CN101955256B - Method for preparing ibuprofen-degrading bacterial agent - Google Patents

Method for preparing ibuprofen-degrading bacterial agent Download PDF

Info

Publication number
CN101955256B
CN101955256B CN2010102862885A CN201010286288A CN101955256B CN 101955256 B CN101955256 B CN 101955256B CN 2010102862885 A CN2010102862885 A CN 2010102862885A CN 201010286288 A CN201010286288 A CN 201010286288A CN 101955256 B CN101955256 B CN 101955256B
Authority
CN
China
Prior art keywords
ibuprofen
bacterial agent
degrading bacterial
preparation
degrading
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Expired - Fee Related
Application number
CN2010102862885A
Other languages
Chinese (zh)
Other versions
CN101955256A (en
Inventor
杨兴
薛罡
赵晓祥
刘亚男
史锦
程起跃
周宁娟
孙超
田世烜
罗来盛
阮俊杰
高品
张丹丹
毛菲菲
梅述芳
邢聆君
王贝灵
徐冰洁
何梦琦
刘畅
李姗珊
侍宽
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Donghua University
Original Assignee
Donghua University
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Donghua University filed Critical Donghua University
Priority to CN2010102862885A priority Critical patent/CN101955256B/en
Publication of CN101955256A publication Critical patent/CN101955256A/en
Application granted granted Critical
Publication of CN101955256B publication Critical patent/CN101955256B/en
Expired - Fee Related legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Images

Classifications

    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02WCLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
    • Y02W10/00Technologies for wastewater treatment
    • Y02W10/10Biological treatment of water, waste water, or sewage

Landscapes

  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

The invention relates to a method for preparing an ibuprofen-degrading bacterial agent, which comprises the following steps of: (1) inoculating activated sludge in a secondary sedimentation tank of a sewage treatment plant into a culture medium, domesticating by using a gradient pressure domestication method, standing, taking an upper-layer water sample, gradually diluting, streaking for separation, culturing, purifying, inoculating the separated strain into a beef extract peptone culture medium, and performing shake culture until a logarithmic phase; and (2) inoculating the cultured strain into the culture medium to prepare the ibuprofen-degrading bacterial agent. The method has the advantages of simpleness and low cost; and the prepared ibuprofen-degrading bacterial agent has the advantages of convenient use, high efficiency of degrading ibuprofen, easy inactivation of the strain, and application to ibuprofen removal in water treatment.

