Nothing Special   »   [go: up one dir, main page]

CN111269848B - Rhodococcus ruber JJ-3 and application thereof in acrylic acid degradation - Google Patents

Rhodococcus ruber JJ-3 and application thereof in acrylic acid degradation Download PDF

Info

Publication number
CN111269848B
CN111269848B CN201911393310.3A CN201911393310A CN111269848B CN 111269848 B CN111269848 B CN 111269848B CN 201911393310 A CN201911393310 A CN 201911393310A CN 111269848 B CN111269848 B CN 111269848B
Authority
CN
China
Prior art keywords
acrylic acid
rhodococcus ruber
culture
culture medium
value
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.)
Active
Application number
CN201911393310.3A
Other languages
Chinese (zh)
Other versions
CN111269848A (en
Inventor
陈浚
戴璐瑶
胡利勇
何金佳
张士汉
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.)
Zhejiang University of Technology ZJUT
Original Assignee
Zhejiang University of Technology ZJUT
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 Zhejiang University of Technology ZJUT filed Critical Zhejiang University of Technology ZJUT
Priority to CN201911393310.3A priority Critical patent/CN111269848B/en
Publication of CN111269848A publication Critical patent/CN111269848A/en
Application granted granted Critical
Publication of CN111269848B publication Critical patent/CN111269848B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Images

Classifications

    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20Bacteria; Culture media therefor
    • C12N1/205Bacterial isolates
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12RINDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00Microorganisms ; Processes using microorganisms
    • C12R2001/01Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • CCHEMISTRY; METALLURGY
    • C02TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02FTREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F3/00Biological treatment of water, waste water, or sewage
    • C02F3/34Biological treatment of water, waste water, or sewage characterised by the microorganisms used
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20Bacteria; Culture media therefor
    • CCHEMISTRY; METALLURGY
    • C02TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02FTREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F2101/00Nature of the contaminant
    • C02F2101/30Organic compounds
    • C02F2101/34Organic compounds containing oxygen

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • Zoology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Genetics & Genomics (AREA)
  • Wood Science & Technology (AREA)
  • Microbiology (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Biochemistry (AREA)
  • Medicinal Chemistry (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Virology (AREA)
  • Biomedical Technology (AREA)
  • Biodiversity & Conservation Biology (AREA)
  • Hydrology & Water Resources (AREA)
  • Environmental & Geological Engineering (AREA)
  • Water Supply & Treatment (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Purification Treatments By Anaerobic Or Anaerobic And Aerobic Bacteria Or Animals (AREA)

Abstract

The invention disclosesA new strain-Rhodococcus ruber JJ-3 and its application for degrading acrylic acid are disclosed, the Rhodococcus ruber JJ-3 is preserved in China center for type culture Collection with the address: china, wuhan university, zip code: 430072, deposit number: CCTCC NO: m2019546, date of deposit 2019, month 07, day 12; the invention provides Rhodococcus erythropolis JJ-3 with acrylic acid degradation performance and application thereof in degrading acrylic acid wastewater, wherein the strain can react with initial concentration of 1000 mg.L within 16h‑1The acrylic acid degradation rate reaches 100%, and the discovery of the degrading bacteria has important significance for the efficient purification of acrylic acid in industrial wastewater.

