 4.01(1H, d, J =11.64), δ 4.07(1H, d, J = 11.5); δ 4.10(2H, d, J = 4.44); the 6 methine signals were δ 6.12(1H, d, J =2.28), respectively; δ 6.20(1H, d, J = 2.28); δ 6.69(1H, m); δ 6.76(1H, dd, J =0.78, 7.92); δ 7.02(1H, m); δ 7.11(1H, dd, J = d =)1.5,7.5). According to the H-H COSY spectrum, two methine groups delta 6.12 and delta 6.20 are coupled, four methine groups delta 06.69, delta 16.76, delta 2,7.02 and delta 37.11 are coupled, and the existence of two benzene rings is indicated. According to the correlation peak of the HMBC spectrum, H delta 3.78 in the methoxyl is coupled with C-4 ʹ, H delta 3.80 is coupled with C-2 ʹ, and C-4 ʹ (delta 162.3) and C-2 ʹ (delta 165.4) are positioned in a low field region, indicating that the connection with O is realized, and the two methoxyl are respectively connected with C-4 ʹ and C-2 ʹ on a benzene ring; according to NOE spectrum, H in two methoxy groups is coupled with H-1 ʹ, H-3 ʹ, which indicates that H is associated with C-1 ʹ and C-3 ʹ; h-1 ʹ was coupled to C-3, and C-3 (delta 163.9) was located in the low field region, indicating a connection to O; h-1 ʹ and H-3 ʹ were coupled to C-4, H-1 ʹ and H-5 were coupled to C-3, and C-5 (. delta.72.5) was located in the low field region, suggesting a link to O, indicating that C-4 and 6-O are linked through a C-5 methylene group; h-5 is coupled to C-9, H-7 is coupled to C-8 carbonyl C and C-9, and C-7 (delta 72.1) is located in the low field region, suggesting a connection to O, while H-9 is coupled to C-8 carbonyl C and to C-7, indicating that C-7 is connected to O at position 6 and to C-9 through C-8 carbonyl C; h-1 ʹ ʹ, H-2 ʹ ʹ and H-3 ʹ ʹ are all coupled to C-10, H-9 is coupled to C-10 and C-4 ʹ ʹ, H-4 ʹ ʹ is coupled to C-9, indicating that C-9 is connected to another benzene ring; h-1 ʹ ʹ, H-2 ʹ ʹ, H-3 ʹ ʹ and H-4 ʹ ʹ are all coupled to C-11, and C-11 (delta 155.7) is located in the low field region, indicating a connection to O; according to the NOE spectrum, the methines on the two benzene rings are homogeneously coupled and [ M + H ] is given by HRESI (+) TOFMS]⁺The quasi-molecular ion peak proves that the O at the 1 position is connected with the O at the 2 position to form a peroxy bond. From the above information, the novel compounds can be identified as having the above structure.

The invention also provides an extraction and separation method of the compound, which comprises the following specific steps.

Step 1: weighing 150kg of dry purslane medicinal material, extracting with water under reflux, wherein the water dosage (v/v) is 10 times of that of the medicinal material, extracting under reflux twice, each time for 2h, heating and concentrating at 90-100 ℃, and cooling to room temperature to obtain liquid medicine for later use.

Step 2: evaporating the liquid medicine obtained in the step 1 to dryness, performing chromatographic separation by using a silica gel column, isocratically eluting by using ethyl acetate (115L), wherein the silica gel is 100-200 meshes, and recovering the ethyl acetate to obtain an extract under reduced pressure below 40 ℃ to obtain an ethyl acetate extract.

And step 3: separating the ethyl acetate extract obtained in the step 2 by using a polyamide column, performing gradient elution by using ethanol-water (0/100, 30/70, 50/50, 70/30, 100/0, v/v), evaporating to dryness at 90-100 ℃ by using 50% ethanol, performing chromatographic separation by using a silica gel column, wherein the silica gel is 200-300 meshes, performing gradient elution by using ethyl acetate-methanol (5: 1, 2:1, 1:2, v: v) in sequence to obtain 19 parts (namely 19 bottles are obtained in total and 300 mL in each bottle), detecting by using a thin-layer chromatography, developing, combining the developed 1-14 elution parts, and concentrating the combined 1-14 parts at the temperature of below 40 ℃ under reduced pressure until the parts are dried for later use.

