Compound with anti-tumor effect in Sinkiang salvia officinalis, separation method and application

Abstract

The invention relates to a compound with an anti-tumor effect in Sinkiang salvia officinalis as well as a separation method and application thereof, and belongs to the technical field of medicines. The invention particularly discloses a separation and preparation method and application of a compound which is separated from Sinkiang salvia officinalis and has an anti-tumor effect. Researches prove that the compound has inhibitory activity on proliferation of tumor cells such as lung cancer A-549, breast cancer MCF-7, colon cancer SW480, liver cancer SMMC-7721, leukemia HL-60 and the like; and the dose dependence reduces the proliferation and metastasis of zebra fish transplanted tumors. The dimerized terpenoids can be used as lead compounds for development of antitumor drugs, and are of great significance to comprehensive utilization and development of Sinkiang salvia officinalis.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical technology, and more specifically, relates to compounds with anti-tumor effects in Salvia deserta Schang, a separation method thereof, and applications thereof. Background Art

[0002] Malignant tumors are characterized by uncontrolled growth and spread of cells, and their incidence and mortality rates have been increasing year by year, posing a great threat to human life, health and safety. At present, chemotherapy remains the main means of treating malignant tumors, and the main chemotherapy drugs clinically are cytotoxic drugs such as alkylating agents, antimetabolites, and antibiotics. Traditional cytotoxic drugs non-specifically block cell division, thereby causing cell death. While killing tumor cells, they also damage normal human cells, and most of these drugs have disadvantages such as poor selectivity, strong toxic and side effects, and drug resistance, affecting the quality of life of patients. Therefore, accelerating the development of safe, effective, and low-toxic anti-cancer drugs has become a common goal pursued by researchers. Active ingredients of Chinese herbal medicines have the advantages of novel skeletal structures and structural diversity, and show advantages such as good curative effects, small side effects, and significantly improving the quality of life of patients in the prevention and treatment of tumor diseases. Therefore, active ingredients of Chinese medicines are an important source of lead compounds for anti-cancer innovative drugs.

[0003] Salvia deserta Schang is a perennial herbaceous plant growing in the Gobi wilderness area of Xinjiang. Due to its deep roots, it plays an active role in wind prevention and sand fixation. Salvia deserta Schang has high utility in the agricultural field. It can not only improve the orchard environment and provide green organic fertilizers, but also be used as an ornamental plant for landscaping. The stems and leaves of Salvia deserta Schang are usually brewed as tea by local people, and its roots are also used to make medicinal wine, which is favored because it is said to be beneficial to enhancing physical health. Salvia deserta Schang is also known as "Xinjiang Salvia miltiorrhiza" and is a famous traditional medicine, with the effects of clearing heat and detoxifying, relieving phlegm and cough, and detumescence and diuresis. When the resources of Salvia miltiorrhiza are scarce, the roots of Salvia deserta Schang can be used as a substitute. However, there is little research on the active substance basis of Salvia deserta Schang. Summary of the Invention

[0004] The purpose of the present invention is to provide terpene dimer compounds with anti-tumor effects in Salvia deserta Schang and a method for separating and preparing the same, and to clarify the application value of the terpene dimer compounds in the preparation of anti-tumor drugs, thereby solving the problems of poor selectivity, strong toxic and side effects, and drug resistance of chemotherapy drugs.

[0005] According to the first aspect of the present invention, there is provided a terpene dimer compound, and the structural formula of the terpene dimer compound is shown as follows:

[0006]

[0007] R in Formula I 1-R 4 、R in formula II 1 -R 6 、R in formula III 1 -R 6 、R in formula IV 1 -R 6 and R in formula V 1 -R 2 are each independently selected from hydrogen, hydroxyl, methylene, carbonyl, halogen, acyl, acyl halide, nitro, cyano, benzene ring and indole ring.

[0008] Preferably, formula I is compound 1 or compound 2, formula II is compound 3, compound 4, compound 5 or compound 6, formula III is compound 7, formula IV is compound 8, compound 9 or compound 10, and formula V is compound 11;

[0009]

[0010]

[0011] On the other hand, according to the present invention, a method for separating the terpene dimer compound is provided, including the following steps:

[0012] (1) Dry and pulverize the roots of Salvia deserta Schang, extract with ethanol, and concentrate under reduced pressure to obtain a total extract;

[0013] (2) Suspend the total extract obtained in step (1) in water, extract with petroleum ether, and concentrate under reduced pressure to obtain an extract of the petroleum ether fraction;

[0014] (3) Extract the extract obtained in step (2) with a mixture of methanol / water to obtain an extract A of the petroleum ether layer and an extract B of the methanol / water layer; The extract A of the petroleum ether layer is mixed with silica gel and subjected to normal-phase silica gel column chromatography. The eluent is gradient elution with petroleum ether / acetone. Combining TLC spotting detection, 8 components A1 - A8 with increasing polarity are obtained by combining the same components; Component A3 is subjected to reverse-phase C 18 silica gel column chromatography, and the eluent is gradient elution with a mixture of methanol / water to obtain 8 sub-components A3a - A3h with decreasing polarity; Component A3h is subjected to normal-phase silica gel column chromatography, and the eluent is gradient elution with petroleum ether / acetone. Combining the same components, 7 components A3h1 - A3h7 with increasing polarity are obtained;

