A pair of caged oxadiazine compounds that are enantiomers of each other, and their preparation method and use

Abstract

The present invention relates to the field of natural medicine technology, and specifically to a caged ketone compound extracted from Garcinia macrobracte leaves, as well as a preparation method and use thereof. An insulin resistance model was established by inducing HepG2 cells with palmitic acid, and the hypoglycemic activity of the compound was tested using an in vitro glucose oxidase method. The results showed that the caged ketone compound could significantly increase glucose consumption in HepG2-IR cells and improve the insulin resistance state of HepG2-IR cells to varying degrees. In vitro anti-proliferative activity experiments showed that the caged ketone compound had significant anti-A549 cell proliferation activity. The present invention provides alternative compounds for the development of new hypoglycemic and anti-tumor drugs, which is of great significance to the comprehensive development and utilization of Garcinia macrobracte leaves.

Description

Technical Field

[0001] The present invention relates to the technical field of natural medicines, and in particular to a cage-shaped Ketone compounds, preparation methods and uses thereof. Background Art

[0002] Garcinia bracteata CYWu ex YHLi, also known as flower bark tree and Wannian fruit (Malipo, Yunnan), is a plant of the genus Garcinia in the family Guttiferae. The fruit of Garcinia bracteata is edible and has extremely high nutritional value. It is rich in vitamins, proteins, minerals and other nutrients. In recent years, phytochemical studies on its stem bark, leaves, branches and fruits have found that the plant contains polycyclic polyprenylated acylphloroglucinols (PPAPs), benzophenone derivatives, There are many types of chemical components such as ketones, flavonoids, triterpenes, steroids, biphenyls, etc. Ketones and PPAPs are its characteristic and active ingredients. The research team previously isolated a series of novel PPAPs and Ketone compounds, and exhibit multiple biological activities such as anti-tumor, anti-inflammatory and hypoglycemic activities.

[0003] Therefore, extracting, isolating and discovering new compounds with medicinal value from Garcinia macrobracte leaves is of great significance to the comprehensive development and utilization of Garcinia macrobracte leaves. Summary of the Invention

[0004] In view of this, the present invention provides a pair of cage-shaped enantiomers separated and purified from the leaves of Garcinia macrophylla. Ketone compounds, preparation methods and applications thereof.

[0005] The cage-shaped A new ketone compound, the structural formula of which is shown in formula (I):

[0006]

[0007] The present invention provides a caged enantiomer Ketone compounds, the structural formulas are shown in Formula A and B:

[0008]

[0009] The present invention also provides the above-mentioned cage The preparation method of ketone compounds comprises the following steps:

[0010] S1. Weigh and crush dried Garcinia macrobracta leaves, extract with ethanol, and then concentrate the extract under reduced pressure to obtain an ethanol extract;

[0011] S2. Dissolve the ethanol extract with methanol, then add petroleum ether for extraction, and concentrate the lower methanol portion under reduced pressure to obtain a methanol extract;

[0012] S3, dissolving the methanol extract in a 9:1 (v / v) mixed solvent of water and methanol, then adding ethyl acetate for extraction, and concentrating the ethyl acetate fraction under reduced pressure to obtain an ethyl acetate extract;

[0013] S4. The ethyl acetate extract was subjected to normal phase silica gel column chromatography with gradient elution, and similar fractions were combined by TLC detection to obtain 13 fractions, which were labeled A to M in order of increasing polarity;

[0014] S5. Subject the obtained H fraction to medium-pressure ODS reverse-phase column chromatography with gradient elution, and combine similar fractions using TLC detection to obtain 10 fractions, which are labeled H-01 to H-10 in descending order of polarity;

[0015] S6. Component H-06 was subjected to normal phase silica gel column chromatography with gradient elution. Similar components were combined using TLC to obtain 13 components, which were labeled H-06-01 to H-06-13 in order of increasing polarity.

[0016] After the S7 and H-06-07 components were separated by semi-preparative high performance liquid chromatography, the cage-shaped racemates of ketone compounds;

[0017] S8. Perform chiral resolution on the racemate obtained in step S7 to obtain optically pure levorotatory compound A and dextrorotatory compound B.