Description

A kind of preparation method of ibuprofen-degrading bacterial agent
Technical field
The invention belongs to the preparation field of ibuprofen-degrading bacterial agent, particularly relate to a kind of preparation method of ibuprofen-degrading bacterial agent.
Background technology
Medicine and personal care articles (Pharmaceutical and Personal Care Products is called for short PPCPs) receive people's concern day by day as a kind of emerging pollutent.Even the Pharmaceuticals and personal care products trace exists, still have the biological activity of certain degree, cause uncertain impact to environment, and may produce potential hazard to HUMAN HEALTH by approach such as tap water and foods.
In common PPCPs compound, microbiotic, antiphlogistic drug, antiepileptic drug, adjusting blood fat medicine in all kinds of surrounding mediums to detect frequency higher.Wherein non-steroidal antipyretic-antalgic and anti-inflammatory drug Ibuprofen BP/EP (Ibuprofen, molecular structure such as Fig. 1) as nonprescription drugs, are used in a large number, are one of PPCPs pollutants at present of greatest concern.
Traditional activated sludge process, different because of the region, the degradation efficiency of Ibuprofen BP/EP is differed.The product of photo-Fenton method degraded Ibuprofen BP/EP, toxicity strengthens on the contrary.Studies show that clearance and the microbial strains of Ibuprofen BP/EP are closely related.
Summary of the invention
Technical problem to be solved by this invention provides a kind of preparation method of ibuprofen-degrading bacterial agent, and the method is simple, and cost is low; The gained ibuprofen-degrading bacterial agent is easy to use, the efficient degradation Ibuprofen BP/EP, and bacterial strain is easy to deactivation.
A kind of efficient degradation Ibuprofen BP/EP bacterial strain for strengthening the ability of removing the sewage Ibuprofen BP/EP among the present invention is a kind of gramstaining reaction negative bacterial strain BL-1, through being accredited as serratia marcescens (Serratia marcescens).Main Biological is G -, thalline is rod-short, without the brood cell, and about 1.54 * 1.08 μ m of size, the tool peritrichous, facultative aerobic; V-P reaction, catalase, nitrate reaction, the gelatin liquification test positive, methyl red test, oxydase, phenylalanine reaction, hydrogen sulfide production test feminine gender; Can utilize sucrose, glucose, wood sugar, N.F,USP MANNITOL, pectinose to produce acid, can not utilize lactose to produce acid; This bacterial strain all loses activity behind uviolizing 2h.The Genbank accession number of this bacterial strain 16S rRNA is HM136577.
The preparation method of a kind of ibuprofen-degrading bacterial agent of the present invention comprises:
(1) get in the inoculum size access substratum of active sludge by 5%-10% of sewage work's second pond, with the domestication of gradient pressure type method for domesticating, ibuprofen concentration is 10-35mg/L; After the domestication, leave standstill, get the upper strata water sample, line separates behind the gradient dilution, then cultivates 48h-60h at 30 ℃-35 ℃, purifying, and in beef-protein medium, shaking culture is to logarithmic phase with isolated inoculation;
(2) above-mentioned cultured bacterial classification is pressed the inoculum size access culture medium culturing of 10%-20%, and be get final product;
The air flow of sterile air turns to 1 in the described culturing process: 0.6-1.2, and stirring velocity is 180-240 rev/min, culture temperature is 30-35 ℃; Incubation time is 48-60 hour, after the fermentation ends thalline quantity reach 1,000,000,000/more than the mL, nutrient solution directly was distributed into liquid dosage form or adopts adsorption by peat to be distributed into the solid fungicide formulation with packing bag with plastic barrel or packing bottle after fermentation was finished.
Culture medium prescription in the described step (1) is: K 2HPO 44.35g, KH 2PO 41.7g, NH 4Cl 2.1g, MgSO 40.2g, MnSO 40.05g, FeSO 47H 2O 0.01g, CaCl 22H 2O 0.03g, distilled water 1000mL, pH=6.8-7.0.
Domestication condition in the described step (1) is stirring velocity 160-240 rev/min, temperature 30-35 ℃, and time 1-2 month.
Beef-protein medium in the described step (1) is: extractum carnis 3.0g, peptone 10.0g, NaCl 5.0g, distilled water 1000mL, pH=7.0~7.2.
The concentration of the Ibuprofen BP/EP in the described step (1) is 10mg/L, 15mg/L, 20mg/L, 25mg/L, 30mg/L, 35mg/L.
Culture medium prescription in the described step (2) is: sucrose 10g/L, extractum carnis 3g/L, KH 2PO 41.7g/L, NH 4Cl2.1g/L, MgSO 40.2g/L, CaCO 30.3g/L, pH=7.0-7.5, Ibuprofen BP/EP 10-35mg/L.
Obtained strains is applied to the removal of the Ibuprofen BP/EP in the water treatment.
The present invention behind domestication, concentration and separation, purifying, obtains a strain has the efficient degradation effect to Ibuprofen BP/EP bacterium by the thought of biological reinforcing technology from the active sludge of municipal sewage plant's second pond.This bacterium source is in Waste Water Treatment, so this bacterial strain can adapt to complicated physical condition, and this is so that the having a extensive future of this bacterium.