And 4, step 4: and (3) performing pretreated ODS (octadecylsilane chemically bonded silica) medium-pressure column chromatography separation on the product obtained in the step (3), wherein the granularity of the filler is 20-40 mu m, performing gradient elution (pressurizing to enable the flow rate to be 1mL/min and the temperature to be room temperature) by using methanol-water (50/50, 60/40, 70/30, 80/20, 100/0, v/v) to obtain 16 parts (namely performing gradient elution to obtain 16 bottles, wherein each bottle is 100 mL), detecting by using thin-layer chromatography, developing, retaining 7-9 parts for developing, and concentrating under reduced pressure below 50 ℃ until the parts are dry for later use.

And 5: and (3) subjecting the product obtained in the step (4) to chromatographic separation by a pretreated Sephadex column (Sephadex LH-20), eluting by methanol to obtain 26 elution parts (namely, 26 bottles are obtained in total and each bottle is 50 mL), detecting by thin-layer chromatography, developing, reserving the developed 5-11 parts, and concentrating under reduced pressure below 50 ℃ until the parts are dry for later use to obtain a new compound. The pretreatment process of the ODS and the sephadex comprises the steps of soaking in methanol for 24 hours, loading on a column, washing with methanol until no turbidity exists in dropping water, and then balancing with an initial mobile phase.

Step 6: the new compound obtained in step 5 was prepared by HPLC separation with acetonitrile: 0.1% formic acid (40: 60, v/v) is used as a mobile phase, the detection wavelength is 210nm and 280nm, the new compound is obtained by separation and preparation, and the purity measured by a normalization method is 90-99%.

The compounds of the present invention have anti-tumor effects.

1 main material.

1.1 drugs and reagents: the new compound used in the experiment is prepared by the method, the purity of the new compound is 90-99%, the new compound is precisely weighed and diluted by DMSO to be a solution required by each dosage group. DMEM high-glucose medium, fetal bovine serum (Hyclone, usa); penicillin and streptomycin (Hangzhou Sijiqing Co., Ltd.).

1.2 cell lines: human colon cancer cell Caco-2, human breast cancer cell MCF-7, human gastric cancer cell BGC-823, human lung adenocarcinoma cell SPC-A1, human liver cancer cell BEL-7402, human cervical cancer cell Hela-229, ovarian cancer cell Ho-8910, and human oral epidermoid carcinoma cell KB (Shanghai cell Bank of China academy of sciences).

1.3 grouping: divided into a control group, an experimental group and a zero-adjustment group (culture solution containing DMSO solvent).

2 experimental methods.

2.1 cell culture, DMEM high sugar medium, added with l 0% fetal bovine serum, l% antibiotics (100U/mL penicillin and 100. mu.g/mL streptomycin), placed at 37 ℃ with 5% CO₂Culturing in an incubator.

2.2 MTI method for detecting cell proliferation, inoculating cells in logarithmic growth phase into 96-well culture plate with cell density of 1 × 104/mL, 100 μ L per well, temperature of 37 deg.C, and 5% CO₂After overnight culture under the conditions, the experimental groups were added with the novel compounds of the present invention at different concentrations, each group was provided with 3 multiple wells, and after adding the drug, the mixture was placed at 37 ℃ and 5% CO₂Culturing in an incubator for 48 h. Absorbing the culture solution containing the medicine, and adding the mixture into the culture solution in a volume ratio of 4: 1 and MTT (5 mg/mL) for 4 hours, carefully absorbing the supernatant, adding 150 mu L of DMSO into each hole, placing the hole on a shaker to shake so as to completely dissolve crystals (5 min), and detecting the absorbance (A) value of each hole by a microplate reader at the wavelength of 570 nm. Then, the inhibition rate of each concentration of compound on cell growth is calculated, and the inhibition rate formula is as follows: cell growth inhibition rate = (1-A)_{Medicine feeding hole}／A_{Control well}) X 100%, processing data with SPSS software, plotting inhibition rate against drug concentration, and calculating IC₅₀The value is obtained.

3, experimental results.

The experimental result shows that the novel compound containing peroxy bond is fine for human colon cancerThe cell Caco-2, the human breast cancer cell MCF-7, the human gastric cancer cell BGC-823, the human lung adenocarcinoma cell SPC-A1, the human liver cancer cell BEL-7402, the human cervical cancer cell Hela-229, the ovarian cancer cell Ho-8910 and the human oral epidermoid cancer cell KB have an inhibitory effect on the proliferation, and the inhibitory rate is obviously increased along with the increase of the drug concentration, namely the concentration is dependent. The novel compound of the invention is used for treating the eight tumor cells IC₅₀The values are shown in Table 5.

