A kind of guaiane sesquiterpene and uric acid dimer compound and its extraction separation method and application

Abstract

The present application relates to the field of natural products, in particular to a sesquiterpene and uric acid dimer compound of guaiane type and an extraction and separation method and application thereof. A new sesquiterpene and uric acid dimer compound of guaiane type is extracted and separated from Guizhoutianmingjing, and it is found that the compound has strong antitumor activity. The compound has strong inhibitory activity on breast cancer, prostate cancer, nasopharyngeal carcinoma, liver cancer, lung cancer, leukemia, pancreatic cancer, glioma, osteosarcoma, skin cancer, cervical cancer, ovarian cancer, kidney cancer and esophageal cancer, and provides an alternative for broad-spectrum anticancer drugs. Moreover, in the paclitaxel-resistant human prostate cancer cell PC3, human cervical cancer cell Hela, human hepatoma cell HepG2 and human glioma cell U87, the compound also exhibits strong anticancer activity, indicating that it has the potential to treat paclitaxel-resistant patients.

Description

Technical Field

[0001] This invention relates to the field of natural products, and more particularly to a dimer compound of guaiacane-type sesquiterpenes and uric acid, as well as its extraction and separation method and application. Background Technology

[0002] Cancer is one of the leading causes of death worldwide, posing a serious threat to human health. Over the past three decades, significant progress has been made in the development of anti-tumor drugs, with treatment strategies gradually shifting from traditional chemotherapy to molecularly targeted therapy and immunotherapy, forming a new treatment landscape with multiple mechanisms. In this process, natural products, due to their diverse chemical structures, broad biological activities, and low toxicity, continue to be an important source for the discovery of anti-tumor drugs. Statistics show that of the 174 widely used anti-tumor drugs globally, small molecule drugs account for approximately 78%, of which about 83% are directly or indirectly derived from natural small molecule compounds or their structural derivatives. Therefore, screening for highly effective and low-toxicity active ingredients from natural products remains a feasible path for anti-tumor drug development.

[0003] Tianming Jingshu ( Carpesium Plants are widely used in Southeast Asian traditional medicine, often for their anti-inflammatory, hemostatic, and heat-clearing and detoxifying properties. Guizhou Tianming Essence ( Carpesium faberi *Gynostemma pentaphyllum* is a perennial herb belonging to the genus *Gynostemma* in the family Asteraceae. It is mainly distributed in southwestern China and Japan, and is commonly used in traditional medicine to treat infections and bleeding disorders. This plant has a complex and diverse chemical composition, among which sesquiterpene lactones have shown significant antitumor potential in in vitro and in vivo studies and are widely used as drug skeletons for structural optimization. In addition to sesquiterpenes, this plant also contains other types of terpenes and secondary metabolites, possessing the potential to be developed into novel antitumor drugs.

[0004] Although some active ingredients in Guizhou Tianming Essence have been reported in previous studies, its complete chemical composition and anti-tumor mechanism still need to be systematically explored. This invention successfully isolated a novel class of guaiacane-type sesquiterpenes and uric acid dimers from Guizhou Tianming Essence, and preliminary activity evaluation showed that these compounds have strong anti-tumor effects. Summary of the Invention

[0005] The purpose of this invention is to provide dimer compounds of guaiacane-type sesquiterpenes and uric acid from Guizhou Tianming Essence, as well as their extraction and separation methods and their application in the preparation of cancer treatment drugs. The compounds provided by this invention have strong inhibitory activity against breast cancer, prostate cancer, nasopharyngeal carcinoma, liver cancer, lung cancer, leukemia, pancreatic cancer, glioma, osteosarcoma, skin cancer, cervical cancer, ovarian cancer, kidney cancer, and esophageal cancer, providing an option for broad-spectrum anticancer drugs.

[0006] The technical scheme of the present application is as follows: a kind of dimer compound of guaiane type sesquiterpene and uric acid, the compound is as shown in formula I:

[0007] .

[0008] The present application also provides the use of the dimer compound of guaiane type sesquiterpene and uric acid in the preparation of a drug for preventing and / or treating cancer, wherein the cancer is at least one of breast cancer, prostate cancer, nasopharyngeal carcinoma, liver cancer, lung cancer, leukemia, pancreatic cancer, glioma, osteosarcoma, skin cancer, cervical cancer, ovarian cancer, kidney cancer and esophageal cancer.