[0018] Furthermore, in step S4, the conditions for gradient elution of the ethyl acetate extract are: gradient elution using petroleum ether and ethyl acetate at a volume ratio of 10:1, 8:2, 7:3, 6:4, 1:1, 3:7, and 0:1.

[0019] Furthermore, in step S5, the conditions for gradient elution of component H are: gradient elution using water and methanol at a volume ratio of 7:3, 5:5, 4:6, 3:7, 2:8, and 0:1.

[0020] Furthermore, in step S6, the conditions for gradient elution of component H-06 are: using petroleum ether-ethyl acetate as eluent, and performing gradient elution according to the volume ratio of the two: 15:1, 10:1, 6:1, 8:2, 7:3, 6:4, and 1:1.

[0021] Furthermore, in step S7, the conditions for high performance liquid chromatography separation of component H-06-07 are as follows: using a mixed solvent of methanol and water as the eluent, isocratic elution is performed according to a volume ratio of methanol:water of 77:23, a flow rate of 3 mL / min, and a detection wavelength of 254 nm.

[0022] Furthermore, in step S8, the chromatographic conditions for chiral separation are: a detection wavelength of 254 nm, a mixed solvent of n-hexane and isopropanol as the eluent, isocratic elution according to a volume ratio of n-hexane: isopropanol of 95:5, and a flow rate of 2 mL / min to obtain compound A and compound B.

[0023] The above cage Ketone compounds can serve as lead compounds for hypoglycemic and anti-tumor drugs and have further research and development value.

[0024] The advantages and beneficial effects of the technical solution provided by the present invention are as follows: the present invention obtains a pair of new compounds that are mutually enantiomers by separating and purifying the ethanol extract of Garcinia macrobracte leaves, and comprehensively applies a variety of spectral analysis methods to determine that the new compounds are cage-shaped. Ketone compounds; by conducting in vitro anti-proliferation and glucose consumption experiments on the obtained compounds, it was found that they had anti-proliferation activity on A549 and significantly promoted the glucose consumption of HepG2-IR cells; the present invention provides a lead compound for the development of new anti-tumor and hypoglycemic drugs, and is of great significance to the comprehensive development and utilization of Garcinia macrobracte leaves. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 Prepare cage-shaped Flow chart of extraction and separation of ketone compounds;

[0026] Figure 2 The product obtained in step S7 of Example 1 1 H-NMR (500 MHz, CD3OD) spectrum;

[0027] Figure 3 The product obtained in step S7 of Example 1 13 C-NMR (125 MHz, CD3OD) spectrum;

[0028] Figure 4 DEPT (Distortionless Enhancement by Polarization Transfer) (θ=90°) spectrum of the product obtained in step S7 of Example 1;

[0029] Figure 5DEPT (θ=135°) spectrum of the product obtained in step S7 of Example 1;

[0030] Figure 6 HSQC of the product obtained in step S7 of Example 1 ( 1 H-detected Heteronuclear SingleQuantum Coherence (H-detected Heteronuclear SingleQuantum Coherence) spectrum;

[0031] Figure 7 is the HMBC of the product obtained in step S7 of Example 1 ( 1 H-detected Heteronuclear MultipleBond Correlation spectrum;

[0032] Figure 8 The product obtained in step S7 of Example 1 1 H- 1 H COSY (Correlation Spectroscopy) spectrum;

[0033] Figure 9 ROESY (Rotating Frame Overhauser Effect Spectroscopy, rotating coordinate system NOE spectrum) spectrum of the product obtained in step S7 of Example 1;

[0034] Figure 10 HR-ESI-MS spectrum of the product obtained in step S7 of Example 1;

[0035] Figure 11 This is the chiral separation chromatogram of step S8 in Example 1;

[0036] Figure 12 CD spectra of compounds A and B prepared in Example 1;

[0037] Figure 13 The experimental ECD and calculated ECD spectra of compounds A and B prepared in Example 1;

[0038] Figure 14 The antiproliferative activities of the compound of Example 1, Rubescensine A and Paclitaxel at different concentrations on A549 cells in Experiment (I) of Example 2;