[0015] When component A3h5 is subjected to Sephadex LH-20 gel column chromatography with methanol as the eluent, 2 sub-components A3h5a and A3h5b with decreasing molecular weight are obtained; Component A3h5b is separated by semi-preparative high-performance liquid chromatography phenyl column, and the eluent is a mixture of acetonitrile / water to obtain compound 10;

[0016] When component A3h7 was subjected to Sephadex LH-20 gel column chromatography with methanol as the eluent, four sub-components A3h7a - A3h7d with decreasing molecular weights were obtained; component A3h7b was separated by semi-preparative high performance liquid chromatography on a phenyl column with a mixture of acetonitrile / water as the eluent to obtain compound 8 and compound 9 respectively;

[0017] When component A4 was subjected to reverse phase C 18 silica gel column chromatography with a gradient elution of a mixture of methanol / water as the eluent, 16 sub-components A4a - A4p with decreasing polarities were obtained; component A4l was further subjected to Sephadex LH-20 gel column chromatography and reverse phase C 18 silica gel column chromatography to obtain 6 sub-components A4l1 - A4l6 with decreasing polarities; component A4l6 was separated by semi-preparative high performance liquid chromatography on a phenyl column with a mixture of methanol / water as the eluent to obtain compound 5 and three other sub-components A4l6a, A4l6c and A4l6d;

[0018] When component A4l6a was purified by semi-preparative high performance liquid chromatography on a phenyl column with a mixture of acetonitrile / water as the eluent, compound 11 was obtained;

[0019] When component A4l6d was purified by semi-preparative high performance liquid chromatography on a phenyl column with a mixture of acetonitrile / water as the eluent, compound 2 was obtained;

[0020] When component A4l6c was purified by semi-preparative high performance liquid chromatography on a phenyl column with a mixture of acetonitrile / water as the eluent, compound 1 and compound 6 were obtained;

[0021] When component A4m was subjected to normal phase silica gel column chromatography and Sephadex LH-20 gel column chromatography, two sub-components A4m1 and A4m2 with decreasing molecular weights were obtained; component A4m2 was subjected to reverse phase C 18 silica gel column chromatography with a gradient elution of methanol / water to obtain 6 sub-components A4m2a - A4m2f with decreasing polarities; component A4m2b was separated by semi-preparative high performance liquid chromatography on a C 18 column with a mixture of acetonitrile / water / formic acid as the eluent to obtain compound 4 and compound 7 respectively;

[0022] When component A4m2c was separated by semi-preparative high performance liquid chromatography on a C 18 column with a mixture of acetonitrile / water as the eluent, compound 3 was obtained.

[0023] According to another aspect of the present invention, there is provided the use of the terpene dimer compound in the preparation of anti-tumor drugs.

[0024] Preferably, the anti-tumor drug is a drug for treating lung cancer, a drug for treating breast cancer, a drug for treating colon cancer, a drug for treating liver cancer, and / or a drug for treating leukemia.

[0025] According to another aspect of the present invention, there is provided the use of Compound 1 in the preparation of a drug for treating lung cancer, a drug for treating breast cancer, a drug for treating colon cancer, a drug for treating liver cancer, and / or a drug for treating leukemia.

[0026]

[0027] According to another aspect of the present invention, there is provided the use of Compound 1 and / or Compound 2 in the preparation of a drug for treating lung cancer;

[0028]

[0029] According to another aspect of the present invention, there is provided the use of Compound 4 in the preparation of a drug for treating breast cancer;

[0030]

[0031] Generally speaking, compared with the prior art, the above technical solutions conceived by the present invention mainly have the following technical advantages:

[0032] (1) Compounds 1-11 provided in the present invention are terpene dimer compounds, which are all first prepared and discovered from Salvia deserta Schang. Compounds 1 and 2 are the first heterologous terpene dimers formed by abietane-type diterpenes and eremophilane sesquiterpenes. Compounds 3-7 and 11 are terpene dimers formed by two molecules of abietane-type diterpenes with different structures. Compounds 8-10 are heterologous terpene dimers formed by abietane-type diterpenes and oleanane-type triterpenes.

[0033] (2) The terpene dimer compounds provided in the present invention have anti-tumor cell proliferation activity. In particular, Compounds 1 and 2 have significant anti-proliferation effects on A-549 lung cancer cells, stronger than the positive drug cisplatin; Compound 4 has significant anti-proliferation effects on MCF-7 breast cancer cells, stronger than cisplatin.

