CN100590119C - Antineoplastic compound of red pineapple flower alkali A, preparation method and application thereof - Google Patents

Abstract

The invention pertains to the medical technology, in particular to an anti-tumor compound of incarvillea delavayi base A. The compound is a new alkaloid compound, which is extracted from the Chinese herbal medicine incarvillea delavayi. The precise mass number of the compound is 278.1546 (M plus H) <plus>, the molecular formula is C19H19NO by calculation; and the ultraviolet spectrum has strong absorption at 273nm and 249nm. Experiments show that the compound has good activity for inhibiting various tumor cells and can be used for the preparation of anti-tumor drugs.

Description

A kind of antineoplastic compound of red pineapple flower alkali A and its production and application

Technical field

The invention belongs to medical technical field, be specifically related to a kind of new anti-tumor biological bases compound of red pineapple flower alkali A (Delavatine A) that separation obtains from the plant red pineapple flower and its production and application.

Background technology

Tumour is to cause one of human main causes of death.Ministry of Health of China is announced urban and rural residents' major causes of death in 2006 a few days ago.Statistics shows that malignant tumour has become the primary cause of the death.As seen the prevention of tumour and treatment are very urgent.Pharmacological agent is one of main treatment means of tumour.At present, though developed tens kinds of antitumor drugs, effectively prolonged patient's life or improved patient's life quality.Some anti-tumor drugs treatment curative effect highly significant wherein is as the pharmacological agent acute leukemia of children etc.But the drug research of tumour and exploitation also face huge challenge, mostly are cell toxicity medicament as antitumor drug, and its side effect is obvious, has limited the performance of these curative effect of medication.Famous tumor aetiology man, member of Chinese Academy of Engineering's journey book an ancient unit of weight are thought: current medical science still is in the junior stage for the control of tumour.Chinese medicine has the basis of human toxicity experiment in several thousand, and perhaps Chinese medicine can be walked prostatitis in the world at anti-tumor aspect.The lead compound of seeking the anti-tumor activity of high-efficiency low-toxicity from natural product is the focus of new drug research always.

The purple section Chinese Incarvilla Herb platymiscium Incarvillea L. whole world of rivering bend has 14 kinds, China has 10 kinds, be distributed in the southwest more, areas such as northeast, North China, northwest in addition, wherein 5 kinds (Chinese Incarvilla Herb, Westerner's grass, red pineapple flower, Root of Delavay Incarvillea is hidden the POLO flower) hyoscine, in treatment hepatitis, the bacillary dysentery etc. of being mainly used among the people.The research of diagonal angle artemisia is less, and jade is bright late since nineteen ninety waits the people that Chinese Incarvilla Herb I.sinensis Lam. and Westerner's grass I.arguta (Royle) Royle have been carried out the research of chemical ingredients and pharmacologically active.Red pineapple flower (Incarvillea delavay) is that the Bignoniaceae Chinese Incarvilla Herb belongs to (Incarvillea L) plant, and perennial acaulescence draft mainly is distributed in Yunnan and Sichuan.The root of red pineapple flower is used as medicine as Root of Delavay Incarvillea.It is sweet, light, warm in nature to distinguish the flavor of.Invigorating qi and benefiting blood.Cure mainly insufficiency of vital energy and blood after being ill, dizziness and fatigue, postpartum few breast (" China's book on Chinese herbal medicine " editorial committee, " China's book on Chinese herbal medicine ", the .1999 of Shanghai science tech publishing house, the 7th fascicle, 426-427).

Separate from medicinal plant at present that to have antineoplastic activeconstituents be one of focus of chemist.And at present few to the chemical constitution study of red pineapple flower, so we have carried out the chemical constitution study of system to red pineapple flower, in the hope of from wherein separating the natural product that obtains having good antineoplastic activity.

Summary of the invention

The object of the present invention is to provide a kind of natural compounds and its production and application with good resistance tumor effect.

The present invention is a kind of new alkaloid compound that extraction separation obtains from the red pineapple flower that is grown in the Chinese yunnan Eryuan County, called after Delavatine A, and its chemical structural formula is as follows:

The preparation method of above-claimed cpd is as follows:

(1) extracts: the dry herb of red pineapple flower, pulverize, with 80% alcohol reflux 2-4 time, each 1.5-2.5 hour, the extracting solution concentrating under reduced pressure became medicinal extract, transfer Ph to 2-3 with HCl solution behind the thin up, filter, get filtrate and filter residue, filtrate adds NaOH solution and transfers Ph to 10-11, use chloroform extraction, obtain chloroform extract;

(2) separate: above-mentioned chloroform extract applying silicon plastic column chromatography, it with volume chloroform-acetone system gradient elution of 20: 1～2: 1, thin-layer chromatography detects, collection contains the flow point of red pineapple flower alkali A, again through the C18 reversed phase column chromatography, with weight ratio is 50: 100-70: 100 methanol-water carries out wash-out, and thin-layer chromatography detects, and gets the pure product of red pineapple flower alkali A.

Alkaloid compound DelavatineA of the present invention is a yellow oil, [α] ²⁰ _D+ 56 ° (c 0.05, CH ₃Cl); The bismuth potassium iodide reacting positive; By electron spray(ES) ion massspectrum signal m/z 278[M+H] ⁺, high resolution mass spectrum must the accurate total mass number of this compound is 278.1546[M+H] ⁺, calculate that its molecular formula is C ₁₉H ₁₉NO; UV spectrum has strong absorption at 273nm and 249nm. ¹H and ¹³The C nuclear magnetic resonance data sees Table 1.

Experiment shows that this compound thing has good inhibition activity to kinds of tumor cells, can be used for preparing antitumor drug.

