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Taxus cuspidata endophytic fungus for high yield of 10-deacetylbaccatin III

An endophytic fungus and deacetylation technology, which is applied in the field of biopharmaceuticals and microbial technology, can solve the problems of few reports and low yields

Pending Publication Date: 2022-03-15
NORTHEAST FORESTRY UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, endophytic fungal studies on 10-deacetylbaccatin III (10-DAB), a semi-synthetic drug substance of docetaxel and paclitaxel, are generally neglected, with fewer reports and lower yields

Method used

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  • Taxus cuspidata endophytic fungus for high yield of 10-deacetylbaccatin III
  • Taxus cuspidata endophytic fungus for high yield of 10-deacetylbaccatin III
  • Taxus cuspidata endophytic fungus for high yield of 10-deacetylbaccatin III

Examples

Experimental program
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Effect test

Embodiment 1

[0021] Example 1: Isolation and identification of Penicillium oxalicum R8-3-4 in the present invention.

[0022] In this embodiment, Penicillium oxalicum (Penicillium oxalicum) R8-3-4 is isolated from the healthy Northeast Taxus in the Botanical Garden of Northeast Forestry University, and its sample is currently stored in the Key Laboratory of Forest Plant Ecology, Ministry of Education, Northeast Forestry University; The experimental steps are as follows:

[0023] (1) Preparation of Potato Dextrose Agar Medium (PDA): Wash and peel fresh ripe potatoes, weigh 200g of potatoes and cut into pieces, boil in water for 30min, add 20g of glucose, Agar powder 20g, add distilled water to make up to 1L, pH is natural, sterilize under high temperature and high pressure at 121℃ for 20min;

[0024] (2) Take a healthy Northeast Taxus plant, clean the plant surface with distilled water, rinse with 70% alcohol for 1min, rinse with 5% sodium hypochlorite solution for 5min, and rinse with 10%...

Embodiment 2

[0029] Example 2: Screening of 10-deacetylbaccatin III from fermentation culture of Penicillium oxalicum R8-3-4 strain in the present invention.

[0030] (1) Preparation of medium: potato 200g / L, glucose 20g / L, (NH 4 ) 2 SO 4 3.0g / L, MgSO 4 0.5g / L,KH 2 PO 4 0.5g / L, CuSO 4 0.10mg / L, salicylic acid 10mg / L, sodium acetate 8g / L and vitamin B 1 0.001g / L, pH6.0, sterilized at 121℃ for 15min at high temperature and high pressure;

[0031] (2) Get a number of fungal blocks with a diameter of about 5 mm in the northeast yew endophytic fungus R8-3-4 in a good growth state and inoculate them into 200 mL of PDB (potato dextrose broth) fermentation medium, place on a constant temperature shaker at 30 ° C, 160 rpm, Culture for 14 days;

[0032] (3) The mycelia is separated from the fermentation broth by filtration, the fermentation broth is extracted 3 times with an equal volume of ethyl acetate, the mycelium is washed and dried with deionized water, and then ultrasonically ext...

Embodiment 3

[0040] Example 3: Qualitative and quantitative analysis of 10-deacetylbaccatin III fermented by Penicillium oxalicum (Penicillium oxalicum) R8-3-4 strain in the present invention.

[0041] (1) Get a number of fungal blocks with a diameter of about 5 mm in the northeast yew endophytic fungus R8-3-4 in a good growth state and inoculate them into 200 mL of PDB (potato dextrose broth) fermentation medium, place on a constant temperature shaker at 30 ° C, 160 rpm, Culture for 14 days;

[0042] (2) Mycelia is separated from the fermentation broth by filtration, the fermentation broth is extracted 3 times with equal volumes of ethyl acetate, the mycelium is washed and dried with deionized water, and then ultrasonically extracted 3 times with ethyl acetate, and the organic phases are combined. The organic phase was removed by rotary evaporation to obtain a dry crude extract, which was dissolved in 1 mL of mass spectrometric methanol after removal of impurities by a silica gel column f...

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Abstract

The invention relates to endophytic fungus Penicillium oxalicum R8-3-4 capable of realizing high yield of 10-deacetylbaccatin III, which is separated from Taxus cuspidate, and the endophytic fungus Penicillium oxalicum R8-3-4 can be used for artificially synthesizing paclitaxel and docetaxel. The endophytic fungus Penicillium oxalicum R8-3-4 can be used for producing 10-deacetylbaccatin III with high yield. The invention reports that the penicillium fungus produces the 10-deacetylbaccatin III taxane active substance for the first time, the yield is 979.52 mu g / L, and the penicillium fungus has the advantages of short fermentation period and multiple product types, and has a better industrial prospect. The taxane active substance is stably produced by applying a magnetic immobilization technology for the first time, and a way is opened up for commercially producing the most important anti-cancer drug by fungal fermentation.

Description

technical field [0001] The invention belongs to the field of microbial technology and biopharmaceuticals, and particularly relates to a plant endophytic fungus isolated from Taxus chinensis. The liquid fermentation liquid of the strain can produce 10-deacetylbaccatin III at a high rate, and has the ability to artificially synthesize docetaxel Or paclitaxel provides sustainable resource effects. Background technique [0002] Cancer is the main cause of serious threats to human life and social development in the world. In recent years, the global cancer burden has continued to increase. According to statistics from the World Health Organization in 2018, there were 18.1 million new cancer patients worldwide, and about 9.6 million cancer deaths . China is a country with a large population and a high incidence of cancer. In 2018, the number of new cases in my country was 3.804 million, accounting for more than 20% of the number of new cancer cases in the world. In the past five ...

Claims

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Application Information

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IPC IPC(8): C12N1/14C12N11/14C12N11/10C12P17/02C12R1/80
CPCC12N11/14C12N11/10C12P17/02
Inventor 付玉杰李弘琨王欣宇郭娜寇萍
Owner NORTHEAST FORESTRY UNIVERSITY
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