Beneficial effect
(1) preparation method of the present invention is simple, and cost is low; The gained ibuprofen-degrading bacterial agent is easy to use, the efficient degradation Ibuprofen BP/EP, and bacterial strain is easy to deactivation, guaranteed the ecological security that uses; The present invention is for improving and preserving the ecological environment, and the protection people's is healthy significant;
(2) small-scale sewage treatment facility (BAF) result shows, can obviously improve the removal efficient of Ibuprofen BP/EP with the microbial inoculum of the present invention's production.
Description of drawings
The chemical structure of Fig. 1 Ibuprofen BP/EP;
Fig. 2 thalline electromicroscopic photograph;
Fig. 3 do not degrade (a) and degrade GC-MS total ion chromatogram of (b) sample;
Fig. 4 aeration biological filter device figure.
Embodiment
Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiment only to be used for explanation the present invention and be not used in and limit the scope of the invention.Should be understood that in addition those skilled in the art can make various changes or modifications the present invention after the content of having read the present invention's instruction, these equivalent form of values fall within the application's appended claims limited range equally.
Embodiment 1
1. the separation of bacterial strain and evaluation
(1) active sludge of taking from municipal sewage plant's second pond is inoculated in the minimal medium by 5% access amount, tame with gradient pressure type method for domesticating, ibuprofen concentration is 10,15,20,25,30,35mg/L, at 30 ℃, 160r/min, tame cultivation under the condition of loadings 100mL in the 250mL Erlenmeyer flask, the always domestication time is 2 months.The minimal medium prescription is K 2HPO 44.35g, KH 2PO 41.7g, NH 4Cl 2.1g, MgSO 40.2g, MnSO 40.05g, FeSO 47H 2O0.01g, CaCl 22H 2O 0.03g, distilled water 1000mL, pH=6.8-7.0.
(2) get the sample of domestication after 2 months, left standstill 30 minutes, get upper strata water sample 1mL, separate in the flat board line behind the gradient dilution.Then in 30 ℃ of incubators, cultivate 48h.The picking form is single bacterium colony clearly, and numbering, and purifying repeatedly to slant medium, are stored in isolated inoculation under 4 ℃ and treat the follow-up Degrading experiment that carries out in the refrigerator.Wherein enrichment medium is extractum carnis 3.0g, peptone 10.0g, NaCl 5.0g, distilled water 1000mL, pH=7.0~7.2.The agar powder that adds 17g/L in the solid medium.
(3) a certain amount of single bacterium colony after separation and purification of difference picking is in minimal medium separately, the Ibuprofen BP/EP that all contains 15mg/L in each minimal medium, at 30 ℃, 160r/min, to cultivate under the condition of loadings 100mL in the 250mL Erlenmeyer flask, incubation time is 48h.Measure the residual concentration of Ibuprofen BP/EP in the solution after cultivating, thereby determine the bacterial strain BL-1 that Ibuprofen BP/EP is had the efficient degradation effect;
Described inorganic medium is sucrose 10g/L, extractum carnis 3g/L, KH 2PO 41.7g/L, NH 4Cl 2.1g/L, MgSO 40.2g/L, CaCO 30.3g/L, pH=7.0-7.5, Ibuprofen BP/EP 15mg/L.
(4) identify and 16S rRNA sequential analysis and homology comparison that through Physiology and biochemistry bacterial strain BL-1 is initially identified as serratia marcescens (Serratia marcescens).Main Biological is G, and thalline is rod-short, without the brood cell, and about 1.54 * 1.08 μ m of size, the tool peritrichous, facultative aerobic; V-P reaction, catalase, nitrate reaction, the gelatin liquification test positive, methyl red test, oxydase, phenylalanine reaction, hydrogen sulfide production test feminine gender; Can utilize sucrose, glucose, wood sugar, N.F,USP MANNITOL, pectinose to produce acid, can not utilize lactose to produce acid; This bacterial strain all loses activity behind uviolizing 2h.The Genbank accession number of this bacterial strain 16S rRNA is HM136577.
2. laboratory Ibuprofen BP/EP Degrading experiment
A strain Ibuprofen BP/EP efficient degrading bacteria provided by the present invention is about 7. in initial pH value, temperature is 30 ℃, oscillation rate is 160r/min, the liquid amount of 250mL Erlenmeyer flask is under the condition of 100mL, the inoculation amount is 0.6%, the Ibuprofen BP/EP starting point concentration is 15mg/L, incubation time 48h, and the degradation rate of Ibuprofen BP/EP is up to 90%.
3. the strenuous test of small-scale sewage treatment facility (BAF)
The bacterium liquid of fermentation by 2% ratio, is being intake 15.6 ℃~24.5 ℃ of water temperatures, and under the condition of pH 6.88~7.27, influent load is 0.2m 3/ m 2H, gas-water ratio is 1.5: 1 o'clock, the clearance that has added Ibuprofen BP/EP in the BAF of microbial inoculum is 86%~91%.And the clearance that does not add Ibuprofen BP/EP in the BAF of microbial inoculum is 48%~53%.

