CN102977019A

CN102977019A - New C19 diterpenoid alkaloid in Aconitum hemsleyanum var. janyuanum and applications thereof in preparation of anticancer drugs

Info

Publication number: CN102977019A
Application number: CN2011102662258A
Authority: CN
Inventors: 高峰; 任丽红
Original assignee: Individual
Current assignee: Individual
Priority date: 2011-09-06
Filing date: 2011-09-06
Publication date: 2013-03-20

Abstract

The present invention discloses a new C19 diterpenoid alkaloid separated from Aconitum hemsleyanum var. janyuanum which is a new variant of Aconitum hemsleyanum Pritz., Aconitum, Ranunculaceae, and applications thereof in the preparation of anticancer drugs. By using roots of Aconitum hemsleyanum var. janyuanum which is a new variant of Aconitum hemsleyanum Pritz., Aconitum, Ranunculaceae are used as a raw material, a new C19 diterpenoid alkaloid is separated and identified by using a cation exchange resin method, a solvent method, silica gel column chromatography, centrifugal thin layer chromatography and spectroscopic methods. Anticancer experiments in vitro show that the compound has a significant inhibitory activity on liver cancer and non-small cell lung cancer, which lays foundation for the research and development of novel anticancer drugs from such plants.

Description

A kind of novel C in the rhizome of Chinese monkshood of Hanyuan 19Diterpene alkaloid and the application in the preparation antitumor drug thereof

Technical field

The present invention relates to a kind of a kind of novel C that obtains of from the rhizome of Chinese monkshood of Ranunculaceae Aconitum melon leaf rhizome of Chinese monkshood mutation Hanyuan, separating ₁₉Diterpene alkaloid and antitumor application thereof.

Background technology

Aconitum (Aconitum) plant belongs to Ranunculaceae (Ranunculaceae), is distributed in the north temperate zone, about 350 kinds of the whole world, and China has 170 kinds approximately, mainly is distributed in the alpine belt of Northern Yunnan, western Sichuan and Eastern Tibet.The melon leaf rhizome of Chinese monkshood (Aconitum hemsleyanum Pritz.) claims again rattan crow, radix aconiti agrestis, crow poison, salted aconite root etc., mainly is distributed in the ground such as Sichuan, the north, Hubei, Hunan, in growth and the height above sea level 1700-2200 mountain region woods or in the shrub.The Hanyuan rhizome of Chinese monkshood (Aconitum hemsleyanum var.hanyuanum) is a new variant of the Yaan special product melon leaf rhizome of Chinese monkshood.Modern pharmacology studies show that its main active ingredient is C ₁₉The type diterpene alkaloid has widely physiologically active, is mainly manifested in anti-inflammatory, analgesia, the aspect such as antitumor.Therefrom exploitation successfully goes on the market without addicted analgesics lappaconitine, bulleyaconitine A and 3-acetylaconitine.

Yet there are no from extracting, isolate the C shown in structural formula (I) the rhizome of Chinese monkshood plant of Hanyuan ₁₉The report of diterpene alkaloid and uses thereof aspect.

Summary of the invention

The invention provides a kind of antitumor drug of structure novel, or as precursor or the lead compound of cancer therapy drug, have good DEVELOPMENT PROSPECT.

The object of the present invention is to provide a kind of C of novel texture ₁₉Type diterpene alkaloid and at the medicine of preparation prevention or medicine for treating tumor thing tumour, or as the precursor of cancer therapy drug or the application in the lead compound.

The invention provides the C shown in the following structure ₁₉The type diterpene alkaloid:

The object of the present invention is to provide a kind of pharmaceutical composition that comprises described diterpene alkaloid and pharmaceutically acceptable carrier.

The present invention uses the hydrochloric acid diafiltration, use the cation exchange resin processes extract total alkaloids, use solvent method that total alkaloids has been carried out rough segmentation according to the difference of polarity, then mainly adopt solvent method, silica gel column chromatography, centrifugal thin layer chromatography to carry out the total alkali separation and obtain structural formula (I) compound, it comprises following concrete steps:

(1) gets the Hanyuan rhizome of Chinese monkshood (Aconitum hemsleyanum var.hanyuanum) dried root, pulverize;

(2) with 0.05mol/L HCl diafiltration, percolate is used the cation exchange resin processes extract total alkaloids, then soak resin with 10% ammoniacal liquor and make its alkalization, use ethanol elution, the collection ethanol eluate;

(3) with the elutriant decompression, concentrated, be acidified to pH 1 with 10%HCl again, suction filtration is collected filtrate, obtains the total alkaloids extracting solution, uses ether continuous backflow method to extract filtrate, and last concentrate drying obtains structural formula (I) compound.