[0009] As a preferred, the compound as shown in structural formula I or a pharmaceutically acceptable salt thereof is used as a pharmaceutically active component.

[0010] As a preferred, the content of the pharmaceutically active component is 0.1-99.9% (m / m).

[0011] As a preferred, the compound is used in combination with a pharmaceutically acceptable carrier.

[0012] The present application also provides the use of the compound in the preparation of a drug for resisting paclitaxel-resistant cancer, wherein the paclitaxel-resistant cancer is selected from prostate cancer, cervical cancer, liver cancer or glioma. The compound can effectively inhibit the growth of cancer cells resistant to paclitaxel.

[0013] The present application also provides an extraction and separation method of the dimer compound of guaiane type sesquiterpene and uric acid, and the operation of the extraction and separation method is as follows:

[0014] Step (1) reflux extraction is performed on the above-ground parts of Guizhou Tianmingjing by using methanol, the extract is collected and concentrated under reduced pressure to obtain an extract; the extract is dispersed in water, and then extracted with petroleum ether and ethyl acetate in sequence to obtain a petroleum ether extract and an ethyl acetate extract; the ethyl acetate extract is dissolved and subjected to silica gel column chromatography, and gradient elution is performed with petroleum ether-ethyl acetate and dichloromethane-methanol in sequence, and 9 parts of eluate are collected and named as Fr. A-Fr. I, respectively;

[0015] Step (2) the Fr. H part is subjected to small-pore resin chromatography separation, and gradient elution is performed with methanol solution with a volume fraction of 50%-100% in sequence, and 6 parts of eluate are collected and named as Fr. Ga-Fr. Gf in sequence; the Fr. Gd part is subjected to C 18The eluent was collected to obtain five parts, which were named Fr. Gd1~Fr. Gd5 in turn; the Fr. Gd3 part was separated by gel column chromatography, and the eluent was collected to obtain five parts, which were named Fr. Gd3a~Fr. Gd3e in turn; the 50%~100% or 50%~90% methanol solution in step (2) refers to the volume fraction of methanol in the methanol-water mixed solvent being 50%~100% or 50%~90%;

[0016] Step (3) The Fr. Gd3d part was separated by normal phase silica gel column chromatography, eluted with dichloromethane-methanol, and the eluent was collected to obtain three parts, which were named Fr. Gd3d I~Fr. Gd3d III in turn; the Fr. Gd3d II was purified by preparative HPLC to obtain the dimer compound of the guaiane-type sesquiterpene and uric acid.

[0017] As preferred, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate in step (1) is 50:1~5:1; the volume ratio of dichloromethane to methanol in the dichloromethane-methanol is 50:1~1:1.

[0018] As preferred, the gel column chromatography separation conditions in step (2) are as follows:

[0019] Eluent: analytical pure methanol;

[0020] Gel column size: 15 mm*160 mm;

[0021] The flow rate of the eluent: 0.5 mL / min;

[0022] The amount of the eluent: 1.8 L.

[0023] The volume ratio of dichloromethane to methanol in the dichloromethane-methanol in step (3) is 20:1~8:1;

[0024] The preparative HPLC purification conditions in step (3) are as follows:

[0025] Column specifications: 4.6 mm*250 mm, 5 μm;

[0026] Mobile phase: acetonitrile-water 63:37;

[0027] Flow rate: 2 mL / min.

[0028] Compared with the prior art, the application has the following beneficial effects: a new diterpene and uric acid dimer compound is extracted and separated from Guizhou Tianmingjing, and it is found that the compound has strong inhibitory activity on breast cancer, prostate cancer, nasopharyngeal carcinoma, liver cancer, lung cancer, leukemia, pancreatic cancer, glioma, osteosarcoma, skin cancer, cervical cancer, ovarian cancer, kidney cancer and esophageal cancer, and the compound provides an alternative for broad-spectrum anticancer drugs.