[0034] (3) The terpene dimer compounds provided in the present invention have anti-tumor cell proliferation activity. In particular, in the zebrafish tumor transplantation model, Compound 2 can dose-dependently inhibit the proliferation and metastasis of tumor cells, and the inhibitory effect is stronger than the positive drug etoposide. At a concentration of 1 μM, the inhibitory effect of Compound 2 on zebrafish tumor metastasis is also stronger than that of etoposide at a concentration of 10 μM. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 is the X-ray single crystal diffraction pattern of Compound 3.

[0036] Figure 2Results of the anti-tumor activity of Compound 2 in zebrafish Detailed implementation mode

[0037] In order to make the objectives, technical solutions and advantages of the present invention clearer, the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention. In addition, the technical features involved in the various embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0038] The present invention specifically relates to the separation and purification process of terpene dimer compounds in Salvia deserta Schang., structure determination, in vitro anti-tumor cell proliferation effect and in vivo anti-tumor effect on zebrafish, etc.

[0039] The structural formula of the terpene dimer compound of the present invention is shown as follows:

[0040]

[0041] R in Formula I 1 -R 4 R in Formula II 1 -R 6 R in Formula III 1 -R 6 R in Formula IV 1 -R 6 R in Formula V 1 -R 2 Each independently selects hydrogen, hydroxyl group, methylene group, carbonyl group, halogen, acyl group, acyl halide, nitro group, cyano group, benzene ring or indole ring.

[0042] Formula I represents Compounds 1 and 2, Formula II represents Compounds 3, 4, 5 and 6, Formula III represents Compound 7, Formula IV represents Compounds 8, 9 and 10, and Formula V represents Compound 11;

[0043]

[0044] Compound 1: Heterodesertone A

[0045]

[0046] Compound 2: Heterodesertone B

[0047]

[0048] Compound 3: Heterodesertone C

[0049] Compound 4: Heterodesertone D

[0050]

[0051] Compound 5: Heterodesertone E

[0052]

[0053] Compound 6: Heterodesertone F

[0054]

[0055] Compound 7: Heterodesertone G

[0056]

[0057] Compound 8: Heterodesertone H; Compound 9: Heterodesertone I

[0058]

[0059] Compound 10: Heterodesertone J

[0060]

[0061] Compound 11: Broussonetone A

[0062] The compounds of the present invention are isolated and purified from the medicinal plant Salvia deserta of the genus Salvia in the family Lamiaceae. Through repeated column chromatography separation and purification of the petroleum ether extract of the 95% ethanol extract of Salvia deserta, the above-mentioned terpenoid dimeric compounds 1-11 are all prepared and discovered from Salvia deserta for the first time. Compounds 1 and 2 are the first heteroterpenoid dimers formed by a abietane-type diterpenoid and an elemolane sesquiterpenoid. Compounds 3-7 and 11 are terpenoid dimers formed by two molecules of abietane-type diterpenoids with different structures. Compounds 8-10 are heteroterpenoid dimers formed by an abietane-type diterpenoid and an oleanane-type triterpenoid.

[0063] The present invention uses a variety of spectroscopic analysis methods, chemical derivatization methods, quantum chemical calculation methods, and X-ray single crystal diffraction and other methods to determine their structures.

[0064] The above-mentioned compounds 1-11 provided by the present invention can significantly inhibit the proliferation of tumor cells and can be used for preparing anti-tumor drugs, specifically for preparing drugs for treating lung cancer, breast cancer, colon cancer, liver cancer, leukemia and the like.

[0065] The present invention provides a study on the in vitro anti-proliferation effects of compounds 1-11 on 5 kinds of tumor cells (A-549, MCF-7, SW480, HL-60 and SMMC-7721). The in vitro anti-proliferation activity of tumor cells was determined by the MTS method. The results showed that compounds 1-11 had varying degrees of anti-proliferation effects on the 5 kinds of tumor cells (A-549, MCF-7, SW480, HL-60 and SMMC-7721). Compound 1 had significant in vitro anti-proliferation activity against the 5 kinds of tumor cells (lung cancer, breast cancer, colon cancer, liver cancer and leukemia). Compounds 1 and 2 had significant anti-proliferation effects on A-549 lung cancer cells, stronger than the positive drug cisplatin. Compound 4 had significant anti-proliferation effects on MCF-7 breast cancer cells, stronger than the positive drug cisplatin.

[0066] The present invention constructed a zebrafish lung cancer cell A549 xenograft tumor model by microinjection method to study the in vivo anti-tumor activity of compound 2. The results showed that compound 2 inhibited the proliferation and metastasis of tumor cells in a dose-dependent manner, and its inhibitory effect was stronger than that of the positive drug etoposide at the same concentration. At the concentration of 1 μM, the inhibitory effect of compound 2 on zebrafish tumor metastasis was also stronger than that of etoposide at the concentration of 10 μM.

