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Method for promoting microbes to synthesize ascomycin by using exogenous additive and preparation of seed liquor and culture medium

A technology of exogenous additives and ascomycin, applied in the field of microbial fermentation, can solve problems such as gaps, and achieve the effects of low additional input, promotion of metabolism, and enhancement of accumulation

Inactive Publication Date: 2013-05-29
TIANJIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In 2007, Indian scholar Parveen et al. applied for patent WO2007 / 029082 and reported that the unit of ascomycin reached 350-400 mg / L, but domestic research on ascomycin is still in its infancy, and the fermentation titer is generally 150-200 mg / L. Compared with foreign fermentation titers, there is a big gap
However, the combination of exogenous additives soybean oil and isoleucine and shikimic acid has not been seen in the optimization of ascomycin fermentation medium.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Take the mature slant spores of Streptomyces hygroscopicus subspecies (ATCC14891) at room temperature to make a spore suspension with sterile water; Glucose 32g / L, peptone 5g / L, yeast powder 5g / L, calcium carbonate 1g / L, initial pH 7.0) in a 250mL shaker flask, cultured in a shaker at 30°C and 180rpm for 48h; , added to the fermentation medium (composition and content: starch 22g / L, dextrin 40g / L, gluten powder 3g / L, peptone 4g / L, yeast powder 12g / L, corn steep liquor 4g / L, cold Pressed bean cake powder 4g / L, dipotassium hydrogen phosphate 0.5g / L, ammonium sulfate 1g / L, magnesium sulfate 0.5g / L, calcium carbonate 0.5g / L, shikimic acid 0.5g / L, initial pH7.0) 7.5LDE fermenter; fermentation conditions: 30°C, controlled ventilation 0.6v / v / m, rotating speed 500rpm, pH 7.0, fermented for 192h. The concentration of ascomycin in the fermentation broth was detected by HPLC. Ascomycin fermentation unit increased from 170mg / L to 265mg / L, an increase of 55.9%.

Embodiment 2

[0019]Take the mature slant spores of Streptomyces hygroscopicus subspecies (ATCC14891) at room temperature to make a spore suspension with sterile water; Glucose 32g / L, peptone 5g / L, yeast powder 7g / L, calcium carbonate 2g / L, initial pH 7.0) in a 250mL shake flask, cultured in a shaker at 30°C and 180rpm for 48h; , added to the fermentation medium (composition and content: starch 18g / L, dextrin 42g / L, gluten powder 2g / L, peptone 6g / L, yeast powder 10g / L, corn steep liquor 5g / L, cold Pressed bean cake powder 4g / L, dipotassium hydrogen phosphate 1.5g / L, ammonium sulfate 0.5g / L, magnesium sulfate 0.8g / L, calcium carbonate 1.2g / L, initial pH7.0) automatic 7.5LDE fermentation tank. Add 0.5g / L shikimic acid after 36h of fermentation. Fermentation conditions: 30°C, controlled ventilation 0.6v / v / m, rotation speed 500rpm, pH 7.0, fermented for 180h. The concentration of ascomycin in the fermentation broth was detected by HPLC. Ascomycin fermentation unit increased from 170mg / L to 2...

Embodiment 3

[0021] Take the mature slant spores of Streptomyces hygroscopicus subspecies (ATCC14891) at room temperature to make a spore suspension with sterile water; Glucose 29g / L, peptone 4.5g / L, yeast powder 6.5g / L, calcium carbonate 1.5g / L, initial pH7.0) in a 250mL shaker flask, cultured in a shaker at 26°C and 220rpm for 48h; with 10% The amount of inoculum was added to the fermentation medium (composition and content: starch 21g / L, dextrin 41g / L, gluten powder 2.2g / L, peptone 5.5g / L, yeast powder 11g / L, corn steep liquor 5.5g / L, cold-pressed bean cake powder 5.5g / L, dipotassium hydrogen phosphate 0.8g / L, ammonium sulfate 0.8g / L, magnesium sulfate 0.6g / L, calcium carbonate 0.7g / L, isoleucine 2g / L L, in an automatic 7.5LDE fermenter with an initial pH of 7.0). Fermentation conditions: 26°C, 1.0v / v / m aeration, 500rpm rotation speed, pH 7.0, 144h fermentation. The concentration of ascomycin in the fermentation broth was detected by HPLC. Ascomycin fermentation unit increased from 1...

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Abstract

The invention discloses a method for promoting microbes to synthesize ascomycin by using an exogenous additive and a preparation of a culture medium, and belongs to the technical field of biologic synthesis of ascomycins. The method is characterized in that, on the basis of the characteristic that the ascomycin biologic synthesis process refers to the assembly of various precursors, at the beginning of the aerobic fermentation or in the process of aerobic fermentation, 0.5-3.0 g of shikimic acid, 0.5-3.0 g of isoleucine and 1-5 g of soybean oil are added in every one liter of a culture mediumfor enhancing the accumulation of ascomycin synthesis precursors DHCHC, malonyl and methyl malonyl, promoting the metabolism of the ascomycin synthesis pathway, thereby improving the yield of the ascomycin. The method provided by the invention is simple to operate and is low in cost because no extra device and labor is increased and only low additional investment is input; in the step of fermenting and culturing streptomyces hygroscopicus ascomycota subspecies, the fermenting unit of the ascomycin is improved by 55.9-114.7% as compared with a control group.

Description

technical field [0001] The invention belongs to the technical field of microbial fermentation, and in particular relates to a method for exogenous additives to promote the synthesis of ascomycin by microorganisms and the preparation of culture medium. Background technique [0002] Ascomycin (FK-520) is a macrolide antibiotic composed of a twenty-three-membered ring produced by the fermentation of Streptomyces hygroscopicus var.ascomyceticus. Its chemical name is 17α-ethyl-1β, 14α-dihydroxy-12β[2-(4α-hydroxy-3β-methoxy-1β-1-(E)-methylvinyl)]-23α, 25β- Dimethoxy-13α, 19, 21α, 27β-tetramethyl-11,28-dioxa-4-azatricyclo[22,3,1,O]octadecano-18-2,3 , 10,16-tetraketone, monohydrate. Relative molecular mass is 822.5, molecular formula C 44 h 69 NO 12 ·H 2 O. Soluble in methanol, ethanol, acetone, ethyl acetate, ether, benzene, soluble in tert-butyl methyl ether, slightly soluble in water, cyclohexane, n-hexane, insoluble in heptane. [0003] Ascomycin is an ethyl analog of ta...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P17/18C12R1/55
Inventor 闻建平齐海山
Owner TIANJIN UNIV
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