Terpenoid derivatives in Elephantopus scaber L. and their preparation methods and applications

Abstract

Terpenoid compounds in Elephantopus scaber, their preparation methods and applications belong to the field of pharmaceutical technology. Specifically, it involves three terpenoid compound derivatives extracted and isolated from the plant Elephantopus scaber L. of the genus Elephantopus in the Compositae family. Compounds 2 and 3 have good anti-acetylcholinesterase activity. The preparation method of the present invention is simple and easy to implement, with good reproducibility and high purity.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical technology, and particularly relates to three terpene compound derivatives prepared from Elephantopus scaber, their preparation methods, and the application of such compounds in the anti-Alzheimer's disease activity. Background Art

[0002] Elephantopus scaber is the dried whole herb of the plant Elephantopus scaber Elephantopus scaber L. in the genus Elephantopus of the Compositae family, also known as bitter Elephantopus and Elephantopus head. It is widely distributed in regions such as the Americas, Asia, and Africa, and is produced in Zhejiang, Jiangxi, Fujian, Hunan, Guangdong, Guangxi, Guizhou, and Yunnan in China.

[0003] Elephantopus scaber is bitter in taste and cold in nature, and has the effects of clearing heat and cooling blood, and detoxifying and promoting diuresis. It is used to treat colds, bacterial dysentery, gastroenteritis, tonsillitis, pharyngitis, nephritis edema, conjunctivitis, boils and other diseases. Modern research shows that Elephantopus scaber contains a variety of chemical components, including sesquiterpene lactones, triterpenoids, flavonoids, steroids, quinones, aryls, lipids, and others.

[0004] Regarding the compounds involved in the present invention and their effects on anti-acetylcholinesterase activity and the prevention or treatment of Alzheimer's disease, there have been no reports in patent or non-patent literature so far. Summary of the Invention

[0005] The purpose of the present invention is to solve the deficiencies of the prior art, and provides three terpene compounds with novel structures in Elephantopus scaber, their preparation methods, and their application in the preparation of drugs with anti-acetylcholinesterase activity and for the prevention or treatment of Alzheimer's disease.

[0006] To achieve the purpose of the present invention, the following technical solutions are adopted:

[0007] In the first aspect, the present invention provides three terpene derivatives.

[0008] The three terpene derivatives isolated from the plant Elephantopus scaber Elephantopus scaber L.] in the genus Elephantopus of the Compositae family have the following structures:

[0009] 。

[0010] In the second aspect, the present invention provides a preparation method for the above-mentioned terpene derivatives in Elephantopus scaber, and the preparation method includes the following steps:

[0011] Take the powder of the dried whole herb of Elephantopus scaber, combine the extraction solutions and concentrate to obtain an extract. The extract is extracted with petroleum ether, ethyl acetate and n-butanol, and the obtained components are separated by silica gel column chromatography with a gradient elution of dichloromethane-methanol system from 50:1 to 1:1 to obtain 5 components, Fr. A-E;

[0012] Use HP20 and ODS column chromatography to elute component Fr. A with an ethanol-water system from 10:90 to 50:50 to obtain 2 components, Fr. A1-Fr. A2;

[0013] Separate Fr. A1 and Fr. A2 by eluting with a petroleum ether-acetone system from 50:1 to 0:1 on silica gel column chromatography and using a mobile phase of acetonitrile-water on preparative reverse-phase high performance liquid chromatography to obtain Compound 1 and Compound 3;

[0014] Use HP20 and ODS column chromatography to elute component Fr. B with an ethanol-water system from 10:90 to 50:50 to obtain 3 components, Fr. B1-Fr. B3;

[0015] Separate Fr. B3 by eluting with a petroleum ether-acetone system from 50:1 to 0:1 on silica gel column chromatography and using a mobile phase of acetonitrile-water on preparative reverse-phase high performance liquid chromatography to obtain Compound 2.

[0016] As an alternative, in the above preparation method, the Elephantopus scaber used is Elephantopus scaber L. Elephantopus scaber L.]

[0017] As an alternative, in the above preparation method, take the dried whole herb of Elephantopus scaber and reflux extract it with 70% industrial ethanol 3 times, 2-3 h each time.

[0018] As an alternative, in the above preparation method, separate Fr.A1, Fr. A2 and Fr. B3 with an acetonitrile-water mobile phase of 50:50-20:80.

[0019] The obtained compounds are identified by systematic structure determination, and the results are as follows:

[0020] Use high-resolution mass spectrometry, one-dimensional NMR and two-dimensional NMR to identify the structures of Compounds 1-3, and the corresponding spectra and structure characterization data are as Figure 1-19 shown in Table 1.