[0029] The compound provided by the application also exhibits strong anticancer activity in paclitaxel-resistant human prostate cancer cells PC3, human cervical cancer cells Hela, human liver cancer cells HepG2 and human glioma cells U87, which indicates that the compound has the potential to treat paclitaxel-resistant cancers. BRIEF DESCRIPTION OF DRAWINGS

[0030] Figure 1 The H NMR spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure: 1 The H NMR spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure:

[0031] Figure 2 The C NMR spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure: 13 The C NMR spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure:

[0032] Figure 3 The HSQC spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure:

[0033] Figure 4 The HMBC spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure:

[0034] Figure 5 The H H COSY spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure: 1 The H H COSY spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure: 1 The H H COSY spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure:

[0035] Figure 6 The ROESY spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure:

[0036] Figure 7 The HR-ESI-MS spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure:

[0037] Figure 8 The IR spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure:

[0038] Figure 9 The UV spectrum of the diterpene and uric acid dimer compound 1 is shown in the following figure:

[0039] Figure 10CD spectrum of a dimeric compound 1 of guaiane sesquiterpene and uric acid;

[0040] Figure 11 Survival rate inhibition curve and half growth inhibition concentration IC 50 (μM) value. DETAILED DESCRIPTION

[0041] The present application provides a dimeric compound 1 of guaiane sesquiterpene and uric acid isolated and purified from Guizhou tianmingjing, having a structure shown in formula I:

[0042]

[0043] The human breast cancer cells in the following experimental results are human breast cancer cells MDA-MB-231, the human prostate cancer cells are human prostate cancer cells PC3, the human liver cancer cells are human liver cancer cells HepG2, the human lung cancer cells are human lung cancer cells H460, the human cervical cancer cells are human cervical cancer cells Hela, the human leukemia cells are human leukemia cells Jurkat, the human esophageal cancer cells are human esophageal cancer cells Bac-109, the human nasopharyngeal cancer cells are human nasopharyngeal cancer cells HNE1, the human renal cancer cells are human renal cancer cells 786-0, the human ovarian cancer cells are human ovarian cancer cells SKOV3, the human glioma cells are human glioma cells U87, the human pancreatic cancer cells are human pancreatic cancer cells SW1990, the human osteosarcoma cells are human osteosarcoma cells MG63, and the human skin cancer cells are human skin cancer cells A375. The conventional cancer cells used above are all from Kunming Medical University.

[0044] Unless otherwise specified, the reagents used in the embodiments of the present application can be purchased through commercial channels. RPMI1640 medium, DMEM medium and fetal bovine serum are purchased from Biological Industries; phosphate buffer (PBS) and 0.25% trypsin (containing EDTA) are purchased from Gibco Company.

[0045] The cell strains are cultured in DMEM or RPMI1640 medium containing 10% fetal bovine serum at 37℃, 5% CO2 and 90% humidity.

[0046] Example 1 Extraction and separation of dimeric compound 1 of guaiane sesquiterpene and uric acid

[0047] (1) The aerial parts of *Guizhou Tianmingjing* were extracted three times by reflux with methanol. The extracts were collected and concentrated under reduced pressure to obtain an extract. The extract was dispersed in an appropriate amount of water and then extracted with petroleum ether and ethyl acetate in sequence to obtain petroleum ether extract and ethyl acetate extract. The ethyl acetate extract was dissolved and mixed with 40-80 mesh silica gel. The sample was subjected to column chromatography with 100-200 mesh separating silica gel. The sample was eluted with a gradient of petroleum ether-ethyl acetate and dichloromethane-methanol in sequence. Nine fractions of eluent were collected and named Fr. A to Fr. I, respectively. The volume ratio of petroleum ether to ethyl acetate was 50:1 to 5:1. The volume ratio of dichloromethane to methanol in the dichloromethane-methanol mixture was 50:1 to 1:1.

[0048] (2) The Fr. H fraction was separated by MCI small-pore resin chromatography, and gradient elution was performed sequentially with methanol aqueous solution of 50% to 100% (v / v). The eluents were collected to obtain 6 fractions, which were named Fr. Ga to Fr. Gf, respectively. The Fr. Gd fraction was separated by C 18 Reversed-phase silica gel column chromatography was performed, with gradient elution using methanol-water solutions of 50%–90% (v / v), collecting five eluent fractions, named Fr. Gd1–Fr. Gd5. The Fr. Gd3 fraction was eluted using Sephadex LH-20 methanol gel column chromatography, collecting 4 mL of the chromatogram to obtain five eluent fractions, named Fr. Gd3a–Fr. Gd3e. The conditions for Sephadex LH-20 methanol gel column chromatography included:

[0049] Eluent: Analytical grade methanol;

[0050] Gel column dimensions: 15 mm * 160 mm;

[0051] The flow rate of the eluent is 0.5 mL / min;

[0052] The amount of elution solution used: 1.8 L;

[0053] (3) The Fr. Gd3d fraction was separated by normal-phase silica gel column chromatography, eluted with dichloromethane-methanol, and the eluents of the three fractions were collected and named Fr. Gd3d I to Fr. Gd3d III, wherein the volume ratio of dichloromethane to methanol was 20:1 to 8:1; the Fr. Gd3d II fraction was purified by preparative HPLC to obtain a dimer compound 1 of guaiacol-type sesquiterpenes and uric acid, wherein the preparative HPLC conditions included:

[0054] Column specifications: 4.6 mm * 250 mm, 5 μm;

[0055] Mobile phase: Acetonitrile - water 63:37;

[0056] Flow rate: 2 mL / min.