[0039] Figure 15 The effect of different concentrations of the compound of Example 1 on the viability of HepG2 cells in Experiment (II) of Example 2;

[0040] Figure 16The effect of the compound of Example 1 on glucose consumption in HepG2 / IR cells, where NC is the control group; MC is the model group; and Met is metformin. Statistical data are presented as mean ± SEM. ### p<0.001, # indicates control group vs. model group; ***p<0.001, **p<0.01, *p<0.05, * indicates model group vs. drug-treated group. DETAILED DESCRIPTION

[0041] In order to make the objectives, technical solutions and advantages of the present invention more clear, the embodiments of the present invention are further described below in conjunction with specific examples.

[0042] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0043] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0044] Example 1

[0045] This example provides a method for extracting and separating a pair of cage-like enantiomers from the leaves of Garcinia macrophylla. The preparation process of ketone compounds includes the following specific steps:

[0046] Step S1. Weigh 20 kg of dried Garcinia bracteata leaves, crush them, and extract them 3 times with 95% (v / v) ethanol, soaking them at room temperature (25°C, the same below) for 24 hours each time. Filter them after each soaking, collect the extract, combine the extracts after 3 filtrations, and concentrate under reduced pressure to obtain a total extract (2.29 kg); The Garcinia bracteata leaves used in this embodiment were collected in Xichou County, Wenshan Prefecture, Yunnan Province in September 2021, and were identified by Professor Liu Xinqiao of the School of Pharmacy of South-Central University for Nationalities as Garcinia bracteata leaves (Garcinia bracteata CYWu ex YHLi) of the Garcinia family. The plant specimens are preserved in the medicinal plant specimen room of the School of Pharmacy of South-Central University for Nationalities, and the specimen number is 20210901.

[0047] Step S2, dissolving the total extract with methanol, then adding petroleum ether for extraction, concentrating the upper petroleum ether portion under reduced pressure to obtain a petroleum ether extract (100 g), and concentrating the lower methanol portion under reduced pressure to obtain a methanol extract;

[0048] Step S3: The methanol extract was redissolved in a 9:1 (v / v) mixed solvent of water and methanol, and then extracted with ethyl acetate. The ethyl acetate portion was concentrated under reduced pressure to obtain an ethyl acetate extract (430 g).

[0049] Step S4, the ethyl acetate extract was subjected to normal phase silica gel column chromatography, and petroleum ether-ethyl acetate gradient elution (petroleum ether: ethyl acetate = 10:1, 8:2, 7:3, 6:4, 1:1, 3:7, 0:1, v / v) was used to detect and combine similar components using TLC (developing solvent: dichloromethane: methanol = 20:1, v / v) to obtain 13 components, which were labeled AM in ascending order of polarity. The eluates with petroleum ether-ethyl acetate volume ratios of 8:2, 7:3 and 6:4 were collected and combined, numbered H, and concentrated to dryness under reduced pressure for later use.

[0050] Step S5, dry-load the H component (14.27 g), utilize medium-pressure ODS reverse-phase column chromatography, use methanol and water (water: methanol = 7:3, 5:5, 4:6, 3:7, 2:8, 0:1, v / v) for gradient elution, and detect and combine similar components by TLC (developing solvent is dichloromethane: methanol = 20:1, v / v) to finally obtain 10 components, which are labeled H-01 to H-10 in descending order of polarity. The eluate with a water: methanol volume ratio of 3:7 is collected, numbered H-06, and concentrated to dryness under reduced pressure for later use.

[0051] Step S6, H-06 was subjected to normal phase silica gel column chromatography, and petroleum ether-ethyl acetate gradient elution (petroleum ether: ethyl acetate = 15:1, 10:1, 6:1, 8:2, 7:3, 6:4, 1:1, v / v) was used. Similar components were detected and combined by TLC (developing solvent: dichloromethane: methanol = 20:1, v / v) to obtain 13 components, which were labeled (H-06-01 to H-06-13) in ascending order of polarity. The eluates with petroleum ether-ethyl acetate volume ratios of 8:2 and 7:3 were collected and combined, numbered H-06-07, and concentrated to dryness under reduced pressure for later use.