[0067] The following are specific examples:

[0068] Example 1

[0069] (1) Isolation and preparation of compounds 1-11 shown in Formula I

[0070] 1. Plant materials

[0071] The roots of Salvia deserta Schang were collected from Manas County, Xinjiang Uygur Autonomous Region, China (east longitude 85°34′-86°43′, north latitude 43°28′-45°38′) in September 2015, and were identified as Salvia deserta Schang of the genus Salvia in the family Lamiaceae by the Xinjiang Technical Institute of Physics and Chemistry, Chinese Academy of Sciences.

[0072] 2. Isolation and preparation process

[0073] The dried roots of Salvia deserta Schang (35.0 kg) were pulverized and extracted 5 times with 95% ethanol at room temperature for 7 days each time. The filtrates were combined by reduced pressure concentration to obtain a crude extract (3.0 kg). The crude extract paste was suspended with warm water and extracted with petroleum ether, and the petroleum ether fraction extract paste (710 g) was obtained by reduced pressure concentration. The petroleum ether fraction extract paste was further extracted with methanol / water (80:20, V / V) to obtain petroleum ether layer extract A (457 g) and methanol / water layer extract B (253 g). Extract A was mixed with silica gel of 100 - 200 mesh and subjected to normal phase silica gel column chromatography. The eluent was gradient elution with petroleum ether / acetone (200:1 to 2:1, V / V). Combining TLC spotting detection, 8 fractions (A1 - A8) with increasing polarity were obtained by combining the same components. Fraction A3 was subjected to reverse phase C 18 silica gel column chromatography. The eluent was gradient elution with methanol / water (70:30 to 100:0, V / V), and 8 fractions A3a - A3h with decreasing polarity were obtained. Fraction A3h was subjected to normal phase silica gel column chromatography. The eluent was gradient elution with petroleum ether / acetone (150:1 to 30:1, V / V), and 7 fractions (A3h1 - A3h7) with increasing polarity were obtained by combining the same components. Fraction A3h5 was subjected to Sephadex LH - 20 gel column chromatography with methanol as the eluent, and 2 sub - fractions A3h5a and A3h5b with decreasing molecular weight were obtained. Fraction A3h5b was separated by semi - preparative high performance liquid chromatography on a phenyl column. The eluent was acetonitrile / water (92.5:7.5, V / V), the flow rate was 2.0 mL / min, and compound 10 (7.2 mg, yield 0.0000206%) with a retention time of 44.5 min was obtained. Fraction A3h7 was subjected to Sephadex LH - 20 gel column chromatography with methanol as the eluent, and 4 sub - fractions A3h7a - A3h7d with decreasing molecular weight were obtained. Fraction A3h7b was separated by semi - preparative high performance liquid chromatography on a phenyl column. The eluent was acetonitrile / water (92:8, V / V), the flow rate was 2.0 mL / min, and compound 8 (10.4 mg, yield 0.0000297%, retention time 71.3 min) and compound 9 (5.5 mg, yield 0.0000157%, retention time 74.7 min) were obtained. Fraction A4 was subjected to reverse phase C 18 silica gel column chromatography. The eluent was gradient elution with methanol / water (80:20 to 90:10, V / V), and 16 sub - fractions A4a - A4p with decreasing polarity were obtained. Fraction A4l was further subjected to Sephadex LH - 20 gel column chromatography (eluent: methanol) and reverse phase C 18Silica gel column chromatography (eluent: methanol / water, 75:25 to 90:10, V / V) was performed to obtain six sub-components A4l1 - A4l6 with decreasing polarity. Component A4l6 was separated by semi-preparative high performance liquid chromatography on a phenyl column. The eluent was methanol / water (90:10, V / V), and the flow rate was 2.0 mL / min, yielding compound 5 (7.2 mg, yield 0.0000206%, retention time 55.3 min) and three other sub-components A4l6a, A4l6c, and A4l6d. Component A4l6a was purified by semi-preparative high performance liquid chromatography on a phenyl column. The eluent was acetonitrile / water (85:15, V / V), and the flow rate was 2.0 mL / min, yielding compound 11 (10.8 mg, yield 0.0000309%, retention time 42.8 min). Component A4l6d was purified by semi-preparative high performance liquid chromatography on a phenyl column. The eluent was acetonitrile / water (80:20, V / V), and the flow rate was 2.0 mL / min, yielding compound 2 (10.8 mg, yield 0.0000309%, retention time 75.0 min). Component A4l6c was purified by semi-preparative high performance liquid chromatography on a phenyl column. The eluent was acetonitrile / water (82:18, V / V), and the flow rate was 2.0 mL / min, yielding compound 1 (6.4 mg, yield 0.0000183%, retention time 56.0 min) and compound 6 (4.8 mg, yield 0.0000137%, retention time 72.0 min). Component A4m was subjected to normal phase silica gel column chromatography (eluent: petroleum ether / ethyl acetate, 100:1 to 0:1, V / V) and Sephadex LH-20 gel column chromatography (eluent methanol) to obtain two sub-components A4m1 and A4m2 with decreasing molecular weight. Component A4m2 18 Silica gel column chromatography with gradient elution using methanol / water (85:15 to 100:0, V / V) as the eluent was carried out to obtain six sub-components A4m2a - A4m2f with decreasing polarity. Component A4m2b was separated by semi-preparative high performance liquid chromatography on a C 18 column. The eluent was acetonitrile / water / formic acid (95:5:0.1, V / V), and the flow rate was 2.0 mL / min, yielding compound 4 (2.8 mg, yield 0.0000080%, retention time 39.5 min) and compound 7 (1.0 mg, yield 0.0000029%, retention time 52.5 min). Component A4m2c was separated by semi-preparative high performance liquid chromatography on a C 18 column. The eluent was acetonitrile / water (85:15, V / V), and the flow rate was 2.0 mL / min, yielding compound 3 (3.0 mg, yield 0.0000086%, retention time 50.5 min).

