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Method for increasing yield of anidulafungin precursor compound Echinocandin B

A precursor compound, the technology of Anifungin, applied in the field of increasing the yield of Echinocandin B, the precursor compound of Anifungin, can solve the problems of low level of fermentation preparation and achieve the effect of improving the level of fermentation

Active Publication Date: 2014-01-15
ZHEJIANG UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] At present, the main problem is that the level of fermentation preparation of domestic ECB is still low. Therefore, it is of great significance for the industrial production of Anifungin to optimize the fermentation process and increase the yield of ECB by means of fermentation metabolism regulation.

Method used

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  • Method for increasing yield of anidulafungin precursor compound Echinocandin B
  • Method for increasing yield of anidulafungin precursor compound Echinocandin B
  • Method for increasing yield of anidulafungin precursor compound Echinocandin B

Examples

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Embodiment 1

[0025] The strain used in the examples of the present invention is Aspergillus nidulans ZJB09223 (CCTCC NO: M2012300). This strain was deposited in the China Type Culture Collection on July 25, 2012, and has been filed separately as a new strain.

[0026] (1) Aspergillus nidulans ZJB09223 seed culture:

[0027] Take a ring of dark green colonies from the PDA slant culture medium and connect it to the seed culture medium. The culture temperature is 25°C, the shaker speed is 220r / min, and the culture is 2 days to obtain the seed liquid; the seed culture medium has the following composition :Glucose 10g / L, glycerol 10g / L, cottonseed powder 25g / L, solvent is water, pH6.8~7.0, sterilized at 115℃ for 30min;

[0028] (2) Aspergillus nidulans ZJB09223 fermentation culture:

[0029] The composition of the fermentation medium is as follows: peanut oil 20g / L, glycerol 10g / L, peptone 10g / L, L-proline 2g / L, mannitol 90g / L, soybean meal 40g / L, K 2 HPO 4 ·3H 2 O8g / L, MgSO 4 ·7H 2 O0.5g / L, MnSO 4 ·H ...

Embodiment 2

[0032] According to the results of Example 1, we chose to change the temperature during the culture process. The experiment started with the fermentation culture at 25°C, 28°C, and 37°C, and then adjusted the temperature after 3 days, 6 days, and 9 days. The adjustment method is: adjust 28°C and 37°C to 25°C and cultivate until the end of fermentation; adjust 25°C to 28°C to cultivate until the end of fermentation, and determine the ECB content. The results are shown in Table 1. High temperature (28~37°C) cultivation and subsequent low temperature (25°C) cultivation can greatly increase ECB production. Using 37°C for the first 3 days and 25°C for the next 9 days can increase the ECB fermentation level to 1237mg / L, which is 69.9% higher than the constant temperature culture at 25℃.

[0033] Table 1: The influence of different temperature adjustment methods and adjustment time on ECB output

[0034]

Embodiment 3

[0036] It is reported in related literature that proline plays an important role in the growth and metabolism of fungal microorganisms. The experiment examined the effects of different addition times and different concentrations of proline on the fermentation results.

[0037] The composition of the original fermentation medium of ECB in the experiment is: peanut oil 20g / L, glycerol 10g / L, peptone 10g / L, L-proline 5g / L, mannitol 90g / L, soybean meal 40g / L, K 2 HPO 4 ·3H 2 O8g / L, MgSO 4 ·7H 2 O0.5g / L, MnSO 4 ·H 2 O0.1g / L, FeSO 4 ·7H 2 O0.05g / L, CaCl 2 0.3g / L, solvent is water, pH7.0. The seeds cultured for 2 days were put into the fermentation medium at a 10% inoculum amount, and the culture time was 12 days under the conditions of 25°C and a shaker rotation speed of 220r / min.

[0038] Taking the above fermentation medium as a control, proline was added to the fermentation broth at different times to make the final concentrations of 2g / L, 5g / L, and 8g / L, respectively, to investigate t...

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Abstract

The invention provides a method for increasing the yield of an anidulafungin precursor compound Echinocandin B (ECB). The method comprises the following steps: preparing a fermentation culture medium for culturing Aspergillus nidulans for synthesizing ECB, inoculating aspergillus nidulans, culturing for 3 days at 28-37 DEG C, and continuously culturing at 25 DEG C till the fermentation is accomplished. Compared with constant temperature culture, the method adopts the strategy that the ECB is cultured at 37 DEG C at the early three days and is cultured at 25 DEG C at the later 9 days, so that the yield of the ECB is increased to be 1,237mg / L from 728mg / L; in addition, the yield of the ECB is increased to be 850mg / L from 540mg / L by adding 2g / L proline on the sixth day; and the yield of the ECB can be respectively increased by 21.9% and 55.2% by adding 2g / L threonine and 2g / L ornithine into the culture medium.

Description

(1) Technical field [0001] The invention relates to a culture method for increasing the yield of anidulafungin precursor compound Echinocandin B. (2) Background technology [0002] In recent years, organ transplantation has led to the massive use of immunosuppressive agents, chemotherapy and the application of more aggressive medical methods, and many other reasons have led to an increase in immunocompromised patients, and the incidence of fungal infections has increased significantly, especially the incidence of deep fungal infections. The rate and fatality rate are increasing year by year. According to statistics from the World Health Organization (WHO) in 2004, the number of systemic fungal infections worldwide is 335,000 annually. The most common is systemic candidiasis, which accounts for about 70% of all such infections, and the mortality rate caused by it is as high as 40%. With the continuous advancement of medicine, after the 1990s, various antifungal drugs have come o...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/04C12R1/66
Inventor 郑裕国牛坤邹树平毛健钟伟沈寅初
Owner ZHEJIANG UNIV OF TECH
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