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Pseudomonas putida capable of metabolizing nicotine and application thereof

A technology of Pseudomonas putida and nicotine, which is applied in the direction of bacteria, microbial-based methods, biological water/sewage treatment, etc., can solve the problems of weak resistance to nicotine toxicity, limited application, and weak metabolism

Inactive Publication Date: 2008-11-19
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although these microorganisms can metabolize nicotine, their metabolic ability is weak, and their resistance to nicotine toxicity is weak, which limits their application in actual production, and there are no relevant reports on the treatment of nicotine in tobacco processing wastewater

Method used

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  • Pseudomonas putida capable of metabolizing nicotine and application thereof
  • Pseudomonas putida capable of metabolizing nicotine and application thereof
  • Pseudomonas putida capable of metabolizing nicotine and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] Example 1: Screening of Pseudomonas putida (Pseudomonas putida) XPSN CCTCC NO.M 205038 strain

[0074] Collect soil samples from long-term tobacco-growing fields in Weifang, Shandong, etc., take 2-4g and add them to the liquid basic medium containing 1g / L nicotine, shake and cultivate on a shaker at 30°C for 2-4 days, and then transfer to Then transfer 5mL to the same culture medium and transfer 3 to 5 times in succession according to this method. Dilute the obtained culture solution and spread it on the basic medium of solid plate containing 1g / L nicotine. Culture at 30℃ for 1 ~3 days, after the colonies grow, pick large colonies and streak them continuously on the same nicotine solid plate basic medium to confirm that the strains have the ability to degrade nicotine and are pure cultures, and then pick single colonies to contain 1g / L nicotine on a solid slant basic medium, cultured at 30°C for 1 to 3 days to obtain the Pseudomonas putida XPSN of the present invention,...

Embodiment 2

[0080] Example 2: Acquisition of the 16S rDNA of the above-mentioned Pseudomonas putida XPSN CCTCC NO.M 205038 strain

[0081] Collect 2 mL of the culture of Pseudomonas putida (Pseudomonas putida) XPSN CCTCC NO.M205038 that has grown to the stationary phase, centrifuge at 6,000 rpm for 5 minutes, and remove the supernatant; add 565 μL of TE solution to the precipitate, and the formula of the TE solution is : Tris (Tris) of 10mmol / L, ethylenediaminetetraacetic acid (EDTA) of 1mmol / L, adjust the pH to 8.0 with hydrochloric acid; blow repeatedly with a pipette to suspend the sediment, and then add 30 μL of mass-volume ratio Concentration of 10% sodium dodecylsulfonate (SDS) and 5 μL 20 mg / mL proteinase K (Merck, USA), mix well, and put in a water bath at 37 ° C for 1 hour; add 100 μL 5mol / L NaCl, fully Mix well, then add 80 μL of CTAB / NaCl solution, the formula of CTAB / NaCl solution is: the mass volume ratio concentration is 10% cetyltriethylammonium bromide (CTAB) is dissolved ...

Embodiment 3

[0108] Embodiment 3: Utilize the method for the intact cell of Pseudomonas putida (Pseudomonas putida) to process tobacco processing wastewater, wherein the final concentration of nicotine in the tobacco processing wastewater is 0.8g / L

[0109] (1) Microbial strain: select Pseudomonas putida XPSN CCTCC NO.M205038;

[0110] (2) Slant culture: inoculate the above-mentioned strains on a solid slant basic medium containing 0.02% nicotine in a mass volume ratio, and culture at 20°C for 24 hours;

[0111] (3) Seed culture: the bacterial strain cultivated in step (2) was placed in 20 mL of nicotine liquid basic medium containing 0.02% mass volume ratio with an inoculation loop under aseptic conditions, and was shaken at 20° C. Shake culture on the bed for 6 hours to obtain first-grade seeds;

[0112] (4) Expansion cultivation: with the inoculum amount of 5% by volume, connect the primary seeds in 200 mL of nicotine liquid basic medium containing 0.02% by mass volume ratio, and culture...

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Abstract

The invention discloses a pseudomonas putida to metabolite nicotine, which is characterized by the following: preserving in China Typical Culture Preservation Center (Wuhan university, china Wuhan) with keeping number as CCTCC NO.M 205038; possessing stronger nicotine metabolic ability and nicotine toxin immunity ability; growing highest at 6g / L nicotine density; culturing a good deal of bacterial strain; using intact cell as biological activator; degrading nicotine from tobacco industrial waste water completely. This bacterium possesses simple operation and quick disposing waste water speed, which can be used to dispose the waste water from tobacco industrial widely.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a strain of Pseudomonas putida capable of metabolizing nicotine and its application. Background technique [0002] Nicotine is the main alkaloid in many tobaccos, with a content of 0.5-12%. Because it mainly acts on the central nervous system, many people have a strong dependence on tobacco. Yet nicotine is a kind of " poisonous dangerous substance ", and when one-time inhalation amount reaches 40ml / person, can suppress central nervous system, cause heart paralysis, have fatal danger; And smoking moment will be nitrosated to generate tobacco-specific N-nitrosamines (tobacco-specific nitrosamines, TSNA), such as nitrosamine4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), they are Strong lung carcinogen. [0003] Because nicotine has good water solubility, a large amount of waste water is produced in tobacco processing. These waste water contain toxic substances ma...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C02F3/34C12R1/40
Inventor 王书宁许平
Owner SHANDONG UNIV
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