Table 1Delavatine A's ¹H and ¹³The C nuclear magnetic resonance data

¹³C?and? ¹H?δppm?recorded?at?600MHz

J: coupling constant

Embodiment

The following examples can make those skilled in the art more fully understand the present invention, but do not limit the present invention in any way.

Embodiment 1:

Red pineapple flower (Incarvillea delavay) herb 3kg, with 80% ethanol 20L refluxing extraction 3 times, each 2 hours, united extraction liquid, concentrating under reduced pressure gets 60g medicinal extract, use 2%HCl behind the thin up, transfer PH to 2-3, filter, get filtrate and filter residue, filtrate adds 20%NaOH and transfers PH to 11, uses chloroform extraction, obtains chloroform extract; Concentrating under reduced pressure extraction position obtains crude product 25g.Crude product mixed with 50g silica gel mix sample, be added to the silicagel column top that has installed, with chloroform-acetone (volume ratio is 20: 1～2: 1) system's gradient elution, collection contains the flow point of Delavatine A, again through the C18 reversed phase column chromatography, carry out wash-out with methanol-water (weight ratio is 50%-70%), thin-layer chromatography detects, and gets the pure product 212mg of Delavatine A.

The cytotoxic activity experiment of embodiment 2:Delavatine A

1, experiment material

1.1, given the test agent

After Delavatine A dissolves with DMSO (Merck), add solution or uniform suspension that PBS (-) is made into 1000 μ g/ml, then with PBS (-) dilution that contains DMSO.

1.2, cell strain

A549 (human lung carcinoma cell)

LOVO (people's colon-cancer cell)

6T-CEM (human T cell leukemia cell)

HL-60 (human leukemia cell)

HepG-2 (human liver cancer cell)

1.3, nutrient solution

RPMI1640+15%NBS+ is two anti-

1.4, other materials

Full-automatic microplate reader: model: WellscanMK-2, production firm: Labsystems

Import 96 well culture plates etc.

2, test method

Mtt assay: it is 4～6 * 10 that the every hole of 96 orifice plates adds concentration ⁴The cell suspension 100 μ l of individual/ml put 37 ℃, 5%CO ₂In the incubator.Behind the 24h, add sample liquid, two multiple holes are established in 10 μ l/ holes, and 37 ℃, 5%CO ₂Effect 72h.Every hole adds the MTT solution 20 μ l of 5mg/ml, adds lysate behind the effect 4h, and put in the incubator in 100 μ l/ holes, and 570nm OD value is surveyed with the full-automatic microplate reader of MK-2 in the dissolving back.

3, test-results

The results are shown in Table 2, the result shows that Delavatine A is to human lung carcinoma cell, people's colon-cancer cell, the human T cell leukemia cell, human leukemia cell and human liver cancer cell all have better inhibited activity, Delavatine A has favorable anti-tumor effect, has the excellent development prospect.

Embodiment 3:DelavatineA is to the efficacy experiment of mouse S180 sarcoma (solid-type)

1, experiment material

1.1 given the test agent

Delavatine A uses the 0.5%CMC wiring solution-forming after with a small amount of tween-80 hydrotropy.

1.2 animal

Strain: Kunming mouse

The source: the The 2nd Army Medical College Experimental Animal Center provides.

Conformity certification number: Shanghai is moving closes the card word No. 107

Body weight: 18-20g

Sex: female.

1.3 transplanted tumor

Mouse S180 sarcoma is gone down to posterity by Shanghai Institute of Pharmaceutical Industry and to keep.

2, experimental technique

Get well-grown mouse S180 sarcoma ascites, dilute with 1: 4 with physiological saline, every mouse armpit subcutaneous vaccination 0.2ml, random packet is divided into control group, endoxan group (CTX group, 20mg/kg, ip * 7), LID-15-1 (5mg/kg) group, next day is played administration in the inoculation back, the administration volume is the 0.5ml/20g body weight, continuous irrigation stomach 7 days.Inoculate back 10 days and take off neck execution animal, dissect behind the title the weight of animals and get the knurl piece, claim knurl heavy.The result judges according to following formula:

3, experimental result

Delavatine A has significant tumor-inhibiting action.Experimental result sees Table 3.

Table 2Delavatine A is to the in-vitro multiplication restraining effect of human body tumour cell

Table 3DelavtineA is to the restraining effect of mouse S180 sarcoma

Compare with control group: * P＜0.05, * * P＜0.01.

Claims (3)

1, a kind of antineoplastic compound of red pineapple flower alkali A is characterized in that chemical structural formula is as follows:

2, the preparation method of antineoplastic compound of red pineapple flower alkali A as claimed in claim 1 is characterized in that concrete steps are as follows:

(1) extracts: the dry herb of red pineapple flower, pulverize, with 80% alcohol reflux 2-4 time, each 1.5-2.5 hour, the extracting solution concentrating under reduced pressure became medicinal extract, transfer pH to 2-3 with HCl solution behind the thin up, filter, get filtrate and filter residue, filtrate adds NaOH solution and transfers pH to 10-11, use chloroform extraction, obtain the chloroform part;

(2) separate: above-mentioned chloroform certain applications silica gel column chromatography, it with volume ratio chloroform-acetone system gradient elution of 20: 1～2: 1, thin-layer chromatography detects, collection contains stream part of red pineapple flower alkali A, again through the C18 reversed phase column chromatography, with weight ratio is 50: 100-70: 100 methanol-water carries out wash-out, and thin-layer chromatography detects, and gets the pure product of red pineapple flower alkali A.

3, the application of antineoplastic compound of red pineapple flower alkali A according to claim 1 in the preparation antitumor drug.