[0057] Structural characterization of the dimeric compound 1 of guaiane sesquiterpene and uric acid:

[0058] Figure 1 Structural characterization of the dimeric compound 1 of guaiane sesquiterpene and uric acid: 1 H NMR spectrum; Figure 2 Structural characterization of the dimeric compound 1 of guaiane sesquiterpene and uric acid: 13 C NMR spectrum; Figure 3 HSQC spectrum of the dimeric compound 1 of guaiane sesquiterpene and uric acid: Figure 4 HMBC spectrum of the dimeric compound 1 of guaiane sesquiterpene and uric acid: Figure 5 Structural characterization of the dimeric compound 1 of guaiane sesquiterpene and uric acid: 1 H- 1 H COSY spectrum; Figure 6 ROESY spectrum of the dimeric compound 1 of guaiane sesquiterpene and uric acid: Figure 7 HR-ESI-MS spectrum of the dimeric compound 1 of guaiane sesquiterpene and uric acid: Figure 8 IR spectrum of the dimeric compound 1 of guaiane sesquiterpene and uric acid: Figure 9 UV spectrum of the dimeric compound 1 of guaiane sesquiterpene and uric acid: Figure 10 CD spectrum of the dimeric compound 1 of guaiane sesquiterpene and uric acid. The 1 H-NMR and 13 C-NMR data are shown in Table 1.

[0059] Table 1. H-NMR and C-NMR data of the dimeric compound 1 of guaiane sesquiterpene and uric acid: 1 H-NMR and 13 C-NMR data (600 / 150 MHz, DMSO- d 6

[0060]

[0061] From Table 1 and Figures 1 to 10 it is known that the dimeric compound 1 of guaiane sesquiterpene and uric acid has the following structure data: white amorphous powder. , 330 (–10.03), nm; UV (MeOH) λ max ​​(log ε) 198 (4.78): IR (KBr) ν max 3435 cm -1 2930cm -1 1762 cm -1 1626 cm -1 1247 cm -1 1124 cm -1 1030 cm -1 912 cm -1 Through HR-EI-MS m / z : 415.1632 [M–H] – (calcd for C) 20 H 23 N4O6:415.1618); combined 13 C1-NMR data confirmed the molecular formula to be C10. 20 H 24 N4O6, with an unsaturation degree of 11. Infrared spectroscopy indicates the presence of hydroxyl groups in its structure (3435 cm⁻¹). -1 ), ester carbonyl (1762 cm) -1 ) and olefin double bond (1626 cm) -1 ). 1 H NMR and 13 C10 NMR data indicate that its structure contains two methyl groups [δ]. H 1.75 (3H, s), 0.97 (3H, s); δ C 20.8 (q), 23.6 (q)], 1 olefin double bond [δ H 5.66 (1H, t, J =1.8 Hz); δ C 125.0 (s), 141.4 (t)] and 1 ester carbonyl group [δ C [175.6 (s)] indicates that compound 1 contains a guaiacol-type sesquiterpene skeleton. Further analysis of the remaining NMR data, δ H 4.34, 10.7, 11.1, 11.7, δ C 97.6, 138.4, 149.5, 152.2, and 152.7 represent a single uric acid structure, linked by HMBC-related H2-12 (δ) H 3.94, d J =7.8 Hz) to C-16 (δ C 138.4) and C-17 (δ C149.5), indicating that the 12-position of the quassinane-type sesquiterpene skeleton was substituted by uric acid, forming a dimer of quassinane-type sesquiterpene and uric acid. Meanwhile, through 1 H- 1 H COSY, HMBC and ROESY correlation signals also confirmed to be a dimer of quassinane-type sesquiterpene and uric acid. Compound 1 was named as Carpespene K.