Structural formula (I) C ₁₉The diterpene alkaloid compound is white amorphous powder, and molecular formula is C ₃₃H ₄₅NO ₁₁, its nuclear magnetic data such as following table 1:

Table 1

^a?Recorded?in?CDCl ₃， ¹H-NMR?at?400MHz， ¹³C-NMR?at?100MHz，δin?ppm，and?Jin?Hz

Through experimental results demonstrate that compound of the present invention has significant anti-tumor activity, especially the aspects such as liver cancer, lung cancer, cancer of the stomach, large bowel cancer and melanoma have good DEVELOPMENT PROSPECT.

Accompanying drawing is each hydrocarbon signal 2D-NMR (1H-1HCOSY, HMQC, HMBC) correlationship of compound of the present invention.

Embodiment:

Embodiment 1

Get the Hanyuan rhizome of Chinese monkshood (Aconitum hemsleyanum var.hanyuanum) dried root, pulverize; With 0.05mol/L HCl diafiltration, percolate is used the cation exchange resin processes extract total alkaloids, then soak resin with 10% ammoniacal liquor and make its alkalization, use ethanol elution, the collection ethanol eluate; With the elutriant decompression, concentrated, being acidified to the pH value with 10%HCl again is 1, and suction filtration is collected filtrate, obtains the total alkaloids extracting solution, uses ether continuous backflow method to extract filtrate, and last concentrate drying obtains structural formula (1) compound, and its nuclear magnetic data sees Table 1.

Embodiment 2

The anti-tumor activity experiment:

(1) inoculating cell: collect logarithmic phase HCT-8, HePG2, BGC-823, A549, A375 experimental cell, adjust concentration of cell suspension and be made into the individual cells suspension, be inoculated into respectively 96 orifice plates with every hole 1000-10000 cell, every pore volume 100 μ L.

(2) cell cultures: 5%CO ₂, cultivated 3-5 days for 37 ℃.If novel C ₁₉Diterpene alkaloid (C ₃₃H ₄₅NO ₁₁) medicine gradient 5-7 (with perfect medium configuration 2 * diluent), every hole 100 μ L drug dilution liquid; Each gradient (group) is established 3-5 multiple hole.

(3) quality: cultivate after 3-5 days, every hole adds 20 μ L MTT solution (5mg/ml, i.e. 0.5%MTT), continues 5%CO ₂, 37 ℃ of cell culture incubators are hatched 4h, stop cultivating, and careful the suction abandoned supernatant liquor in the hole.Every hole adds 150 μ L DMSO, and concussion 10min fully dissolves crystallisate.

(4) colorimetric: select the 570nm wavelength, detect each hole absorbance value at enzyme-linked immunosorbent assay instrument, the record result.Take the time as X-coordinate, light absorption value is that ordinate zou is drawn cell growth curve.

(5) drawing and calculating IC ₅₀: take drug concentration as X-coordinate, inhibiting rate is the ordinate zou mapping, drug concentration when drawing 50% inhibiting rate.

Mtt assay is studied the new compound anti tumor activity in vitro, found that, its to liver cancer, lung cancer and melanoma all tool suppress significantly active, antitumour activity is as shown in table 2.

Table 2:Anti-tumor Activity of Hanyuanine

Claims

1. novel C ₁₉Diterpene alkaloid, (I) is as follows for its structural formula:

2. one kind prepares C claimed in claim 1 ₁₉The method of diterpene alkaloid, it comprises following steps:

(3) with the elutriant decompression, concentrated, being acidified to the pH value with 10%HCl again is 1, and suction filtration is collected filtrate, obtains the total alkaloids extracting solution, uses ether continuous backflow method to extract filtrate, and last concentrate drying obtains structural formula (1) compound.

3. the pharmaceutical composition that comprises the described diterpene alkaloid of claim 1 and pharmaceutically acceptable carrier.

4. the described C of claim 1 ₁₉The application of diterpene alkaloid in preparation prevention or medicine for treating tumor thing.

5. application according to claim 3 is characterized in that described tumour is solid tumor.

6. application according to claim 4 is characterized in that described solid tumor comprises liver cancer, lung cancer, cancer of the stomach, large bowel cancer and melanoma.

7. application according to claim 6 is characterized in that described lung cancer is nonsmall-cell lung cancer.

8. diterpene alkaloid claimed in claim 1 is in conduct prevention or the precursor of medicine for treating tumor thing or the application in the lead compound, and described tumour is liver cancer, lung cancer, cancer of the stomach, large bowel cancer or melanoma.

9. according to claim 8 application is characterized in that described lung cancer is nonsmall-cell lung cancer.