Table 2 inhibitory effect of the novel compounds of the present invention on tumor cells.

The novel compounds of the present invention have anti-inflammatory properties.

1. The main material.

1.1, drugs and reagents: the new compound used in the experiment is prepared by the method, the purity of the new compound is 90-99%, the new compound is precisely weighed and diluted by DMSO to be a solution required by each dosage group. DMEM high-glucose medium, fetal bovine serum (Hyclone, usa); penicillin, streptomycin (Hangzhou Sijiqing Co.); LPS (Sigma, usa); IL-6, TNF-alpha, PGE₂ELISA kit of (A) (Cayman, USA); cell lysate, Griess reagent (bi yun tian biotechnology limited).

1.2 cell lines: RAW264.7 macrophages (us ATCC cell bank).

1.3 grouping: the test group was divided into a control group, an LPS group and an experimental group.

2 experimental methods.

2.1 cell culture, DMEM high-sugar medium, with addition of l 0% fetal bovine serum, l% antibiotics (100U/mL penicillin and 100. mu.g/mL streptomycin), 37.5% CO₂Culturing in an incubator.

2.2 MTT colorimetric method for determining cell viability, inoculating RAW264.7 macrophage in logarithmic growth phase into 96-well culture plate respectively, wherein cell density is 1 × 10⁴one/mL, 100. mu.L per well, temperature 37 ℃, 5% CO₂After overnight incubation under conditions, the experimental groups were added with books of different concentrationsThe new compound Oleracone I (1-50 mu M) is incubated for 1h, then LPS with the final concentration of 1 mu g/mL is added to an LPS group and an experimental group respectively, a zero-adjusting group (culture solution containing DMSO solvent) is arranged, 3 multiple holes are arranged in each group, and the influence on cells after the addition of the drug is examined. After culturing the above groups of cells for 24 hours, 20. mu.L of MTT 5 mg/mL was added to each well of cells at 37 ℃ with 5% CO₂After incubation for 4h, terminating the culture, absorbing the liquid in the wells, adding 100 μ L of dimethyl sulfoxide (DMSO) into each well, oscillating for 10min to dissolve the intracellular crystal, and measuring the light absorption value of each well at 570nm wavelength of an enzyme-labeling instrument.

2.3 measurement of NO content by Griess method, the inhibitory effect of the novel compound of the present invention on the NO production of LPS-induced mouse macrophage RAW264.7 was examined. Mouse macrophage RAW264.7 passage, culturing in high glucose cell culture medium DMEM containing 10% fetal calf serum, adding the new compound Oleracone I (1-20 μ M) of the invention at different concentrations, and culturing at 37 deg.C and 5% CO₂After incubation for 1h under conditions, inflammatory responses were induced with LPS (final concentration 1. mu.g/mL), and after 24h supernatants were collected and 3 wells were repeated for each group. The Griess method measures the content of NO in cell supernatant, and the influence of the novel compound on the NO release of LPS-induced RAW264.7 cells is reflected according to different concentrations so as to reflect the NO level.

2.4 measurement of inflammatory factors IL-6, TNF-alpha and inflammatory mediators PGE by ELISA₂: RAW264.7 macrophages in logarithmic growth phase were seeded in 24-well culture plates at a cell density of 1X 10⁵one/mL, 1mL per well, temperature 37 ℃, 5% CO₂After incubation overnight under these conditions, the experimental groups were incubated with the novel compound Oleracone I (1-20. mu.M) for 1h, LPS (final concentration of 1. mu.g/mL) was added to each well, and incubated for 24h, with 3 replicates per group treatment. ELISA method for measuring IL-6, TNF-alpha and PGE secreted by RAW264.7 macrophage after treatment of purslane-derived novel compound₂The content of (a).

3, experimental results.

The experimental result shows that the novel compound containing the peroxy bond has no influence on the proliferation of macrophage RAW264.7 induced by LPS, and is safe and nontoxic; and can effectively inhibit macrophage RAW264.7 induced by LPSThe produced excessive inflammatory cytokines IL-6, TNF-alpha and inflammatory mediators NO and PGE₂And is concentration dependent.

The results of the cell relative survival experiments are shown in table 3.

Table 3 effect of the invention on relative survival of RAW264.7 macrophages.

Note:^*P<0.05 compared with the control group (significant difference in the high concentration group), the results of the experiments for determining the NO content by Griess method are shown in Table 4.