[0052] Step S7: The obtained H-06-07 component (9.75 mg) was purified by HPLC using a semi-preparative column YMC-Pack ODS-A (250×10 mm, 5 μm). The detection wavelength was set to 254 nm, and the eluent was methanol and pure water in a volume ratio of 77:23. The isocratic elution was performed at a flow rate of 3 mL / min. The retention time t R =21.10min, which is the cage shown in formula (I) of the present invention. The weight of the racemate of the ketone compound was 1.75 mg.

[0053] Step S8: The racemate obtained in step S7 was separated using a Chiralpak AD-H (250 mm × 10 mm, 5 μm) semi-preparative column from Daicel. The detection wavelength was set to 254 nm. The mobile phase was n-hexane and isopropanol. The isocratic elution was performed at a flow rate of 2 mL / min in a volume ratio of n-hexane to isopropanol of 95:5 to obtain optically pure compound A (t R :31.6min) and compound B (t R :36.8min), see Figure 11 .

[0054] The flowchart of the above steps is shown in Figure 1 .

[0055] The product obtained in this example was subjected to high-resolution mass spectrometry, ultraviolet spectroscopy, optical rotation, and nuclear magnetic resonance analysis to determine the structure of the compound. The physicochemical data and spectral data are as follows:

[0056] Racemic form: colorless amorphous powder; HR-ESI-MS m / z 511.26935[M+H] + (calcd forC 30 H 39 O7,511.26903);

[0057] A: (c 0.05, MeOH); ECD (9.80×10 -4 M,MeOH)λ(θ)214(-4.47),235(+3.26),316(-5.21)nm;

[0058] B: (c 0.05, MeOH); ECD (9.80×10 -4 M,MeOH)λ(θ)214(+4.15),237(-3.89),315(+4.95)nm;

[0059] The NMR data of the compounds are shown in Table (2). 1 H NMR (500 MHz, CD3OD) spectrum is shown in Figure 2 , 13 C-NMR (125MHz, CD3OD) spectrum is shown in Figure 3 , DEPT (θ=90°) spectrum is shown in Figure 4 , DEPT (θ=135°) spectrum is shown in Figure 5 , HSQC spectrum see Figure 6 , HMBC spectrum see Figure 7 , 1 H- 1 H COSY spectrum see Figure 8 , ROESY spectrum see Figure 9 , HR-ESI-MS spectrum see Figure 10 Based on the above high-resolution mass spectrometry and various NMR data, the planar structure of the compound can be identified as 1-O-methyl-8-methoxy-8,8a-dihydroforbesione. The racemate of compound (I) was resolved by chiral column, and a pair of enantiomers of compound (I) were obtained under the conditions of 95% n-hexane-5% isopropanol. and In order to further determine the absolute configurations of A and B, the ECD curves of A and B were compared with those of (-) and (+)-8-methoxy-8,8a-dihydrobractatin (J.Nat.Prod.2018,81,749-757), respectively. We found that the ECD curves of A and (-)-8-methoxy-8,8a-dihydrobractatin showed a negative Cotton effect between 300-320 nm, while at 2 A positive Cotton effect appears between 20 and 240 nm, confirming the absolute configuration of A to be (5R, 7S, 8S, 8aR, 10aS, 17S). Conversely, B and (+)-8-methoxy-8,8a-dihydrobractatin exhibit a positive Cotton effect between 300 and 320 nm, and a negative Cotton effect between 220 and 240 nm, confirming the absolute configuration of B to be (5S, 7R, 8R, 8aS, 10aR, 17R). To further validate these results, we used ECD calculations to verify these conclusions.

[0060] Table 1: Chinese and English names of compounds represented by formula (I)

[0061]

[0062] Table 2: Compounds 1 H-NMR and 13 C-NMR data

[0063]

[0064] (Recorded in CD3OD)

[0065] Example 2: In vitro antiproliferative activity test

[0066] The compound prepared in Example 1 was subjected to an in vitro antiproliferative activity test.