[0021] Elephantopuscaber A(1): White amorphous powder. α +72.2 ( c 0.1, methanol); HRESIMS: [M+Na] + (​m / z ): 357.2031 was determined to have the molecular formula C 20 H 30 O4. By analyzing the 1 H NMR, 13 C NMR, HSQC spectrum, and HMBC spectrum of elephantopuscaber A, the structure of elephantopuscaber A was determined to be a new compound.

[0022] Elephantopuscaber B(2): White amorphous powder. α +35.2( c 0.1, methanol); HRESIMS: [M+Na] + ( m / z ): 399.1411 was determined to have the molecular formula C 20 H 24 O7. By analyzing the 1 H NMR, 13 C NMR, HSQC spectrum, and HMBC spectrum of elephantopuscaber B, the structure of elephantopuscaber B was determined to be a new compound.

[0023] Elephantopuscaber C(3): White amorphous powder. α -62.4 ( c 0.1, methanol); HRESIMS: [M+H] + ( m / z ): 441.3359 was determined to have the molecular formula C 29 H 45 O3. By analyzing the 1 H NMR, 13 C NMR, HSQC spectrum, HMBC spectrum, 1 H- 1 H COSY spectrum of elephantopuscaber C, the structure of elephantopuscaber C was determined to be a new compound.

[0024] The anti-Alzheimer's activities of the three new compounds described in the present invention were investigated. Compounds 2 and 3 exhibited good anti-acetylcholinesterase activities. Therefore, the new terpene compound derivatives described in the present invention have the prospect of being further developed into drugs for preventing and treating Alzheimer's disease.

[0025] ​​In a third aspect, the present invention provides a pharmaceutical composition comprising the terpenoid derivative prepared from Eleutherococcus scoparia or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier or excipient.

[0026] In a fourth aspect, the present invention further provides the use of the terpenoid derivatives prepared from Eleutherococcus elegans or pharmaceutical compositions comprising the terpenoids in the preparation of drugs for inhibiting acetylcholinesterase activity or preventing or treating Alzheimer's disease.

[0027] Compared with the prior art, the present invention has the following beneficial effects:

[0028] The advantages of the present invention are that the compounds are all new compounds, terpene compounds with novel structures, and compound 2 and compound 3 have better anti-Alzheimer's disease activity and are worthy of further development.

[0029] Table 1. Compounds 1-3 1 H NMR and 13 C NMR data (600 MHz, in CDCl3, δ in ppm)

[0030]

[0031] BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 HRESIMS of compound 1;

[0033] Figure 2 Compound 1 1 H-NMR spectrum;

[0034] Figure 3 Compound 1 13 C-NMR spectrum;

[0035] Figure 4 HSQC spectrum of compound 1;

[0036] Figure 5 HMBC spectrum of compound 1;

[0037] Figure 6 NOESY spectrum of compound 1;

[0038] Figure 7 HRESIMS of compound 2;

[0039] Figure 8 Compound 2 1 H-NMR spectrum;

[0040] Figure 9 For Compound 2 13 C-NMR spectrum;

[0041] Figure 10 HSQC spectrum of Compound 2;

[0042] Figure 11 HMBC spectrum of Compound 2;

[0043] Figure 12 " NOESY spectrum of Compound 2;

[0044] Figure 13 HRESIMS of Compound 3;

[0045] Figure 14 For Compound 3 1 H-NMR spectrum;

[0046] Figure 15 For Compound 3 13 C-NMR spectrum;

[0047] Figure 16 HSQC spectrum of Compound 3;

[0048] Figure 17 HMBC spectrum of Compound 3;

[0049] Figure 18 For Compound 3 1 H- 1 H COSY spectrum;

[0050] Figure 19 NOESY spectrum of Compound 3. Detailed implementation manners

[0051] The following examples are helpful for those skilled in the art to better understand the present invention, but do not limit the present invention in any way.

[0052] Example 1: Preparation of Compounds 1-3

[0053] Take the dried powder of the whole herb of Elephantopus scaber, combine the extraction solutions and concentrate to obtain an extract. The extract is extracted with petroleum ether, ethyl acetate and n-butanol, and the obtained components are subjected to silica gel column chromatography, and gradient elution is carried out with a dichloromethane-methanol system of 50:1 - 1:1 to obtain 5 components, Fr. A - E;

[0054] Use HP20 and ODS column chromatography to perform gradient elution on component Fr. A with an ethanol-water system of 10:90 - 50:50 to obtain 2 components, Fr. A1 - Fr. A2;

[0055] Compound 1 and compound 3 were obtained by eluting through silica gel column chromatography with a petroleum ether-acetone system of 50:1 - 0:1 and separating Fr. A1 and Fr. A2 using a mobile phase of acetonitrile-water on preparative reversed-phase high performance liquid chromatography;

[0056] The fraction Fr. B was eluted by gradient elution with an ethanol-water system of 10:90 - 50:50 using HP20 and ODS column chromatography to obtain three fractions Fr. B1 - Fr. B3;

[0057] Compound 2 was obtained by eluting through silica gel column chromatography with a petroleum ether-acetone system of 50:1 - 0:1 and separating Fr. B3 using a mobile phase of acetonitrile-water on preparative reversed-phase high performance liquid chromatography.