[0074] 2. Structure identification of compounds 1 - 11 shown in Formula I

[0075] Combined with a variety of spectroscopic analysis methods (high-resolution mass spectrometry HRESIMS, ultraviolet spectroscopy UV, infrared spectroscopy IR, and nuclear magnetic resonance spectroscopy NMR), chemical derivatization method, quantum chemical calculation method ( 13 C-NMR DP4+ probability analysis and ECD), and X-ray single crystal diffraction and other methods for comprehensive analysis to determine the structures of new compounds 1-10. The structure of known compound 11 was determined by comparing literature data. Among them, the absolute configuration of compound 3 was determined by X-ray single crystal diffraction, as Figure 1 shown.

[0076] Compound 1 (heterodesertone A): Brown powder; optical rotation value [α] 25 D -91.0 (c 0.1, methanol); ultraviolet (methanol) λ max (logε): 201 (4.41), 267 (3.88) nm; infrared (potassium bromide) ν max 3433, 2958, 2927, 1690, 1635, 1458, 1375, 1248, 1153, 1077, 758 cm -1 ; CD (methanol) λ max (Δε): 309 (+29.56), 402 (-15.83) nm; 1 H and 13 C NMR spectral data are shown in Table 1 and Table 2; high-resolution mass spectrometry m / z 551.3730 [M+H] + (calculated value C 35 H 51 O 5 , 551.3736) and 573.3538 [M+Na] + (calculated value C 35 H 50 O 5 Na, 573.3556).

[0077] Compound 2 (heterodesertone B): Brownish-yellow powder; optical rotation value [α] 25 D -35.0 (c 0.1, methanol); ultraviolet (methanol) λ max (logε): 270 (3.68) nm; infrared (potassium bromide) 3406, 2933, 2869, 1706, 1636, 1460, 1376, 1247, 1154, 1068, 900, 757 cm -1 ; CD (methanol) λ max(Δε): 219 (-29.84), 304 (+47.02), 404 (-29.56) nm; 1 H and 13 C NMR spectral data are shown in Tables 1 and 2; High-resolution mass spectrometry HRESIMS m / z 573.3548 [M+Na] + (calcd for C 35 H 50 O 5 Na, 573.3556) and 1123.7233 [2M+Na] + (calcd for C 70 H 100 O 10 Na, 1123.7214).

[0078] Compound 3 (heterodesertone C): Colorless prismatic crystals (methanol:dichloromethane = 1:1, V / V); Melting point: 205 - 206 °C; Optical rotation [α] 25 D -4.0 (c 0.1, methanol); Ultraviolet (methanol) λ max (logε): 204 (4.68), 277 (4.05) nm; Infrared (KBr) ν max 3439, 2961, 1636, 1385, 1251, 1073, 769 cm -1 ; CD (methanol) λ max (Δε): 249 (-13.28), 299 (+49.52), 403 (-15.24) nm; 1 H and 13 C NMR spectral data are shown in Tables 1 and 2; High-resolution mass spectrometry m / z 615.4047 [M-H] - (calcd for C 40 H 55 O 5 , 615.4049).

[0079] Compound 4 (heterodesertone D): Brownish-yellow powder; Optical rotation [α] 25 D +6.0 (c 0.1, methanol); Ultraviolet (methanol) λ max (logε): 207 (4.72) and 278 (4.29) nm; Infrared (KBr) ν max 3395, 2962, 2871, 1722, 1643, 1461, 1379, 1257, 1163, 1085, 892, and 758 cm -1 ; CD (methanol) λ max(Δε): 264 (+65.26) and 318 (-19.95) nm; 1 H and 13 C NMR spectral data are shown in Tables 1 and 2; High resolution mass spectrometry m / z 639.4019 [M+Na] + (Calculated for C 40 H 56 O 5 Na, 639.4025) and 1255.8092 [2M+Na] + (Calculated for C 80 H 112 O 10 Na, 1255.8135).