[0067] Materials and reagents: DMEM high-glucose medium was purchased from Hyclone, USA; penicillin-streptomycin solution was purchased from Hyclone, USA; fetal bovine serum was purchased from Hangzhou Sijiqing Bioengineering Materials Co., Ltd.; CCK-8 kit was purchased from Shanghai Biyuntian Biotechnology Co., Ltd.; dimethyl sulfoxide (DMSO, BIOSHARP, USA); glucose solution was purchased from Hyclone, USA; 6 mmol / L ready-to-use palmitic acid solution was purchased from Xi'an Kunchuang Technology Development Co., Ltd.; metformin hydrochloride tablets were purchased from Shanghai Bristol-Myers Squibb Pharmaceuticals Co., Ltd.; glucose detection kit was purchased from Nanjing Jiancheng Bioengineering Research Institute; paclitaxel (PTX, product number: B21695, Shanghai Yuanye Biotechnology Co., Ltd.); oridonin (homemade, purity greater than 98%); Multiskan GO multifunctional microplate reader (Thermo Fisher, USA).

[0068] The test tumor cell lines: human non-small cell lung cancer A549 cells, purchased from Nanjing Senbeijia Biotechnology Co., Ltd. (Cat NO: BC-C-HU-009); human hepatocellular carcinoma cell HepG2, purchased from Wuhan Punosai Life Science Co., Ltd. (Cat NO: CL-0103).

[0069] (I) CCK-8 assay to detect the antiproliferative activity of compounds A and B on A549 cells

[0070] Experimental methods:

[0071] Experimental grouping and dosage setting: Compounds A and B (50 μM) were used as the experimental group, Rubescensine A (50 μM) and Paclitaxel (20 μM) were used as positive drugs, and a control group (an equal volume of serum-free culture medium containing cells but no compounds) and a blank control group (an equal volume of serum-free culture medium without compounds and cells) were set up.

[0072] A549 cells in the logarithmic growth phase were cultured in DMEM high glucose medium (containing penicillin and streptomycin) containing 10% fetal bovine serum until the cells reached about 80% of the total growth rate. Then A549 cells (1×10 5Cells (cells / well) were inoculated into 96-well plates in a final volume of 100 μL and cultured in a 37°C cell culture incubator for 6 hours until the cells adhered. Compounds A and B from Example 1 were prepared in DMSO to a 10 mM stock concentration and then diluted to various concentrations in DMEM. After the cells adhered, the culture medium in the 96-well plates was discarded, and after rinsing with PBS (1×, pH=7.6, the same below), 100 μL of the test compound at a concentration of 50 μmol / L was added to each well. The plates were then incubated in a 37°C, 5% CO2 incubator for 48 hours. The supernatant from each well was then discarded, and 100 μL of culture medium containing 10% (volume percentage) CCK-8 working solution was added to each well. The plates were incubated for another 1 hour at 37°C, 5% CO2, and the absorbance (A) of each well at 450 nm was measured using a microplate reader. Six replicate wells were set up for each group. The inhibition rate was calculated based on the measured A value. For compounds with an inhibition rate of 50% or more, a gradient concentration (1, 3.125, 6.25, 12.5, 25, 50 μM) was set and the corresponding IC 50 Value (see Figure 14 ).

[0073] Inhibition rate = [(A 对照 -A 实验 ) / (A 对照 -A 空白 )]×100%

[0074] Experimental conclusion:

[0075] from Figure 14 It can be seen that the anti-proliferative activity screening results showed that compounds A and B had significant anti-proliferative effects on A549 cells at a concentration of 50 μM. The IC 50 The IC values ​​of positive drugs Rubescensine A and Paclitaxel were 8.34±2.53μM and 8.11±1.69μM, respectively. 50 The values ​​were 9.20±1.51μM and 1.37±1.11μM, respectively.

[0076] (II) Glucose consumption activity assay

[0077] 1. Effects of Compounds A and B on HepG2 Cell Viability

[0078] Experimental methods:

[0079] Experimental grouping and dosage setting: Compounds A and B (5 and 10 μM) were used as the experimental group, metformin (10 μM) was used as the positive control group, and a control group (an equal volume of serum-free culture medium containing cells but no compounds) and a blank control group (serum-free culture medium without compounds and cells) were set up at the same time.