Description

Rhodococcus ruber JJ-3 and application thereof in acrylic acid degradation
(I) technical field
The invention belongs to the technical field of biological treatment of environmental pollutants, and particularly relates to a new strain with acrylic acid degradation capacity, namely Rhodococcus ruber JJ-3(Rhodococcus ruber JJ-3), and application thereof.
(II) background of the invention
Acrylic acid is the simplest unsaturated carboxylic acid, consisting of one vinyl group and one carboxyl group, and is a vinyl monomer that polymerizes very quickly. Acrylic acid is a colorless liquid with a characteristic pungent odor, is miscible with water, alcohols, ethers, and chloroform, and is prepared from propylene. Acrylic acid is an important organic synthetic raw material and synthetic resin monomer, and is mostly used for producing acrylic esters such as methyl acrylate, ethyl acrylate, butyl acrylate, hydroxyethyl acrylate and the like. The acrylic acid and acrylic ester can be homopolymerized or copolymerized, and the polymer can be used in the production processes of synthetic resin, synthetic fiber, super absorbent resin, building material, paint and the like. Acrylic acid is one of the 3-class carcinogen lists published by international cancer research institution of world health organization in 2017.
Acrylic acid is an important chemical raw material and is widely applied to industries such as paint, coating, leather, textile, printing and the like. At present, the total yield of acrylic acid in China is about 120 million t, and the total amount of acrylic acid wastewater in China is about 140 million t every year according to 1.2t wastewater generated by producing 1t acrylic acid.
The catalytic wet oxidation method for treating the acrylic acid wastewater has the advantages of high removal efficiency, low energy consumption and small secondary pollution, is a cleaner treatment technology, but has the problems of high operation cost, high control difficulty, secondary pollution of heavy metal catalysts and the like; the supercritical water oxidation method for treating the acrylic acid wastewater has the advantages of short reaction time, thorough pollutant removal, small occupied area, cleanness, environmental protection and the like, but high-concentration dissolved oxygen, equipment corrosion and inorganic salt deposition caused by high temperature and high pressure are the main problems faced by the technology; the fiber adsorption method, the ion exchange fiber method, the Fen-ton oxidation method and the photoelectron decomposition method for degrading the acrylic acid wastewater have the characteristics of mild reaction conditions, high reaction speed and high removal efficiency, but cannot be applied to the field of actual engineering treatment because of less deep research and high treatment cost at present. The method for treating the acrylic acid wastewater by using the biological method has the advantages of low energy consumption, mild reaction conditions and higher efficiency, and is more in line with the national strategy of energy conservation, emission reduction and sustainable development, but the research on the acrylic acid degradation functional strains at home and abroad is only rarely reported, so the research has important significance for screening the available strains for acrylic acid degradation and performing acrylic acid biodegradation.
Disclosure of the invention
The invention aims to provide a Rhodococcus ruber with acrylic acid degradation performance and application thereof, and provides a new method for treating acrylic acid wastewater.
The technical scheme adopted by the invention is as follows:
the invention provides a new strain-Rhodococcus ruber (Rhodococcus ruber) JJ-3, which is preserved in China center for type culture Collection with the preservation number: CCTCC NO: m2019546, date of deposit 2019, month 07, day 12, address: china, wuhan university, zip code: 430072.
