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Hyphomicrobium sp. strain and preparation method for pyrroloquinoline quinone

A technology of pyrroloquinoline quinone and filamentous bacteria, which is applied in the field of microbial engineering, can solve the problems of low fermentation titer, cannot meet the needs of industrialized production, etc., and achieves the effects of stable genetic traits, excellent commercial application value, and easy realization.

Active Publication Date: 2017-01-04
ZHEJIANG HISUN PHARMA CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0015] The technical problem to be solved by the present invention is to provide a kind of hyphaetic microbe 1112-NTG-2318 and its fermentation to obtain pyrroloquine in view of the defects that the fermentation titer of pyrroloquinoline quinone is generally low and cannot meet the needs of industrial production. The method for quinone quinone, the described hyphaetic microbe 1112-NTG-2318 has a strong ability to produce pyrroloquinoline quinone under the supporting fermentation process, the strain performance is stable, and has good commercial value; the described method can be easily Obtain a large amount of pyrroloquinoline quinone, which is beneficial to industrial production

Method used

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  • Hyphomicrobium sp. strain and preparation method for pyrroloquinoline quinone
  • Hyphomicrobium sp. strain and preparation method for pyrroloquinoline quinone
  • Hyphomicrobium sp. strain and preparation method for pyrroloquinoline quinone

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Experimental program
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Embodiment 1

[0044] Embodiment 1, the preparation of high-yield bacterial strain Hyphomicrobium CGMCC No.10709 (Hyphomicrobium sp.)

[0045] Get test bacterial classification 1112-NTG-1953, this bacterial classification obtains TK0415 from the soil sample of Holland (Netherlands) T (No. 1112-O-36) obtained through natural screening and UV mutagenesis screening. (Strain TK 0415 T See Teizi Urakami, Junko Sasaki, Ken-Ichiro Suzuki, etc., Characterization and Description of Hyphomicrobium denitrificans sp.nov., International Journal Of Systematic Bacteriology, Vol.45, No.3, p.528-532, July 1995)

[0046] Draw the suspension of glycerol tube bacteria and spread it on the slant of the test tube. The slant medium is (g / L): ammonium sulfate 1.00, magnesium sulfate heptahydrate 0.20, sodium dihydrogen phosphate 0.50, dipotassium hydrogen phosphate 1.55, methanol 1.20, agar 15.00 , pH7.2, cultured at 26-30°C for 4 days. Scrape the slanted bacteria to prepare the test strain suspension for mutage...

Embodiment 2

[0053] Embodiment 2: Fermentation small-scale test process research of hyphaemicrobial bacteria CGMCC No.10709 on 50L tank

[0054] Based on the process parameters obtained from the 250ml shake flask and referring to the characteristics of the mutant strain CGMCC No.10709, a 50L tank fermentation process was designed and optimized. The growth and metabolism of the mutant strains and the optimal production capacity of pyrroloquinoline quinone were investigated.

[0055] Seed medium (g / L), ammonium sulfate 2.0, potassium dihydrogen phosphate 1.6, disodium hydrogen phosphate 2.8, magnesium sulfate heptahydrate 0.3, methanol 1.2, pH 7.0.

[0056] Seed tank process control: seed medium loading volume is 10 / 15L, culture temperature is 26-30°C, stirring speed is 200-500rpm, dissolved oxygen is greater than 15%, air flow rate is 0.5-2.0, and the culture period is 3 days.

[0057] Fermentation medium (g / L), namely methanol 12.0, ammonium sulfate 5.0, yeast extract powder 5.0, potassiu...

Embodiment 3

[0061] Embodiment 3: Fermentation small-scale test process research of hyphaemicrobial bacteria CGMCC No.10709 on 50L tank

[0062] Based on the process parameters of Example 2, a 50L tank fermentation process was designed and optimized. The growth and metabolism of the mutant strains and the optimal production capacity of pyrroloquinoline quinone were investigated.

[0063] Seed medium (g / L), ammonium sulfate 2.0, potassium dihydrogen phosphate 1.6, disodium hydrogen phosphate 2.8, magnesium sulfate heptahydrate 0.3, methanol 1.2, pH 7.0.

[0064] Seed tank process control: seed medium loading volume is 10 / 15L, culture temperature is 26-30°C, stirring speed is 200-500rpm, dissolved oxygen is greater than 15%, air flow rate is 0.5-2.0, and the culture period is 3 days.

[0065] Fermentation medium (g / L), namely methanol 12.0, ammonium sulfate 5.0, yeast extract powder 5.0, potassium dihydrogen phosphate 2.0, disodium hydrogen phosphate 5.0, magnesium sulfate heptahydrate 0.6,...

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Abstract

The invention discloses a Hyphomicrobium sp. strain and a preparation method for pyrroloquinoline quinone. The Hyphomicrobium sp. strain 1112-NTG-2318 capable of realizing high yield of pyrroloquinoline quinine is preserved in China General Microbiological Culture Collection Center with an accession number of CGMCC No. 10709. The preparation method for pyrroloquinoline quinine comprises the following steps: fermenting the Hyphomicrobium sp. strain CGMCC No. 10709 in a fermentation medium and acquiring pyrroloquinoline quinine from fermentation broth. The fermentation titer of the preparation method in a tank with a volume of 80 m<3> reaches 1783 mg / L, so the preparation method is favorable for industrial production of pyrroloquinoline quinine.

Description

technical field [0001] The invention belongs to the technical field of microbial engineering, and in particular relates to a hyphaetic microorganism and a method for fermenting and producing pyrroloquinoline quinone. Background technique [0002] Pyrroloquinoline quinone (Pyrroloquinoline quinone), referred to as PQQ, is the 14th B vitamin. [0003] The chemical structure of pyrroloquinoline quinone is as follows: [0004] [0005] Japanese scholars confirmed in 2003 that pyrroloquinoline quinone is a new type of water-soluble vitamin that fully meets the characteristics of vitamins. The research results were published in the international authoritative journal "Nature", which is the latest vitamin discovered for the first time in 55 years. Pyrroloquinoline quinone is a new prosthetic group, the third found in membrane-bound bacterial dehydrogenases after flavin nucleotides (FMN, FAD) and nicotinamide nucleotides (MAD, NADP) prosthetic group. [0006] Pyrroloquinoline ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P17/18C12R1/01
Inventor 郑玲辉杜敏娜朱小容周稳程涛张斌白骅
Owner ZHEJIANG HISUN PHARMA CO LTD
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