Claims (6)

1. the preparation method of an ibuprofen-degrading bacterial agent comprises:
(1) get in the inoculum size access substratum of active sludge by 5%-10% of sewage work's second pond, with the domestication of gradient pressure type method for domesticating, ibuprofen concentration is 10-35mg/L; After the domestication, leave standstill, get the upper strata water sample, line separates behind the gradient dilution, then cultivates 48h-60h at 30 ℃-35 ℃, purifying, and in beef-protein medium, shaking culture is to logarithmic phase with isolated inoculation;
(2) above-mentioned cultured bacterial classification is pressed the inoculum size access culture medium culturing of 10%-20%, and be get final product;
The air flow of sterile air turns to 1 in the described culturing process: 0.6-1.2, and stirring velocity is 180-240 rev/min, culture temperature is 30-35 ℃; Incubation time is 48-60 hour, after the fermentation ends thalline quantity reach 1,000,000,000/more than the mL.
2. the preparation method of a kind of ibuprofen-degrading bacterial agent according to claim 1, it is characterized in that: the culture medium prescription in the described step (1) is: K 2HPO 44.35g, KH 2PO 41.7g, NH 4Cl 2.1g, MgSO 40.2g, MnSO 40.05g, FeSO 47H 2O 0.01g, CaCl 22H 2O 0.03g, distilled water 1000mL, pH=6.8-7.0.
3. the preparation method of a kind of ibuprofen-degrading bacterial agent according to claim 1, it is characterized in that: the domestication condition in the described step (1) is stirring velocity 160-240 rev/min, temperature 30-35 ℃, time 1-2 month.
4. the preparation method of a kind of ibuprofen-degrading bacterial agent according to claim 1, it is characterized in that: the beef-protein medium in the described step (1) is: extractum carnis 3.0g, peptone 10.0g, NaCl 5.0g, distilled water 1000mL, pH=7.0~7.2.
5. the preparation method of a kind of ibuprofen-degrading bacterial agent according to claim 1, it is characterized in that: the concentration of the Ibuprofen BP/EP in the described step (1) is 10mg/L, 15mg/L, 20mg/L, 25mg/L, 30mg/L, 35mg/L.
6. the preparation method of a kind of ibuprofen-degrading bacterial agent according to claim 1, it is characterized in that: the culture medium prescription in the described step (2) is: sucrose 10g/L, extractum carnis 3g/L, KH 2PO 41.7g/L, NH 4Cl 2.1g/L, MgSO 40.2g/L, CaCO 30.3g/L, pH=7.0-7.5, Ibuprofen BP/EP 10-35mg/L.
CN2010102862885A 2010-09-19 2010-09-19 Method for preparing ibuprofen-degrading bacterial agent Expired - Fee Related CN101955256B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN2010102862885A CN101955256B (en) 2010-09-19 2010-09-19 Method for preparing ibuprofen-degrading bacterial agent

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN2010102862885A CN101955256B (en) 2010-09-19 2010-09-19 Method for preparing ibuprofen-degrading bacterial agent

Publications (2)

Publication Number Publication Date
CN101955256A CN101955256A (en) 2011-01-26
CN101955256B true CN101955256B (en) 2013-01-16

Family

ID=43482885

Family Applications (1)

Application Number Title Priority Date Filing Date
CN2010102862885A Expired - Fee Related CN101955256B (en) 2010-09-19 2010-09-19 Method for preparing ibuprofen-degrading bacterial agent

Country Status (1)

Country Link
CN (1) CN101955256B (en)

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103255161B (en) * 2013-05-15 2015-07-29 西北农林科技大学 A kind of method knocking out Ibuprofen BP/EP FOX plasmid catechol dioxygenase gene
CN109574399B (en) * 2018-12-27 2021-07-30 南京大学 Method for removing diclofenac in sewage in enhanced manner based on enrichment of nitrobacteria

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0567102A2 (en) * 1992-04-22 1993-10-27 Canon Kabushiki Kaisha Method of biologically decomposing phenol or furan compounds by microorganisms
CN1837356A (en) * 2006-04-21 2006-09-27 哈尔滨工业大学 Method for acclimating, separating and screening highly effective phosphate-accumulating bacteria from intermittent circulating active sludge-membrane biological reaction system
CN101031637A (en) * 2005-04-21 2007-09-05 揖斐电株式会社 Method of treating wastewater containing organic compound
CN101260373A (en) * 2007-10-24 2008-09-10 大连理工大学 Micrococcus luteus with novel metabolic characteristic and application thereof in biological degradation

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0567102A2 (en) * 1992-04-22 1993-10-27 Canon Kabushiki Kaisha Method of biologically decomposing phenol or furan compounds by microorganisms
CN101031637A (en) * 2005-04-21 2007-09-05 揖斐电株式会社 Method of treating wastewater containing organic compound
CN1837356A (en) * 2006-04-21 2006-09-27 哈尔滨工业大学 Method for acclimating, separating and screening highly effective phosphate-accumulating bacteria from intermittent circulating active sludge-membrane biological reaction system
CN101260373A (en) * 2007-10-24 2008-09-10 大连理工大学 Micrococcus luteus with novel metabolic characteristic and application thereof in biological degradation

