Biological preparation method of fermentation medium and rhamnolipid

A fermentation medium and biological preparation technology, applied in the field of efficient and harmless rhamnolipid biological preparation, can solve problems such as decreased expression of pathogenic factors, and achieve the effects of promoted secretion, increased production, and low use cost

Active Publication Date: 2017-04-26
ZHEJIANG GONGSHANG UNIVERSITY
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AI Technical Summary

Problems solved by technology

Most clinically isolated Pseudomonas aeruginosa quorum-sensing lasR deletion mutants have significantly decreased expression of virulence factors

Method used

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  • Biological preparation method of fermentation medium and rhamnolipid
  • Biological preparation method of fermentation medium and rhamnolipid
  • Biological preparation method of fermentation medium and rhamnolipid

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] (1) Cultivate the seed solution for rhamnolipid synthesis for 10h-12h until it is in the logarithmic growth phase or the early stage of the stationary phase. The rhamnolipid producing bacteria used are: 1. Pseudomonas aeruginosa wild strain, 2. Pseudomonas aeruginosa mutant strain lacking lasR gene, 3. Burkholderia cepacia strain.

[0047] (2) prepare the iron-limited medium that promotes rhamnolipid production: c((NH 4 ) 2 SO 4 )=1g / L, c(KH 2 PO 4 )=1.7g / L, c(NaHPO 4 )=1.775g / L, c(EDTA)=0.0025g / L, c(ZnSO 4 ·7H 2 O)=0.011g / L, c(MnSO 4 ·H 2 O)=0.00154g / L, c(CuSO 4 ·5H 2 O)=0.000392g / L, c(Co(NO 3 ) 2 ·6H 2 O)=0.00025g / L, c(Na 2 B 4 o 7 10H 2 O)=0.000177g / L, c(CaCl 2 2H 2 O)=0.0667g / L, c(MgSO 4 )=0.289g / L, c((NH 4 ) 6 Mo 7 o 24 4H 2 O)=0.000185g / L, c(KOH)=0.146g / L, c(nitrilotriacetic acid)=0.2g / L, c(FeSO 4 ·7H 2 O)=0.000698g / L, c (glutamic acid)=10g / L.

[0048] (3) Insert (1) seed solution into (2) culture medium for fermentation and cultivation...

Embodiment 2

[0058] (1) Cultivate the seed solution for rhamnolipid synthesis. The used rhamnolipid producing bacteria is a mutant strain of Pseudomonas aeruginosa lacking the lasR gene. The seed solution was cultured to 1.2h in the early stage of logarithmic growth; 2.12h in the early stage of stationary stage; 3.20h in the late stage of stationary stage.

[0059] (2) prepare the iron-limited medium that promotes rhamnolipid production: c((NH 4 ) 2 SO 4 )=1g / L, c(KH 2 PO 4 )=1.7g / L, c(NaHPO 4 )=1.775g / L, c(EDTA)=0.0025g / L, c(ZnSO 4 ·7H 2 O)=0.011g / L, c(MnSO 4 ·H 2 O)=0.00154g / L, c(CuSO 4 ·5H 2 O)=0.000392g / L, c(Co(NO 3 ) 2 ·6H 2 O)=0.00025g / L, c(Na 2 B 4 o 7 10H 2 O)=0.000177g / L, c(CaCl 2 2H 2 O)=0.0667g / L, c(MgSO 4 )=0.289g / L, c((NH 4 ) 6 Mo 7 o 24 4H 2 O)=0.000185g / L, c(KOH)=0.146g / L, c(nitrilotriacetic acid)=0.2g / L, c(FeSO 4 ·7H 2 O)=0.000698g / L, c (glutamic acid)=10g / L.

[0060] (3) The seed solution of (1) is respectively inserted into the culture medium ...

Embodiment 3

[0069] (1) Cultivate the seed solution for rhamnolipid synthesis for 10h-12h until it is in the logarithmic growth phase or the early stage of the stationary phase. The used rhamnolipid producing bacteria is a mutant strain of Pseudomonas aeruginosa lacking the lasR gene.

[0070] (2) preparation promotes the substratum that rhamnolipid produces to be respectively: 1. ferric glutamate restriction substratum: c((NH4 ) 2 SO 4 )=1g / L, c(KH 2 PO 4 )=1.7g / L, c(NaHPO 4 )=1.775g / L, c(EDTA)=0.0025g / L, c(ZnSO 4 ·7H 2 O)=0.011g / L, c(MnSO 4 ·H 2 O)=0.00154g / L, c(CuSO 4 ·5H 2 O)=0.000392g / L, c(Co(NO 3 ) 2 ·6H 2 O)=0.00025g / L, c(Na 2 B 4 o 7 10H 2 O)=0.000177g / L, c(CaCl 2 2H 2 O)=0.0667g / L, c(MgSO 4 )=0.289g / L, c((NH 4 ) 6 Mo 7 o 24 4H 2 O)=0.000185g / L, c(KOH)=0.146g / L, c(nitrilotriacetic acid)=0.2g / L, c(FeSO 4 ·7H 2 (O)=0.000698g / L, c (glutamic acid)=10g / L; 2. casein iron-limited medium: c((NH 4 ) 2 SO 4 )=1g / L, c(KH 2 PO 4 )=1.7g / L, c(NaHPO 4 )=1.775g / L, c...

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Abstract

The invention discloses a biological preparation method of a fermentation medium and rhamnolipid. The biological preparation method comprises the following steps: (1) inoculating a rhamnolipid producing strain seed solution into an iron limited fermentation medium, carrying out fermentation culturing, and adding nanoparticles into the fermentation medium; and (2) extracting the rhamnolipid of obtained fermentation liquor by using an acid precipitation freeze-drying method. A mutant strain which is weak in pathogenic ability and does not have quorum sensing genes lasR is used, synthesis of the rhamnolipid is stimulated by the suitable nanoparticles under the suitable culture conditions, secretion of the rhamnolipid can be promoted, and moreover, health risks in a production process is greatly reduced. Meanwhile, the biological preparation method has the characteristics of low cost and simplicity in operation.

Description

technical field [0001] The invention relates to the field of biosynthesis and the field of application of nanometer materials, in particular to an efficient and harmless rhamnolipid biological preparation method. Background technique [0002] Surfactant can significantly reduce the surface tension of the target solution. It is an important chemical raw material and is known as "industrial monosodium glutamate". status. At present, almost all surfactants are chemically synthesized from petroleum, and chemically synthesized surfactants often cause serious environmental pollution problems during production and use. Biosurfactant is a rising star in the surfactant family, which is a kind of biomacromolecular substances with surface activity produced by microorganisms. Compared with chemically synthesized surfactants, biosurfactants not only have the same effects of reducing surface tension, stabilizing emulsions and increasing foam, but also have non-toxic and biodegradable pr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P19/44C07H15/04C07H1/06C12R1/385
CPCC07H1/06C07H15/04C12P19/44
Inventor 汪美贞汪丽佳严慧聪沈臻黄超民张芸芸陈健李娜申屠佳丽殷峻沈东升
Owner ZHEJIANG GONGSHANG UNIVERSITY
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