[0080] Compound 5 (heterodesertone E): Brownish yellow powder; Optical rotation [α] 25 D -62.5 (c 0.1, methanol); Ultraviolet (methanol) λ max (logε): 272 (4.03) nm; Infrared (KBr) ν max 3414, 2961, 2929, 2872, 1644, 1601, 1461, 1377, 1318, 1246, 1077, 1012, and 757 cm -1 ; CD (methanol) λ max (Δε): 236 (-1.30), 315 (+79.28), and 406 (-35.45) nm; 1 H and 13 C NMR spectral data are shown in Tables 1 and 2; High resolution mass spectrometry m / z 683.3883 [M+Na] + (Calculated for C 41 H 56 O 7 Na, 683.3924).

[0081] Compound 6 (heterodesertone F): Brownish yellow powder; Optical rotation [α] 25 D +0.7 (c 0.1, methanol); Ultraviolet (methanol) λ max (logε): 206 (4.19), 272 (4.04) nm; Infrared (KBr) ν max 3375, 3359, 2959, 2928, 1642, 1602, 1384, 1325, 1247, and 972 cm -1 ; CD (methanol) λ max(Δε): 212(+38.28), 241(+1.66), 278(+28.25), 343(-8.65) nm; 1 H and 13 C NMR spectral data are shown in Tables 2 and 3; High-resolution mass spectrometry m / z 697.3771 [M+Na] + (Calculated for C 41 H 54 O 8 Na, 697.3716).

[0082] Compound 7 (heterodesertone G): Brownish-yellow powder; Optical rotation [α] 25 D +4.5 (c 0.1, methanol); UV (methanol) λ max (logε): 206(4.85), 276(4.39) nm; IR (KBr) ν max 2955, 2925, 2867, 1722, 1643, 1460, 1376, 1275, 1153, 1075, 769 cm -1 ; CD (methanol) λ max (Δε): 260(-30.86), 305(+66.05), 305(-20.18) nm; 1 H and 13 C NMR spectral data are shown in Tables 2 and 3; High-resolution mass spectrometry m / z 617.4199 [M+H] + (Calculated for C 40 H 57 O 5 , 617.4206) and 639.4011 [M+Na] + (Calculated for C 40 H 56 O 5 Na, 639.4025).

[0083] Compound 8 (heterodesertone H): Brownish-yellow powder; Optical rotation [α] 25 D -3.8 (c 0.1, methanol); UV (methanol) λ max (logε): 271(3.80) nm; IR (KBr) ν max 3423, 2948, 2866, 1731, 1644, 1369, 1249, 1149, 1029 cm -1 ; CD (methanol) λ max(Δε): 226 (-3.40), 269 (+18.32), 410 (-2.72) nm; 1 H and 13 The \(^{1}H\) and \(^{13}C\) NMR spectral data are shown in Tables 2 and 3; high-resolution mass spectrometry m / z 835.5426 [M+Na] + (calcd for C 52 H 76 O 7 Na, 835.5489) and 1649.1006 [2M+Na] + (calcd for C 104 H 152 O 14 Na, 1649.1113).

[0084] Compound 9 (heterodesertone I): Brownish-yellow powder; optical rotation [α] 25 D +10.0 (c 0.1, methanol); UV (methanol) λ max (logε): 276 (2.97) nm; IR (KBr) ν max 3427, 2929, 2854, 1733, 1641, 1462, 1385, 1249, 1158, 1031, 753 cm -1 ; CD (methanol) λ max (Δε): 224 (+3.32), 293 (-9.79), 382 (+2.18) nm; 1 H and 13 The \(^{1}H\) and \(^{13}C\) NMR spectral data are shown in Tables 2 and 3; high-resolution mass spectrometry m / z 835.5390 [M+Na] + (calcd for C 52 H 76 O 7 Na, 835.5489).

[0085] Compound 10 (heterodesertone J): Brownish-yellow powder; optical rotation [α] 25 D +4.0 (c 0.1, methanol); UV (methanol) λ max (logε): 276 (3.14) nm; IR (KBr) ν max 3449, 2959, 1733, 1461, 1385, 1247, 1171, 1035 cm -1 ; 1 H and 13 The \(^{1}H\) and \(^{13}C\) NMR spectral data are shown in Tables 2 and 3; high-resolution mass spectrometry m / z 833.5280 [M+Na] +(Calculated value C 52 H 74 O 7 Na, 833.5332).

