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Production method for increasing yield of D-biotin

A production method and biotin technology, applied in the field of fermentation engineering, can solve the problems of many by-products with similar structures, low D-biotin yield, and inability to meet industrial production

Pending Publication Date: 2022-05-13
ZHEJIANG SHENGDA BIO PHARM +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0033] As mentioned above, at present, the vast majority of commercialized D-biotin is still prepared by chemical synthesis (high cost, high safety hazard, complex process, many by-products with similar structures, heavy environmental burden, etc.) , and a small amount is prepared from animals and plants by direct extraction methods, including enzyme extraction, organic solvent-assisted extraction, etc. (low extraction rate, low safety, etc.). low cost, safer, simple process, and environment-friendly), but because the yield of D-biotin produced by existing fermentation methods is generally low, most of them are still in the research and development stage, and the domestic fermentation level has a large gap compared with the international leading level. Satisfy industrialized production, therefore, need to develop a kind of improved microbial fermentation method that improves D-biotin output, to break the technical barrier of existing chemical synthesis, have very important meaning to improving existing fermentation process, on the other hand, also can Meet the needs of industrial scale-up

Method used

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  • Production method for increasing yield of D-biotin

Examples

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Embodiment 1

[0100] The method of embodiment 1 fermentative production D-biotin

[0101] The Pseudomonas mutans genetically engineered bacteria selected in this example are genetically engineered bacteria containing the DNA sequences shown in SEQ ID NO.1-6.

[0102] The genetically engineered bacteria uses Pseudomonas mutans ATCC31014 as the starting strain for the construction of genetic engineering, and the name of the engineered bacteria is PM / WIABFD, derived from the application number CN201910591872.2, and the name is "Methods for Promoting Biotin Synthesis, "Recombinant Cell and Genetically Engineered Bacteria Promoting Biotin Synthesis" is a Chinese invention patent application, and its specific construction method is derived from Example 1 and Example 2 of the specification.

[0103] The medium composition is as follows:

[0104] Seed medium (g / L): glucose 20, peptone 20, yeast extract 10, Na 2 HPO 4 12H 2 O 18. KH 2 PO 4 4.05, (NH 4 ) 2 SO 4 2. MgSO 4 ·7H 2 O 0.8, pH 7....

Embodiment 2

[0119] The method of embodiment 2 fermentative production D-biotin

[0120] Seed medium (g / L): glucose 21, yeast powder 19, yeast extract 10, Na 2 HPO 4 12H 2 O 18. KH 2 PO 4 4. (NH 4 ) 2 SO 4 2. MgSO 4 ·7H 2 O 0.7, pH 7.0.

[0121] Fermentation medium (g / L): glycerol 22, glucose 21, peptone 19, (NH 4 ) 2 SO 4 2. NaH 2 PO 4 15. K 2 HPO 4 ·3H 2 O 3 , MgSO 4 ·7H 2 O 0.6, FeSO 4 ·7H 2 O 0.01, pimelic acid 1, pH 7.0.

[0122] The bacterial classification preserved in glycerin (bacterial classification: identical with the genetically engineered bacterium of embodiment 1), inserts in the seed culture medium, at 29 ℃, under the condition of 210r / min, shaking table culture, until OD reaches 2.0, Can be made into seed solution.

[0123] Transfer the cultured seed solution to a 10L mechanically stirred fermenter containing 6L fermentation medium with an inoculum size of 1.2% (volume ratio). During the fermentation process, the temperature is controlled at 28°C an...

Embodiment 3

[0125] The method of embodiment 3 fermentative production D-biotin

[0126] Seed medium (g / L): glucose 18, peptone 20, yeast extract 9, NaH 2 PO 4 12. K 2 HPO 4 ·3H 2 O2, NH 4 Cl4, MgSO 4 ·7H 2 O 0.9, pH 7.1.

[0127] Fermentation medium (g / L): glycerin 20, glucose 22, yeast extract 23, NH 4 NO 3 3. NaH 2 PO 4 15. K 2 HPO 4 ·3H 2 O 3.5, MgSO 4 ·7H 2 O 0.7, FeSO 4 ·7H 2 O 0.01, pimelic acid 1.1, pH 7.0.

[0128] The bacterial classification preserved in glycerol (bacterial classification: the same as the genetically engineered bacteria in Example 1) was inserted into the seed medium, and cultured on a shaking table at 28° C. and 220 r / min until the OD reached 2.1. Can be made into seed solution.

[0129] Transfer the cultivated seed solution to a 10L mechanically stirred fermenter containing 6L fermentation medium with an inoculum size of 1.3% (volume ratio). During the fermentation process, the temperature is controlled at 29°C and the dissolved oxygen is ...

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Abstract

The invention relates to a production method for increasing the yield of D-biotin, and belongs to the technical field of biology. The invention discloses a production method for increasing the yield of D-biotin, which adopts pseudomonas mutabilis genetically engineered bacteria, takes glucose and glycerol as composite carbon sources, adopts a two-stage or three-stage temperature control strategy and combines an ethanol feeding process to carry out rapid fermentation production of D-biotin, the yield is increased by 21%-170% compared with that of the original process, and the yield of D-biotin is increased by 20%-30% compared with that of the original process. And the period is shortened by 13%-28%. The invention provides a simple, safe and efficient D-biotin fermentation production method, the existing production process steps are changed, the production intensity of the D-biotin is obviously improved, the fermentation period is shortened, the cost is saved, the emission of fermentation waste liquid and waste gas is reduced, the environmental pollution is reduced, in addition, the process is simple and easy to amplify, and the production cost is reduced. The industrial production is easy to realize.

Description

technical field [0001] The invention belongs to the technical field of fermentation engineering, and in particular relates to an improved method of a liquid fermentation method for producing D-biotin. Background technique [0002] Biotin belongs to the water-soluble B vitamins, also known as vitamin B 7 , vitamin H, coenzyme R (Coenzyme R, CoR), is a ring-shaped substance with a structure ring of thiophenol (Thiophene), this chemical substance has eight isomers, but only dextrorotatory D-biotin exists in nature, and only it has vitamin activity. It is one of the essential vitamins for the nutrition of animals, plants and microorganisms, a coenzyme for carboxylation and decarboxylation reactions, a necessary substance for the synthesis of vitamin C, and an indispensable substance for the normal metabolism of fat and protein. It plays an essential role in normal growth and development, skin health, hair and bone health. Therefore, it is widely used in food, medical, multidi...

Claims

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Application Information

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IPC IPC(8): C12P17/18C12N1/21C07D495/04C12R1/38
CPCC12P17/18C12N1/20C07D495/04
Inventor 郭凤柱李飞龙周斌梅吉雷李光进况嘉铀江根强朱勇刚徐志南赵佳润
Owner ZHEJIANG SHENGDA BIO PHARM