the Rhodococcus ruber JJ-3 strain is characterized in that: the colony is orange yellow, the round opaque surface is rough and dry, the cell shape is spherical, and the cell has no flagellum and regular edges. The optimal growth pH value is 7.0, the optimal growth temperature is 35 ℃, and the 16S rDNA sequence of the strain is shown in SEQ ID No. 1.
The invention also provides application of the Rhodococcus ruber JJ-3 in biodegradation of acrylic acid. Acrylic acid is the simplest unsaturated carboxylic acid, consisting of one vinyl group and one carboxyl group, and is pure, colorless and clear with a characteristic pungent odor.
Further, the application is as follows: inoculating a fermentation liquid, namely a bacterium-containing suspension, obtained by carrying out fermentation culture on Rhodococcus ruber JJ-3 into an acrylic acid liquid selection culture medium, and culturing at the temperature of 20-45 ℃ and the speed of 140-160rpm by taking acrylic acid as a carbon source to realize the degradation of the acrylic acid; the final concentration composition of the acrylic acid liquid selective medium is as follows: NH (NH)4Cl 540mg·L-1,K2HPO41000mg·L-1,KH2PO41000mg·L-1,NaCl 500mg·L -1200 mg/L yeast extract powder-1Acrylic acid 100-2000 mg. L-1,MgSO4·7H2O 16mg·L-1,CaCl24mg·L-1,CuSO4 4mg·L-1,FeSO4 4mg·L-1,MnSO4 4mg·L-1The solvent is water, and the pH value is 5.0-10.0.
Further, the OD of the suspension containing the bacterial cells is preferred600The value is 0.7 to 1.0, preferably 0.8.
Further, the acrylic acid is added at a concentration of 0.1 to 2g/L, preferably 0.1 to 1g/L, and most preferably 1 g/L.
Further, it is preferable that the culture pH is 7.0, 35 ℃ and 160 rpm.
Further, the volume inoculation amount of the bacterial suspension is preferably 3-20%, and preferably 10-15%.
The bacteria-containing suspension is prepared by the following steps:
(1) slant culture: inoculating Rhodococcus ruber JJ-3 to a slant culture medium, and culturing at 30 deg.C for 3 days to obtain a thallus slant; the final concentration of the slant culture medium is as follows: NH (NH)4Cl 540mg·L-1,K2HPO4 1000mg·L-1,KH2PO41000mg·L-1,NaCl 500mg·L -1200 mg/L yeast extract powder-1Peptone 400 mg. L-1 Acrylic acid 1000 mg. L-1,MgSO4·7H2O 16mg·L-1,CaCl24mg·L-1,CuSO4 4mg·L-1,FeSO4 4mg·L-1,MnSO4 4mg·L-1The solvent is water, the pH value is 7.0, and the agar is 18-20 g.L-1(preferably 18 g. L)-1);
(2) Seed culture: selecting colonies from the thallus slant, inoculating the colonies to an LB culture medium, and culturing at 30 ℃ for 18h to obtain a seed solution; the LB culture medium has the following final concentration composition: NaCl 10 g.L-15 g.L of yeast extract powder-1Peptone 10 g. L-1The solvent is water, and the pH value is 7.0;
(3) fermentation culture: inoculating the seed solution to a fermentation culture medium by an inoculation amount with the volume concentration of 3-20%, and culturing at 30 ℃ to obtain a fermentation culture solution, namely a bacterium-containing suspension; the final concentration of the fermentation medium is as follows: NH (NH)4Cl 540mg·L-1,K2HPO41000mg·L-1,KH2PO41000mg·L-1,NaCl 500mg·L -1200 mg/L yeast extract powder-1 Acrylic acid 1000 mg. L-1,MgSO4·7H2O 16mg·L-1,CaCl24mg·L-1,CuSO4 4mg·L-1,FeSO4 4mg·L-1,MnSO4 4mg·L-1The solvent is water, and the pH value is 7.0.
Compared with the prior art, the invention has the following beneficial effects: the invention provides a Rhodococcus erythropolis JJ-3 with acrylic acid degradation performance and application thereof in acrylic acid degradation, wherein the strain can react to the initial concentration of 1000 mg.L in 5d-1The acrylic acid degradation rate reaches 100%, and the degrading bacteria have important significance for treating acrylic acid in industrial wastewater.
(IV) description of the drawings
FIG. 1 is a transmission electron micrograph of strain JJ-3.
FIG. 2 is a phylogenetic tree diagram of strain JJ-3.
FIG. 3 is a comparison of the acrylic acid degradation performance of strain JJ-3 at different pH.
FIG. 4 is a comparison of the acrylic acid degradation performance of strain JJ-3 at different temperatures.
FIG. 5 is a comparison of the acrylic acid-degrading performance of strain JJ-3 at different inoculum sizes.
FIG. 6 is a comparison of the degradation performance of strain JJ-3 at different initial acrylic acid concentrations.