Table 4 effect of the invention on LPS-induced release of NO from RAW264.7 cells (mean ± standard deviation, n = 3).

Note:^*P<0.05 compared with the control group,^#P<0.05 compared to the LPS group.

ELISA method for measuring inflammatory factors IL-6, TNF-alpha and inflammatory mediator PGE₂The results are shown in Table 5.

TABLE 5 IL-6, TNF-alpha and PGE secreted by RAW264.7 cells induced by LPS according to the invention₂Influence of content (mean ± standard deviation, n = 3).

Note:^*P<0.05 compared with the control group,^#P<0.05 compared to the LPS group.

In conclusion, the invention provides the special compound and the extraction and separation method thereof, the new compound is successfully separated and obtained by sequentially adopting water reflux extraction, silica gel column chromatography, polyamide column chromatography, ODS medium-pressure column chromatography and sephadex column chromatography for separation and purification, the method is simple, convenient, rapid and environment-friendly, the purity of the compound obtained by separation by the method is higher, and the obtained compound has unique chemical structure, is extracted from the common traditional Chinese medicine purslane and has the anti-tumor and anti-inflammatory effects, so the special compound, the salt and the derivative thereof can be used as a natural product to develop new traditional Chinese medicines, and have wide prospects.

Claims (3)

1. The purslane contains a peroxy bond compound Oleracone I with a molecular formula of C₁₈H₁₈O₆The chemical structural formula is as follows:

。

2. the method for extracting and separating the compound Oleracone I according to claim 1, which comprises the following steps:

step 1: weighing 150kg of purslane dry medicinal material, performing reflux extraction by adopting water, wherein the using amount of the water is added according to the volume ratio of the water to the medicinal material of 10:1, performing reflux extraction twice, each time for 2 hours, heating and concentrating at 90-100 ℃, and cooling to room temperature to obtain liquid medicine for later use;

step 2, evaporating the liquid medicine obtained in the step 1 to dryness, performing chromatographic separation by using a silica gel column, isocratically eluting by using 115L ethyl acetate, wherein the silica gel is 100-200 meshes, and recovering the ethyl acetate to obtain an extract under reduced pressure at the temperature of below 40 ℃ to obtain an ethyl acetate extract;

step 3, separating the ethyl acetate extract obtained in the step 2 by using a polyamide column, performing gradient elution by using ethanol and water in a volume ratio of 0:100, 30:70, 50:50, 70:30 and 100:0, evaporating to dryness at 90-100 ℃ by using 50% ethanol, performing chromatographic separation by using a silica gel column, wherein the silica gel is 200-300 meshes, performing gradient elution by using ethyl acetate-methanol in a volume ratio of 5:1, 2:1 and 1:2 in sequence to obtain 19 parts in total, detecting by using a thin-layer chromatography, developing, combining the developed 1-14 elution parts, and concentrating the combined 1-14 parts under reduced pressure at 40 ℃ until the parts are dry for later use;

step 4, subjecting the product obtained in the step 3 to pretreated ODS medium-pressure column chromatography separation, wherein the filler particle size is 20-40 μm, performing gradient elution by using methanol-water with the volume ratio of 50:50, 60:40, 70:30, 80:20 and 100:0, pressurizing at room temperature until the flow rate is 1mL/min to obtain 16 parts, detecting by thin-layer chromatography, developing, reserving 7-9 parts for developing, and concentrating under reduced pressure below 50 ℃ until the parts are dry for later use;

step 5, subjecting the product obtained in the step 4 to chromatography separation by a pretreated sephadex column, eluting by methanol to obtain 26 elution parts, detecting by thin-layer chromatography, developing, reserving the developed 5-11 parts, and concentrating under reduced pressure below 50 ℃ to dryness for later use to obtain a new compound; the pretreatment process of the ODS and the sephadex comprises the steps of soaking in methanol for 24 hours, loading on a column, washing with methanol until no turbidity exists in dropwise added water, and then balancing with an initial mobile phase;

and 6, separating and preparing the new compound obtained in the step 5 by HPLC, wherein the volume ratio is 40: 60 parts of acetonitrile: taking 0.1% formic acid as a mobile phase, detecting the wavelength of 210nm and 280nm, separating and preparing the compound Oleracone I as the claim 1, wherein the purity is 90-99% by a normalization method.

3. Use of the compound Oleracone I as claimed in claim 1 for the preparation of an anti-tumor, anti-inflammatory medicament.

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