[0080] The CCK-8 assay was used to detect the effects of compounds on HepG2 cell viability. Cells in the logarithmic growth phase were digested with trypsin and prepared with serum-free DMEM medium at a concentration of 1×10 5 100 μL of cell suspension with a concentration of 5 μM / mL was inoculated into a 96-well plate in each well and cultured at 37°C and 5% CO2. When the cell monolayer adhered to 80%-90%, the supernatant was discarded and 100 μL of serum-free DMEM medium containing different concentrations (5 μM, 10 μM) of the compound was added (the compound was pre-dissolved in DMSO to prepare a 10 mM stock solution). At the same time, a control group (an equal volume of serum-free DMEM medium containing cells but not compounds) and a blank control group (serum-free medium without compounds and cells) were set up. After incubation for 24 hours, the supernatant was discarded and 100 μL of culture medium containing 10% (volume percentage) CCK-8 working solution was added to each well. The plates were cultured at 37°C and 5% CO2 for 1 hour. The absorbance at 450 nm (A value) was measured on a microplate reader and the cell survival rate was calculated. The results are shown in FIG. Figure 15 .

[0081] Cell survival rate = [(A s -A b ) / (A c -A b )]×100%

[0082] A s : Experimental group; A c : control group; A b : Blank control group

[0083] 2. Effects of Compounds on Glucose Consumption in HepG2 / IR Cell Model

[0084] Experimental grouping and dosage setting: the serum-free DMEM group without compounds and cells was used as the blank control group; the serum-free DMEM group without compounds but with cells was used as the control group (NC); the serum-free DMEM group without compounds but with palmitic acid (0.25 mM) and glucose solution (30 mM) was used as the model group (MC); the serum-free DMEM group with palmitic acid (0.25 mM), glucose solution (30 mM) and metformin (10 μM) was used as the positive control group (Met); the serum-free DMEM group with palmitic acid (0.25 mM), glucose solution (30 mM) and the test compound (10 μM) was used as the experimental drug group.

[0085] Based on the cytotoxicity results of the compounds in Experiment 1, glucose consumption experiments were conducted on the compounds that achieved a cell viability of 90% at a concentration of 10 μM. The glucose oxidase method was used to determine the effects of the compounds on glucose consumption in HepG2 / IR model cells induced by palmitic acid (PA).5 A cell suspension of 100 μL / mL was seeded into a 96-well plate, with 100 μL per well. The cells were cultured at 37°C and 5% CO2 until 80%-90% cell monolayer adherence was achieved. The supernatant was discarded, and the cells were washed once with PBS. Then, 100 μL of serum-free medium was added to each well to starve the cells for 12 hours. The old culture medium was discarded, and a blank control group, a control group (NC), a model group (MC), a positive drug group (Met), and an experimental drug group were set up, with a final volume of 100 μL. After incubation at 37°C and 5% CO2 for 24 h, 2 μL of the supernatant was aspirated, and 2 μL of glucose calibration solution was aspirated as a calibration group to eliminate the influence of the glucose kit on absorbance. The supernatant aspirated from each group was added to the wells containing 200 μL of glucose detection reagent. After incubation at 37°C and 5% CO2 for 0.5 h, the absorbance was measured at 505 nm using a microplate reader, and the glucose consumption (GC) of each group was calculated. At the same time, 100 μL of culture medium containing 10% (volume percentage) CCK-8 working solution was added to each well of the original 96-well plate with 2 μL of supernatant aspirated. The cells were incubated at 37°C and 5% CO2 for 1 h. Finally, the absorbance at 450 nm was measured using a microplate reader, and the cell viability CV of each group was determined (calculation method is the same as experiment 1). The GC / CV values ​​were used to evaluate the effect of the monomeric compounds on glucose consumption in HepG2 / IR cells. The experimental results are shown in Figure 16 .