(V) detailed description of the preferred embodiments
The invention will be further described with reference to specific examples, but the scope of the invention is not limited thereto:
example 1: isolation, purification and characterization of Rhodococcus ruber JJ-3
Isolation and purification of Rhodococcus ruber JJ-3
Rhodococcus ruber JJ-3 is screened from sludge in a wastewater treatment tank of Zhejiang satellite energy Co., Ltd, and the specific steps are as follows:
the acrylic acid liquid selective culture medium is prepared by the following method: NH (NH)4Cl 540mg·L-1,K2HPO4 1000mg·L-1,KH2PO4 1000mg·L-1,NaCl 500mg·L -1200 mg/L yeast extract powder-1Acrylic acid 1000 mg. L-1,MgSO4·7H2O 16mg·L-1,CaCl2 4mg·L-1,CuSO4 4mg·L-1,FeSO4 4mg·L-1,MnSO4 4mg·L-1The solvent is water, pH value is 7.0, and sterilization is carried out for 20min at 121 ℃.
Acrylic acid solid selection medium: NH (NH)4Cl 540mg·L-1,K2HPO41000mg·L-1,KH2PO41000mg·L-1,NaCl 500mg·L -1200 mg/L yeast extract powder-1Peptone 400 mg. L-1Acrylic acid 1000 mg. L-1,MgSO4·7H2O 16mg·L-1,CaCl24mg·L-1,CuSO4 4mg·L-1,FeSO4 4mg·L-1,MnSO4 4mg·L-1The solvent is water, the pH value is 7.0, and 18 g.L are added simultaneously-1Agar, sterilized at 121 ℃ for 20 min.
Taking sludge in an acrylic acid wastewater treatment pool of a satellite energy company, standing for 24h, taking 10mL, inoculating into a sterile water culture bottle (250mL) containing 100mL, carrying out shaking culture at 30 ℃ and 160rpm for 30min, standing for 2min after shaking is stopped, taking 5mL of suspension, inoculating into a selective culture medium containing 100mL of acrylic acid liquid, carrying out shaking culture at 30 ℃ and 160rpm for 3d, taking 5mL of suspension, inoculating into a selective culture medium containing 100mL of fresh acrylic acid liquid, carrying out shaking culture at 30 ℃ and 160rpm for 3d, repeating the culture for 3 times, and preparing a bacterial liquid with a certain concentration by using sterile water. And (3) separating and purifying the obtained bacterial liquid by using an acrylic acid solid selective medium through multiple flat plate streaking to obtain a single bacterial colony, namely the acrylic acid degrading strain, which is marked as a strain JJ-3.
2. Identification of Strain JJ-3
a. Physiological and biochemical characteristics of Strain JJ-3
The colony color is orange yellow, the round opaque surface is rough and dry, and has no flagellum and no spore. The shape of the cells was observed as spherical or short rod under a transmission electron microscope (FIG. 1). The optimum pH value for growth is 7.0, and the optimum temperature is 35 ℃.
b. 16S rRNA sequence analysis of Strain JJ-3
The strain JJ-3 is determined to be Rhodococcus ruber by 16S rRNA sequence analysis and physiological and biochemical experimental identification.
The method comprises the following specific steps:
the DNA of the strain JJ-3 was extracted and purified using a 3S column centrifugal type environmental sample DNA recovery kit (V2.2, Biotech, Inc., Zhejiang Tian Ke., Ltd.), and stored at 4 ℃. The purified DNA was PCR amplified using bacterial universal primers F27 and 1492R, the primer sequences were:
F27:5’-AGA GTT TGA TCC TGG CTC AG-3’
1492R:5’-GGT TAC CTT GTT ACG ACT T-3’
the PCR reaction system was (50. mu.L): 1.75. mu.L of template DNA, 1. mu.L each of primer F27 and primer R1492, MgCl2(25mmol·L-1)3μL,Taq enzyme (5U. mu.L)-1) 0.25. mu.L, 10 XPCR buffer 5. mu.L, dNTP (2.5 mmol. multidot.L)-1) mu.L, 34. mu.L of redistilled water.
The PCR reaction program was set as: pre-denaturation at 94 ℃ for 4 min; then denaturation at 94 ℃ for 1min, annealing at 59 ℃ for 1min, extension at 72 ℃ for 1.5min, and circulating for 35 cycles; then extending for 10min at 72 ℃; finally, the temperature is kept at 4 ℃ for 10 min. Sequencing the PCR product (Zhejiang family), and the sequencing result is shown in sequence SEQ ID NO: 1 is shown.