[0086] Table 1 1 H NMR data (chloroform-d, 400 MHz) of Compounds 1 - 5

[0087]

[0088]

[0089] Table 2 13 C NMR data (chloroform-d, 100 MHz) of Compounds 1 - 10

[0090]

[0091]

[0092] Table 3 1 H NMR data (chloroform-d, 400 MHz) of Compounds 6 - 10

[0093]

[0094]

[0095] Compound 11 (broussonetone A): white amorphous powder; optical rotation value [α] 25 D -2.9 (c 0.1, methanol); 1 H NMR (chloroform-d, 400 MHz) δ H : 0.56 (3H, d, J = 6.6 Hz, H 3 -16), 0.97 (3H, d, J = 6.6 Hz, H 3 -17), 0.88 (3H, s, H 3 -18), 0.88 (3H, s, H 3 -19), 1.38 (3H, s, H 3 -20), 0.94 (1H, H-5), 1.54 (1H, sep, J = 6.6 Hz, H-15), 2.65 (1H, s, H-14), 6.00 (1H, s, H-11); 0.80 (3H, s, H 3 -18′), 0.77 (3H, s, H 3 -19′), 0.92 (3H, d, J = 6.6 Hz, H 3 -16′), 1.00 (3H, d, J = 6.6 Hz, H3 -17'), 0.84 (3H, s, H 3 -20'), 0.71 (1H, brd, J = 12.5, H-5'), 1.59 (1H, sep, J = 6.6 Hz, H-15'), 3.02 (1H, s, H-14'), 3.71 (1H, s, H-11'). 13 C NMR (chloroform-d, 100 MHz) δ C : 40.3 (C-1), 18.6 (C-2), 41.1 (C-3), 34.2 (C-4), 54.3 (C-5), 18.8 (C-6), 30.5 (C-7), 48.6 (C-8), 169.1 (C-9), 42.2 (C-10), 123.2 (C-11), 202.5 (C-12), 78.9 (C-13), 46.4 (C-14), 38.3 (C-15), 16.7 (C-16), 15.8 (C-17), 33.7 (C-18), 22.4 (C-19), 22.1 (C-20), 36.5 (C-1'), 18.2 (C-2'), 41.6 (C-3'), 33.4 (C-4'), 52.9 (C-5'), 19.1 (C-6'), 42.2 (C-7'), 140.7 (C-8'), 139.3 (C-9'), 36.7 (C-10'), 59.2 (C-11'), 212.2 (C-12'), 76.1 (C-13'), 44.8 (C-14'), 34.3 (C-15'), 18.3 (C-16'), 17.4 (C-17'), 33.6 (C-18'), 21.7 (C-19'), 19.3 (C-20').

[0096] Example 2

[0097] The in vitro anti-proliferation effects of Compound 1-11 on 5 kinds of tumor cells (A-549, MCF-7, SW480, HL-60 and SMMC-7721) were determined by the MTS method. The results are shown in Table 4.

[0098] Table 4 Anti-tumor cell proliferation activities of Compound 1-11 (IC 50 μM) a,b

[0099]

[0100] a IC of other compounds 50 > 40 μM; b IC 50 values are expressed as mean ± deviation, n = 3; cAbietane diterpenoid 6,7-dehydroroyleanone; d Positive drug cisplatin.

[0101] Conclusion: Compounds 1-11 all had varying degrees of anti-proliferative effects on 5 types of tumor cells (A-549, MCF-7, SW480, HL-60, and SMMC-7721). Among them, compound 1 had significant in vitro anti-proliferative activity against 5 types of tumor cells, with IC 50 values ranging from 3.55 to 10.16 μM. Compounds 1 and 2 had significant anti-proliferative effects on A-549 lung cancer cells, with IC 50 values of 5.73 ± 0.23 and 12.18 ± 0.91 μM respectively, stronger than the positive drug cisplatin (IC 50 value of 13.84 ± 0.47 μM). Compound 4 had a significant anti-proliferative effect on MCF-7 breast cancer cells, with an IC 50 value of 13.46 ± 0.49 μM, stronger than the positive drug cisplatin (IC 50 value of 13.46 ± 0.49 μM).

[0102] Example 3

[0103] Zebrafish is a commonly used animal model in cancer research. The main advantage is that due to the lack of an adaptive immune system in zebrafish larvae, rejection reactions are not likely to occur, and xenograft tumor transplantation is easy. In this example, a zebrafish A549 lung cancer cell xenograft tumor model was constructed using the microinjection method to study the in vivo anti-tumor activity of compound 2, with etoposide as the positive drug. After the drug intervention ended, the tumor size and tumor metastasis in zebrafish were observed under a fluorescence microscope and photographed. The fluorescence intensity of the tumors in zebrafish was calculated using image processing software as an index to evaluate the tumor size. The number of tumor cell lesions was used as an index to evaluate the tumor metastasis situation.

[0104] Conclusion: Compound 2 significantly reduced the red fluorescence intensity of tumors in zebrafish in a dose-dependent manner ( Figure 2 A and B therein) and the number of tumor cell lesions ( Figure 2 A and C therein). Compound 2 had a significant inhibitory effect on the proliferation and metastasis of tumor cells, and its inhibitory effect was stronger than that of the positive drug etoposide at the same concentration. Even at a concentration of 1 μM, the inhibitory effect of compound 2 on zebrafish tumor metastasis was stronger than that of etoposide at a concentration of 10 μM.