[0086] Experimental conclusion:

[0087] After 24 hours of co-incubation with palmitic acid and high glucose, glucose consumption in the MC group decreased significantly compared to the NC group, with a GC / CV value of 2.62±0.23 mM, indicating that HepG2 cells developed IR and the model was successfully established. Compared with the MC group, metformin significantly enhanced glucose consumption. Treatment with 10 μM compounds A and B promoted glucose consumption in HepG2 / IR cells to varying degrees, improving the insulin resistance of HepG2 cells and demonstrating good hypoglycemic activity. Their GC / CV values ​​were 8.93±0.09 mM and 8.74±0.07 mM, respectively, demonstrating superior efficacy compared to metformin and restoring the cells' glucose utilization to the level of the NC group.

Claims

1. A cage-like Ketone compounds, whose structural formula is:

2. The cage-shaped A ketone compound, characterized in that The structural formula of the compound is:

3. A method for preparing the compound according to claim 2, characterized in that: The following steps are involved: S1. Weigh and crush dried Garcinia macrobracta leaves, extract with ethanol, and then concentrate the extract under reduced pressure to obtain an ethanol extract; S2. Dissolve the ethanol extract with methanol, then add petroleum ether for extraction, and concentrate the lower methanol portion under reduced pressure to obtain a methanol extract; S3, dissolving the methanol extract with a mixed solvent of water and methanol, then adding ethyl acetate for extraction, and concentrating the ethyl acetate portion under reduced pressure to obtain an ethyl acetate extract; S4. Subjecting the ethyl acetate extract to normal phase silica gel column chromatography with gradient elution to obtain 13 fractions A to M; S5. Subject the obtained H component to medium-pressure ODS reverse-phase column chromatography and gradient elution to obtain 10 components, H-01 to H-10; S6. Component H-06 was subjected to normal phase silica gel column chromatography with gradient elution to obtain 1313 components, H-06-01 to H-06-1; After fractions S7 and H-06-07 were separated by semi-preparative high performance liquid chromatography, a racemate of the compound represented by formula (I) was obtained; S8. Perform chiral separation on the racemate obtained in step S7 to obtain compounds A and B.

4. The preparation method according to claim 3, characterized in that In step S4, the conditions for gradient elution of the ethyl acetate extract are as follows: using petroleum ether-ethyl acetate as the eluent, gradient elution is performed according to the volume ratio of the two of 10:1, 8:2, 7:3, 6:4, 1:1, 3:7, and 0:1, and similar components are combined by TLC detection, wherein the developing solvent is a mixed solvent of dichloromethane-methanol with a volume ratio of 20:

1.

5. The preparation method according to claim 3, characterized in that In step S5, the conditions for gradient elution of component H are as follows: using water-methanol as the eluent, gradient elution is performed according to the volume ratio of the two of 7:3, 5:5, 4:6, 3:7, 2:8, and 0:1, and similar components are combined by TLC detection, wherein the developing solvent is a mixed solvent of dichloromethane-methanol with a volume ratio of 20:

1.

6. The preparation method according to claim 3, characterized in that In step S6, the conditions for gradient elution of component H-06 are as follows: using petroleum ether-ethyl acetate as the eluent, gradient elution is performed according to the volume ratio of 15:1, 10:1, 6:1, 8:2, 7:3, 6:4, and 1:1 between the two, and similar components are combined by TLC detection, wherein the developing solvent is a mixed solvent of dichloromethane-methanol with a volume ratio of 20:

1.

7. The preparation method according to claim 3, characterized in that In step S7, the conditions for high performance liquid chromatography separation of component H-06-07 are as follows: detection wavelength is 254 nm, a mixed solvent of methanol and water is used as the eluent, isocratic elution is performed according to a volume ratio of methanol:water of 77:23, and a flow rate of 3 mL / min.

8. The preparation method according to claim 3, characterized in that In step S8, the chromatographic conditions for chiral separation are: a detection wavelength of 254 nm, a mixed solvent of n-hexane and isopropanol as the eluent, isocratic elution according to a volume ratio of n-hexane:isopropanol of 95:5, and a flow rate of 2 mL / min.

9. The cage-shaped Application of ketone compounds in the preparation of anti-tumor drugs.

10. The cage-shaped Application of ketone compounds in the preparation of hypoglycemic drugs.

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