[0062] Example 2 Detection of Carpespene K Anti-tumor Activity

[0063] CCK-8 assay was used to detect cell growth inhibition. CCK-8 assay, full name Cell Counting Kit-8, is a commonly used method for cell proliferation and cytotoxicity detection. The principle of CCK-8 assay is based on the fact that WST-8 (2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfonic acid phenyl)-2H-tetrazole monosodium salt) can be reduced to form a highly water-soluble orange yellow formazan product in the presence of electron carrier 1-methoxy PMS (methyl naphthoquinone phosphate). The amount of formazan generated is proportional to the number of living cells, and the color depth can reflect the metabolic activity of cells. The absorbance can be measured by a microplate reader at 450 nm wavelength, which can indirectly reflect the number of living cells. Therefore, CCK-8 can be used to evaluate cell proliferation, cytotoxicity or cell effect of drugs.

[0064] Compound Carpespene K was dissolved in DMSO (dimethyl sulfoxide), and then each compound was prepared into a solution with a concentration of 10 μM, 1 μM, 500 nM, 100 nM, 60 nM, 30 nM, 10 nM and 1 nM using DMSO; the above concentration solutions were used as the test solutions.

[0065] Human breast cancer cells MDA-MB-231 (4×10 3 were inoculated in 96-well plates, and the medium in each well was 99 μL. After incubation at 37°C in a 5% CO2 incubator for 24 h, 1 μL of the test solution was added to each well of the cell culture; the control group was only added with 1 μL of DMSO to the cell culture, and then incubated in the incubator for 72 h. 10 μL of CCK-8 solution was added to each well, and gently mixed to avoid air bubbles; incubation was continued in the incubator for 1 h. The absorbance of each well was measured at 450 nm wavelength by a microplate reader, and the average OD value of each well was calculated. By comparing with the control group (untreated cells), the changes in cell proliferation or toxicity were analyzed. The percentage of the absorbance value of the experimental group relative to the absorbance value of the control group represented the survival rate of the cells or the level of cell proliferation, i.e. the control group was set as 100%.

[0066] Results are shown in Table 2. Figure 11 Carpespene K inhibited the survival of human breast cancer cell MDA-MB-231 in a concentration gradient dependent manner. The half growth inhibition concentration IC 50 (μM) value was calculated, and the IC 50 value of Carpespene K on MDA-MB-231 cell was 0.017 μM, indicating that Carpespene K has strong anti-breast cancer activity.

[0067] Example 3 Evaluation of cytotoxicity of Carpespene K on different tumor cells

[0068] Human breast cancer cell MDA-MB-231 was replaced by human prostate cancer cell PC3, human nasopharyngeal carcinoma cell HNE1, human cervical cancer cell Hela, human ovarian cancer cell SKOV3, human skin cancer cell A375, human renal cancer cell 786-0, human esophageal cancer cell Bac-109, human lung cancer cell H460, human leukemia cell Jurkat, human osteosarcoma cell MG63, human liver cancer cell HepG2, human pancreatic cancer cell SW1990 and human glioma cell U87; paclitaxel was used as positive control, and other conditions were the same as in Example 2. The results of IC 50 values are shown in Table 2.

[0069] Table 2 Half growth inhibition concentration IC 50 (μM) of Carpespene K on various cancer cells

[0070]

[0071] According to the results in Table 2, Carpespene K has very strong cytotoxic activity on human prostate cancer cell PC3, human nasopharyngeal carcinoma cell HNE1, human cervical cancer cell Hela, human ovarian cancer cell SKOV3, human skin cancer cell A375, human renal cancer cell 786-0, human esophageal cancer cell Bac-109, human lung cancer cell H460, human leukemia cell Jurkat, human osteosarcoma cell MG63, human liver cancer cell HepG2, human pancreatic cancer cell SW1990 and human glioma cell U87.

[0072] It is worth noting that the activity of Carpespene K on human prostate cancer cell PC3, human nasopharyngeal carcinoma cell HNE1, human cervical cancer cell Hela, human ovarian cancer cell SKOV3, human skin cancer cell A375, human lung cancer cell H460, human liver cancer cell HepG2, human pancreatic cancer cell SW1990 and human glioma cell U87 is superior to that of paclitaxel, a clinically used chemotherapeutic drug.

[0073] Moreover, Carpespene K also showed strong anticancer activity in paclitaxel-resistant human prostate cancer cell PC3, human cervical cancer cell Hela, human hepatoma cell HepG2 and human glioma cell U87, indicating that Carpespene K has the potential to treat paclitaxel-resistant patients.