Also Published As

Publication number Publication date
CN101955256A (en) 2011-01-26

Similar Documents

Publication Publication Date Title
CN104673715B (en) There is fixed effect to cadmium and enterobacteria and its application of plant growth can be promoted
CN101857847A (en) Pseudomonas aeruginosa strain separating, purifying and domesticating method and use
CN104195090A (en) Compound microbiological preparation and application thereof in waste water treatment
CN102586160B (en) Stenotrophomonas maltophilia DS4
CN108264150A (en) A kind of complex micro organism fungicide applies the method in water body is administered
CN106754578B (en) Microbial inoculum and the application of one plant of chloramphenicol degradation bacteria strains LMS-CY and its production
CN101948782B (en) Rhodopseudomonas spheroides strain and liquid inoculant thereof as well as preparation method and application thereof
CN109706096B (en) Cold-resistant brevibacterium strain with denitrification and efficient flocculation capacity and application thereof
CN107201322A (en) Bacillus subtilis and its application for degrading aflatoxin B 1
CN101519639A (en) Paenibacillus polymyxa for preventing and treating plant fungal diseases and production method thereof
Liu et al. Enhancement of sludge dewaterability with filamentous fungi Talaromyces flavus S1 by depletion of extracellular polymeric substances or mycelium entrapment
CN102583780A (en) Application of Stenotrophomonas maltophilia DS4 for degrading organic pollutants in saponin waste water
CN102206585A (en) Composite microbial preparation for purifying water and preparation method thereof
CN101955256B (en) Method for preparing ibuprofen-degrading bacterial agent
CN103305443A (en) Preparation and method for restoring ammonia nitrogen-containing industrial sewage
CN109576195A (en) One pseudomonas and its application on degradation terramycin
CN110157630A (en) A kind of composite bacteria agent and preparation method thereof applied to high salt organic waste water
CN102515366B (en) Microbiological degradation method for nitrobenzene industrial wastewater
CN104862253B (en) One plant of Norfloxacin degraded acinetobacter calcoaceticus NOR 36 and its application
CN103224899A (en) Preparation method of antibiotic medicine erythromycin high-efficiency degradation microbial inoculum
CN103898002A (en) Method for producing compound bacteria for water environment improvement
CN102757926B (en) Bacillus thuringiensis emulsification fermentation culture medium
Niu et al. Cold-shock resistance of activated sludge microorganisms strengthened by a static magnetic field
CN108102942A (en) One plant of bacterial strain and its application for being used to purify molasses alcohol waste water
CN101691558A (en) Method for extracting efficient degrading bacteria of carbamazepine

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
C53 Correction of patent for invention or patent application
CB03 Change of inventor or designer information

Inventor after: Yang Xing

Inventor after: Luo Laisheng

Inventor after: Ruan Junjie

Inventor after: Gao Pin

Inventor after: Zhang Dandan

Inventor after: Mao Feifei

Inventor after: Mei Shufang

Inventor after: Xing Lingjun

Inventor after: Wang Beiling

Inventor after: Xu Bingjie

Inventor after: He Mengqi

Inventor after: Xue Gang

Inventor after: Liu Chang

Inventor after: Li Panshan

Inventor after: Shi Kuan

Inventor after: Zhao Xiaoxiang

Inventor after: Liu Yanan

Inventor after: Shi Jin

Inventor after: Cheng Qiyue

Inventor after: Zhou Ningjuan

Inventor after: Sun Chao

Inventor after: Tian Shihui

Inventor before: Yang Xing

Inventor before: Luo Laisheng

Inventor before: Ruan Junjie

Inventor before: Xue Gang

Inventor before: Zhao Xiaoxiang

Inventor before: Liu Yanan

Inventor before: Shi Jin

Inventor before: Cheng Qiyue

Inventor before: Zhou Ningjuan

Inventor before: Sun Chao

Inventor before: Tian Shihui

COR Change of bibliographic data

Free format text: CORRECT: INVENTOR; FROM: YANG XING XUE GANG ZHAO XIAOXIANG LIU YANAN SHI JIN CHENG QIYUE ZHOU NINGJUAN SUN CHAO TIAN SHI LUO LAISHENG RUAN JUNJIE TO: YANG XING XUE GANG ZHAO XIAOXIANG LIU YANAN SHI JIN CHENG QIYUE ZHOU NINGJUAN SUN CHAO TIAN SHI LUO LAISHENG RUAN JUNJIE GAO PIN ZHANG DANDAN MAO FEIFEI MEI SHUFANG XING LINGJUN WANG BEILING XU BINGJIE HE MENGQI LIU CHANG LI SHANSHAN SHI KUAN

C14 Grant of patent or utility model
GR01 Patent grant
CF01 Termination of patent right due to non-payment of annual fee

Granted publication date: 20130116

Termination date: 20150919

EXPY Termination of patent right or utility model