Method for cell abduction generation of taxone with Chinese yew

A Chinese yew and cell induction technology is applied in the field of inducing the production of new taxane compounds to achieve the effects of releasing feedback inhibition, reducing costs, and simplifying the operation of extraction, separation and purification

Inactive Publication Date: 2008-12-10
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Abstract
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  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In the suspension culture system of Taxus chinensis, it has not been reported that the production of taxane

Method used

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  • Method for cell abduction generation of taxone with Chinese yew
  • Method for cell abduction generation of taxone with Chinese yew
  • Method for cell abduction generation of taxone with Chinese yew

Examples

Experimental program
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Effect test

preparation example Construction

[0034] (4) Preparation, purification and structural identification of new taxane compounds:

[0035] Collect the adsorbents that adsorbed taxane in several batches of experiments, and ultrasonically extract them at room temperature for 3 hours with 8 times the volume of methanol:dichloromethane 1:1 (v / v), rotate the extract, dissolve in chromatographically pure methanol, filter, and prepare do the preparation. The preparation work was completed at the Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Group 1804 - Medicinal Chemistry Research Group. The preparative chromatograph was watersDP4000, the preparative chromatographic column was CAPCELL PAK C18 column (Type: UG80, SIZE: 20×250mm, 5 μm), the mobile phase was acetonitrile:water=60:40, and the flow rate was 15ml / min.

[0036] Two relatively pure compounds were obtained through preparation, but it was not a single point detected by TLC, and continued to be purified by silica gel column chromatography. T...

Embodiment 1

[0042] Chinese yew suspension cells are used;

[0043] Culture conditions: Suspension culture in 100mL shake flask, using MS medium, add 0.2mg / L 2,4-dichlorophenoxyacetic acid, 0.5mg / L naphthaleneacetic acid, 0.5mg / L 6-benzyl adenine, 100mg / L L ascorbic acid and 30g / L sucrose. Inoculate 2 g of wet cells obtained by filtration into a shake flask containing 20 mL of the above-mentioned culture medium, and culture on a rotary shaker at 100 rpm at 25°C.

[0044] On the seventh day, MJA was added to a final concentration of 100 μM, and 100 g / L XAD-7 was added at the same time, the culture was continued to 21 days, and the cells were harvested and dried at 50°C. The cell density reached its highest level (12.7 g / L dry weight) on day 15. Tc analysis results are as follows (* represents the maximum value):

[0045] Training time

[0046] At the same time, two derivatives of Yunnan taxane Tc were obtained: 2,14-deacetyl Yunnan taxane C and 2-deacetyl Yunnan taxane C.

Embodiment 2

[0048] The culture conditions were the same as in Example 1, DHPJA was added on the 7th day to a final concentration of 100 μM, and 100 g / L XAD-7HP was added at the same time, and the culture was continued until 21 days. The cell density reached its highest level (dry weight 13.2 g / L) on the 12th day. The production capacity of Tc is shown in the table below.

[0049] Training time

[0050] At the same time, two derivatives of Yunnan taxane Tc were obtained: 2,14-deacetyl Yunnan taxane C and 2-deacetyl Yunnan taxane C.

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Abstract

The invention relates to a plant cell culture technology, in particular to a method for inducible production of taxane by utilization of Chinese yew cells. The method comprises the following steps that: the Chinese yew cells are placed into a commonly used culture medium for culture, and the cell inoculum size is between 50 and 200 grams per liter green weight; in the initial stage of the exponential growth of the cells, a revulsant with a concentration of between 1 and 2,000 micrometers, 50 to 200 grams of sterile absorbent and 10 to 50 grams of sugar are added into per liter of the culture medium; and in the intermediate stage of the exponential growth of the cells, the revulsant with a concentration of between 1 and 2,000 micrometers is added for the second time; and the revulsant is an abiological revulsant or a fungal elicitor. Because a large part of taxane compounds are externally secreted to the outsides of the cells and absorbed on the absorbent, the operation of separation and purification is greatly simplified, thereby the cost is reduced and the industrialized production is easy to realize. Moreover, because the method can produce novel taxane compounds by means of induction, the taxane is taken as a precursor compound of a first-line anticarcinogenic drug, namely paclitaxel, and the technology has important commercial application prospect.

Description

technical field [0001] The invention relates to plant cell culture technology, in particular to a method for greatly increasing the yield of taxane produced by Chinese yew cell culture and promoting its secretion, and at the same time inducing the production of new taxane compounds. Background technique [0002] As a first-line drug for the treatment of advanced ovarian cancer and breast cancer, paclitaxel is one of the few natural compounds with the most anticancer activity so far, with billions of dollars in sales each year, but the problem of drug sources is still a problem for paclitaxel researchers. The primary problem to be solved. There are not many types of plants that can produce paclitaxel in the world, and their growth is slow. The content of paclitaxel is extremely low, even in the bark with the highest content, it is only 1.5 / 10,000 on average. Obtaining paclitaxel by cutting down yew plants seriously damages the ecological balance and biodiversity, and will ev...

Claims

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Application Information

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IPC IPC(8): C12N5/04C12P17/02
Inventor 张卫高明波虞星炬金美芳
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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