[0074] In summary, Carpespene K has very strong anticancer effect and can effectively inhibit the proliferation of various cancer cells, and has broad-spectrum anticancer activity.

[0075] It should be noted that the above-described examples are only for the purpose of explaining the present application and do not constitute any limitation on the present application. The present application is described by referring to typical examples, but it should be understood that the words used therein are descriptive and explanatory words, rather than limiting words. Modifications can be made to the present application within the scope of the claims of the present application, and the present application can be revised without departing from the scope and spirit of the present application. Although the present application described therein relates to specific methods, materials and examples, it does not mean that the present application is limited to the specific examples disclosed therein, on the contrary, the present application extends to all other methods and applications having the same function.

Claims

1. A dimeric compound of a guaiane sesquiterpene and uric acid, characterized in that, The compound is shown as formula I: 。 2. Use of the dimeric compound of the quassinoid sesquiterpene and uric acid according to claim 1 for the preparation of a medicament for the prevention and / or treatment of cancer, characterized in that, The cancer is at least one of breast cancer, prostate cancer, nasopharyngeal cancer, liver cancer, lung cancer, leukemia, pancreatic cancer, glioma, osteosarcoma, skin cancer, cervical cancer, ovarian cancer, kidney cancer and esophageal cancer.

3. Use according to claim 2, characterized in that, The compound shown in structural formula I or a pharmaceutically acceptable salt thereof is used as a pharmaceutical active ingredient.

4. Use according to claim 3, characterized in that, The content of the pharmaceutical active ingredient is 0.1-99.9%.

5. Use according to claim 2, characterized in that, The compound is used in combination with a pharmaceutically acceptable carrier.

6. Use of the dimer of a guaiane sesquiterpene and uric acid according to claim 1 for the preparation of a medicament against cancer resistant to paclitaxel, characterized in that, The paclitaxel-resistant cancer is selected from prostate cancer, cervical cancer, liver cancer or glioma.

7. The method of claim 1, wherein the method is characterized by, The extraction and separation method is operated as follows: Step (1) The above-ground parts of Guizhou tianmengjing are reflux extracted with methanol, the extract is collected and concentrated under reduced pressure to obtain an extract; the extract is dispersed in water and then extracted with petroleum ether and ethyl acetate in sequence to obtain petroleum ether extract and ethyl acetate extract; the ethyl acetate extract is dissolved and subjected to silica gel column chromatography, and gradient elution is performed with petroleum ether-ethyl acetate and dichloromethane-methanol in sequence, and 9 parts of eluate are collected and named as Fr. A-Fr. I, respectively; Step (2) Fr. H was separated by small pore resin column chromatography, and gradient elution was performed using methanol solution with volume fraction of 50% to 100% successively, and the eluate of 6 fractions was collected successively and named as Fr. Ga to Fr. Gf; Fr. Gd was separated by C 18 Fr. Gd1 to Fr. Gd5; Fr. Gd3 was separated by gel column chromatography, and the eluate of 5 fractions was collected successively and named as Fr. Gd3a to Fr. Gd3e; Step (3) The Fr. Gd3d part is separated by normal-phase silica gel column chromatography and eluted with dichloromethane-methanol, and 3 parts of eluate are collected and named as Fr. Gd3d I-Fr. Gd3d III in sequence; the Fr. Gd3d II is purified by preparative HPLC to obtain the dimer compound of guaiane-type sesquiterpene and uric acid.

8. The method of claim 7, wherein the method is characterized by, In step (1), the volume ratio of petroleum ether to ethyl acetate in petroleum ether-ethyl acetate is 50:1-5:1; and the volume ratio of dichloromethane to methanol in dichloromethane-methanol is 50:1-1:

1.

9. The method of claim 7, wherein the method is characterized by, In step (2), the gel column chromatography separation conditions are as follows: Eluate: analytical pure methanol; Gel column size: 15 mm*160 mm; Flow rate of eluate: 0.5 mL / min; Eluate amount: 1.8 L.

10. The method for extracting and separating guaiacane-type sesquiterpenes and uric acid dimers according to claim 7, characterized in that, In step (3), the volume ratio of dichloromethane to methanol in dichloromethane-methanol is 20:1-8:

1. In step (3), the preparative HPLC purification conditions are as follows: Column specifications: 4.6 mm*250 mm, 5 μm; Mobile phase: acetonitrile-water 63:37; Flow rate: 2 mL / min.

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