[0058] In the said preparation method, the Elephantopus scaber used is Elephantopus scaber Elephantopus scaber L.] of the genus Elephantopus in the family Compositae.

[0059] In the said preparation method, the dried whole herb of Elephantopus scaber was reflux-extracted 3 times with 70% industrial ethanol for 2 - 3 h each time.

[0060] In the said preparation method, Fr. A1, Fr. A2 and Fr. B3 were separated with an acetonitrile-water mobile phase of 50:50 - 20:80.

[0061] Example 2: Investigation of the anti-Alzheimer's disease activity of compounds 1 - 3

[0062] The compounds were dissolved in DMSO and diluted to 5 series of concentrations with potassium phosphate buffer (KH2PO4 / K2HPO4, 0.1 M, pH 8.0). The reaction system contained 25 μ μl of the test compound, 12.5 μ μl of the enzyme, 125 μ μl of DTNB. The mixture was refrigerated overnight at 4°C. 50 μ μl of ATCI was added and the reaction started. The absorbance was measured 3 times within 10 min after adding the enzyme at a wavelength of 412 nm. The Graph-Pad Prism program was used for non-linear regression analysis of the response concentration (logarithmic) curve to calculate the IC 50 value.

[0063] The experimental results are shown in Table 2.

[0064] Table 2: Inhibitory effects of the terpene compounds in the present invention on acetylcholinesterase

[0065]

[0066] Obviously, those skilled in the art can make various modifications and variations to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalent technologies, the present invention is also intended to include these modifications and variations.

Claims

1. A terpene derivative in Elephantopus scaber, characterized in that, It is any one of the following structures: 。 2. The preparation method of the terpene derivative in Elephantopus scaber described in claim 1, characterized in that, The preparation method includes the following steps: Take the dry whole herb powder of Elephantopus scaber and reflux extract it three times with 70% industrial ethanol for 2 - 3 h each time. Combine the extract and concentrate it to obtain an extract. The extract is extracted with petroleum ether, ethyl acetate and n-butanol, and the obtained components are subjected to silica gel column chromatography and gradient eluted with a dichloromethane-methanol system of 50:1 - 1:1 to obtain five components, Fr. A - E. The Elephantopus scaber used is Elephantopus scaber L. of the genus Elephantopus in the family Compositae Elephantopus scaber L. ; Using HP20 and ODS column chromatography, fraction Fr. A was eluted with an ethanol-water system of 10:90 - 50:50 in a gradient manner to obtain 2 fractions, Fr. A1 - Fr. A2; By silica gel column chromatography with a petroleum ether-acetone system of 50:1 - 0:1 for elution and using a mobile phase of 50:50 - 20:80 acetonitrile-water on preparative reverse-phase high performance liquid chromatography to separate Fr. A1 and Fr. A2, compound 1 and compound 3 were obtained; Using HP20 and ODS column chromatography, fraction Fr. B was eluted with an ethanol-water system of 10:90 - 50:50 in a gradient manner to obtain 3 fractions, Fr. B1 - Fr. B3; By silica gel column chromatography with a petroleum ether-acetone system of 50:1 - 0:1 for elution and using a mobile phase of 50:50 - 20:80 acetonitrile-water on preparative reverse-phase high performance liquid chromatography to separate Fr. B3, compound 2 was obtained.

3. A pharmaceutical composition, characterized in that, The pharmaceutical composition contains the terpene derivatives in Elephantopus scaber L. described in claim 1 or their pharmaceutically acceptable salts and a pharmaceutically acceptable carrier or excipient.

4. Use of the terpene derivatives in Elephantopus scaber L. described in claim 1 or their pharmaceutically acceptable salts or the pharmaceutical composition described in claim 3 in the preparation of a drug with anti-acetylcholinesterase activity.

5. Use of the terpene derivatives in Elephantopus scaber L. described in claim 1 or their pharmaceutically acceptable salts or the pharmaceutical composition described in claim 3 in the preparation of a drug for preventing or treating Alzheimer's disease.

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