[0105] It is easy for those skilled in the art to understand that the above are only the preferred embodiments of the present invention and are not used to limit the present invention. Any modifications, equivalent replacements, and improvements made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A terpene dimer compound, characterized in that: The structural formula of the terpene dimer compound is as follows: R1-R4 in formula I, R1-R6 in formula II, R1-R6 in formula III, R1-R6 in formula IV and R1-R2 in formula V are each independently selected from hydrogen, hydroxyl, methylene, carbonyl, halogen, acyl, acyl halide, nitro, cyano, benzene ring and indole ring.

2. The terpene dimer compound according to claim 1, characterized in that The formula I is compound 1 or compound 2, the formula II is compound 3, compound 4, compound 5 or compound 6, the formula III is compound 7, the formula IV is compound 8, compound 9 or compound 10, and the formula V is compound 11; 3. The method for separating terpene dimer compounds according to claim 2, characterized in that: The following steps are involved: (1) The roots of Xinjiang sage are dried and crushed, extracted with ethanol, and concentrated under reduced pressure to obtain a total extract; (2) suspending the total extract obtained in step (1) in water, extracting with petroleum ether, and concentrating under reduced pressure to obtain a petroleum ether extract; (3) The extract obtained in step (2) is extracted with a mixture of methanol / water to obtain a petroleum ether layer extract A and a methanol / water layer extract B; the petroleum ether layer extract A is mixed with silica gel and subjected to normal phase silica gel column chromatography, with the eluent being a petroleum ether / acetone gradient elution, combined with TLC spot detection, and the same components are combined to obtain 8 components A1-A8 with increasing polarity; component A3 is subjected to reverse phase C 18 Silica gel column chromatography, eluent is a mixture of methanol / water gradient elution, to obtain 8 sub-fractions A3a-A3h with decreasing polarity; component A3h is subjected to normal phase silica gel column chromatography, eluent is petroleum ether / acetone gradient elution, and the same fractions are combined to obtain 7 fractions A3h1-A3h7 with decreasing polarity; When component A3h5 was subjected to Sephadex LH-20 gel column chromatography with methanol as eluent, two subcomponents A3h5a and A3h5b with decreasing molecular weight were obtained; component A3h5b was separated by semi-preparative HPLC phenyl column with acetonitrile / water mixture as eluent to obtain compound 10; When component A3h7 was subjected to Sephadex LH-20 gel column chromatography with methanol as eluent, four subcomponents A3h7a-A3h7d with decreasing molecular weight were obtained; component A3h7b was separated by semi-preparative HPLC phenyl column with acetonitrile / water mixture as eluent to obtain compounds 8 and 9, respectively; When component A4 is subjected to reverse phase C 18 Silica gel column chromatography, with a gradient elution of a mixture of methanol / water as the eluent, yielded 16 subfractions A4a-A4p with decreasing polarity; component A4l was further chromatographed on a Sephadex LH-20 gel column and reversed phase C 18 Silica gel column chromatography yielded six subfractions A4l1-A4l6 with decreasing polarity; Component A4l6 was separated by semi-preparative HPLC phenyl column with a methanol / water mixture as eluent to obtain compound 5 and three other subcomponents A4l6a, A4l6c and A4l6d; When component A416a was purified by semi-preparative HPLC phenyl column with an acetonitrile / water mixture as the eluent, compound 11 was obtained; When component A416d was purified by semi-preparative HPLC phenyl column with an acetonitrile / water mixture as the eluent, compound 2 was obtained; When component A416c was purified by semi-preparative HPLC phenyl column with an acetonitrile / water mixture as the eluent, compound 1 and compound 6 were obtained; When component A4m was subjected to normal phase silica gel column chromatography and Sephadex LH-20 gel column chromatography, two sub-components A4m1 and A4m2 with decreasing molecular weight were obtained; component A4m2 was subjected to reverse phase C 18 Silica gel column chromatography, eluent methanol / water gradient elution, to obtain 6 sub-fractions A4m2a-A4m2f with decreasing polarity; component A4m2b was subjected to semi-preparative high performance liquid chromatography C 18 Column separation, the eluent is a mixture of acetonitrile / water / formic acid, to obtain compound 4 and compound 7 respectively; When component A4m2c was subjected to semi-preparative HPLC 18 Column separation with an acetonitrile / water mixture as the eluent gave compound 3.

4. Use of the terpene dimer compound as claimed in claim 1 or 2 in the preparation of anti-tumor drugs.

5. The use according to claim 4, characterized in that The anti-tumor drug is a drug for treating lung cancer, a drug for treating breast cancer, a drug for treating colon cancer, a drug for treating liver cancer and / or a drug for treating leukemia.

6. Use of compound 1 in the preparation of drugs for treating lung cancer, breast cancer, colon cancer, liver cancer and / or leukemia.

7. Use of Compound 1 and / or Compound 2 in the preparation of drugs for treating lung cancer; 8. Application of compound 4 in the preparation of drugs for treating breast cancer;

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