Method for fermenting high-yield echinocandins B through aspergillus nidulans

An echinocandin and Aspergillus nidulans technology is applied in the field of using Aspergillus nidulans to ferment high-yield echinocandin B, can solve the problems of high viscosity of fermentation broth, hindered dissolved oxygen mass transfer, etc., and achieves good application prospects and fermentation products. Yield improvement effect

Active Publication Date: 2018-07-31
ZHEJIANG UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The object of the present invention is to provide a method for utilizing Aspergillus nidulans to ferment high-yield echinocandin B, and to provide a fermentation medium for controlling the morphology of Aspergillus nidulans fermentation strain A.nidulans ZJB 09223 to overcome echinocandin B During the fermentation process, the viscosity of the fermentation broth is too high, and the mass transfer of dissolved oxygen is hindered. Then, the addition of precursor amino acids to the medium is optimized to further increase the fermentation yield of echinocandin B.

Method used

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  • Method for fermenting high-yield echinocandins B through aspergillus nidulans
  • Method for fermenting high-yield echinocandins B through aspergillus nidulans
  • Method for fermenting high-yield echinocandins B through aspergillus nidulans

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1: Preparation of Echinocandin B (ECB) by Fermentation of Aspergillus nidulans A.nidulans ZJB 09223

[0029] Dig an area of ​​about 1cm with an inoculation shovel 2 A. nidulans ZJB09223 (CCTCC M 2012300, disclosed in patent application CN 103509840A, CN 201310478251.6) plate colony was inoculated into a 250ml Erlenmeyer flask containing 50ml of seed medium, and cultured at 25°C for 48h to obtain seed liquid. Seed medium (g / L) preparation: soybean cake powder 25, glucose 10, glycerin 10, solvent is distilled water, pH value is natural, sterilized at 115°C for 30 minutes.

[0030] In order to investigate the influence of a single carbon source in the medium on the fermentation level and cell morphology of ECB, five different fermentation mediums were designed, the volume of the shake flask was 50ml / 250ml, and the composition of the fermentation medium was as follows:

[0031] Fermentation medium (g / L) I (original medium): mannitol 97, glycerol 10.0, peptone 8.6, ...

Embodiment 2

[0039] Example 2: Fermentation of Aspergillus nidulans A.nidulans ZJB 09223 to prepare ECB

[0040] Dig an area of ​​about 1cm with an inoculation shovel 2 The plate colony of Aspergillus nidulans (A. nidulans) ZJB 09223 was inoculated into a 250ml Erlenmeyer flask containing 50.0ml of seed medium, and cultured at 25°C for 48h to obtain a seed solution. Seed culture medium is with embodiment 1.

[0041] In order to investigate the effect of different concentrations of methyl oleate on the fermentation level of ECB, five different fermentation media were designed, the liquid volume of the shake flask was 50ml / 250ml, and the composition of the fermentation media was as follows:

[0042] Medium (g / L) I: methyl oleate 54, peptone 8.6, soybean cake powder 40.0, peanut oil 20.0, L-threonine 2.1, L-ornithine 6.1, K 2 HPO 4 ·3H 2 O 3.2, CaCl 2 0.5, MgSO 4 ·7H 2 O 0.5, FeSO 4 ·7H 2 O 0.5, MnSO 4 ·H 2 O 0.2, CuSO 4 ·5H 2 O 0.6, the solvent is distilled water, the pH value ...

Embodiment 3

[0050] Example 3: Fermentation of Aspergillus nidulans A.nidulans ZJB 09223 to prepare ECB

[0051] Dig an area of ​​about 1cm with an inoculation shovel 2 The plate colony of Aspergillus nidulans (A. nidulans) ZJB 09223 was inoculated into a 250ml Erlenmeyer flask containing 50.0ml of seed medium, and cultured at 25°C for 48h to obtain a seed solution. Seed culture medium is with embodiment 1.

[0052] In order to investigate the effect of different particle additions on ECB fermentation level and cell morphology (see figure 2 ) influence, based on the culture medium in embodiment 3, design 5 different fermentation mediums, the liquid volume of shaking the flask is 50ml / 250ml, and the fermentation medium composition is respectively:

[0053] Medium (g / L) I (blank control): methyl oleate 114, peptone 8.6, soybean cake powder 40.0, peanut oil 20.0, L-threonine 2.1, L-ornithine 6.1, K 2 HPO 4 ·3H 2 O 3.2, CaCl 2 0.5, MgSO 4 ·7H 2 O 0.5, FeSO 4 ·7H 2 O0.5, MnSO 4 ·H 2...

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Abstract

The invention discloses a method for fermenting high-yield echinocandins B through aspergillus nidulans. The method includes the steps that a fermentation culture medium is inoculated with the aspergillus nidulans, culture is carried out at the temperature of 25 DEG C, fermentation liquor containing echinocandins B is obtained, the fermentation liquor is separated and purified, and echinocandins Bare obtained; by optimizing the varieties of carbon sources, adding particles and the like, the forms of mycelia in the fermentation process of the aspergillus nidulans are controlled, the mycelia are coccoid, and the problems that as the mycelia are too long during fermentation, the viscosity is high, dissolved oxygen is low, and mass transfer is insufficient are solved; by analyzing key metabolism components in the ECB synthesis process and optimizing a precursor adding strategy in the fermentation process, the yield of fermentation products is further increased by one time or above, and broad application prospects are achieved.

Description

(1) Technical field [0001] The invention relates to a method for using Aspergillus nidulans to ferment high-yield echinocandin B. (2) Background technology [0002] During the fermentation process of filamentous fungi, the mycelium morphology is affected by the growth conditions and culture environment. With the change of culture conditions, medium composition, etc., the mycelium morphology will appear in different types, such as non-branched mycelium, branched mycelia, etc. And mycelium with complex structure, mycelium mass, mycelium ball, etc., show different sizes, densities and surface structures. In addition, the internal structure of mycelium in different growth stages will also change with time. The mycelium of filamentous fungi usually presents three forms: flocculent, agglomerate and spherical in liquid submerged fermentation. The shape of mycelium has a great influence on the viscosity of the fermentation broth, and the products of the different mycelium shapes w...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/04C12R1/66
CPCC07K7/56C12P21/02
Inventor 郑裕国牛坤胡晓龙邹树平吴旭萍
Owner ZHEJIANG UNIV OF TECH
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