The 16S rDNA sequence of the strain JJ-3 was uploaded to the Genbank for homology comparison, and found to belong to the genus Rhodococcus, having the highest homology to Rhodococcus ruber of 100%, and FIG. 2 is a phylogenetic tree diagram of the strain. In order to further determine the reliability of the identification result, the strain JJ-3 is finally determined to belong to Rhodococcus ruber through physiological and biochemical experiments, therefore, the strain JJ-3 is named as Rhodococcus ruber JJ-3 and is preserved in China center for type culture Collection with the preservation number: CCTCC NO: m2019546, date of deposit 2019, month 07, day 12, address: china, wuhan university, zip code: 430072.
example 2 Rhodococcus ruber JJ-3 fermentation broth
(1) Slant culture
Inoculating Rhodococcus ruber JJ-3 to a slant culture medium, and culturing at 30 deg.C for 3 days to obtain a thallus slant; the final concentration of the slant culture medium is as follows: NH (NH)4Cl 540mg·L-1,K2HPO41000mg·L-1,KH2PO41000mg·L-1,NaCl 500mg·L -1200 mg/L yeast extract powder-1Peptone 400 mg. L-1Acrylic acid 1000 mg. L-1,MgSO4·7H2O 16mg·L-1,CaCl24mg·L-1,CuSO4 4mg·L-1,FeSO4 4mg·L-1,MnSO4 4mg·L-1The solvent is water, the pH value is 7.0, and 18 g.L are added simultaneously-1Agar.
(2) Seed culture: selecting colonies from the thallus slant, inoculating the colonies to an LB culture medium, and culturing at 30 ℃ for 12h to obtain a seed solution; the LB mediumThe concentration composition is as follows: NaCl 10 g.L-15 g.L of yeast extract powder-1Peptone 10 g. L-1The solvent is water, and the pH value is 7.0;
(3) fermentation culture: inoculating the seed solution to a fermentation medium at an inoculum size of 5 vol%, culturing at 30 deg.C and 160rpm to OD600When the concentration is 0.8, obtaining a fermentation culture solution, namely the bacterium-containing suspension; the final concentration of the fermentation medium is as follows: NH (NH)4Cl 540mg·L-1,K2HPO4 1000mg·L-1,KH2PO4 1000mg·L-1,NaCl 500mg·L -1200 mg/L yeast extract powder-1Acrylic acid 1000 mg. L-1,MgSO4·7H2O 16mg·L-1,CaCl2 4mg·L-1,CuSO4 4mg·L-1,FeSO44mg·L-1,MnSO4 4mg·L-1The solvent is water, the pH value is 7.0, and the solvent is water.
Example 3: rhodococcus ruber JJ-3 acrylic acid degradation performance detection
1. Investigating the acrylic acid degrading performance of Rhodococcus ruber JJ-3 under different initial pH conditions
The experiments of the Rhodococcus ruber JJ-3 on degrading acrylic acid are carried out at different initial pH values, and the pH value of 7.0 is found to be the optimal pH value, and the degradation rate is the highest at the time, and the concrete implementation steps are as follows:
acrylic acid was used as a carbon source (added at a concentration of 1000 mg. multidot.L)-1) OD was inoculated at a volume concentration of 5%600The culture broth was obtained by inoculating 0.8 suspension containing bacteria (prepared by the method of example 2) to an acrylic acid liquid selection medium (having the same composition as in example 2) having different pH values (pH values of 5.0, 6.0, 7.0, 8.0, 9.0, and 10.0, respectively), culturing at 35 ℃ and 160rpm under shaking for 16 hours, and the concentration of acrylic acid was measured by high performance liquid chromatography, i.e., 1.5mL of the culture broth was used as a sample, the sample was centrifuged, the supernatant was extracted by a 5mL disposable syringe, the remaining microorganisms were removed by filtration through a disposable organic syringe filter having a pore size of 0.22 μm, and the concentration of acrylic acid was measured by high performance liquid chromatography.
Chromatographic conditions are as follows:
chromatographyColumn:
Figure BDA0002345591890000051
C18columns (4.6 mm. times.250 mm, 5 μm); mobile phase: 0.02% phosphoric acid (pH 2.5): 70 parts of methanol: 30, of a nitrogen-containing gas; flow rate 0.6 mL/min-1(ii) a Ultraviolet detection wavelength: 210 nm; column temperature: 26 ℃; sample introduction amount: 10 μ L.
As shown in FIG. 3, the environmental suitability of the strain JJ-3 to weak acids is relatively poor, and when the pH is 7.0, the acrylic acid degradation effect of the strain JJ-3 is the best, and reaches 98.0%.
2. Investigating the acrylic acid degrading performance of Rhodococcus ruber JJ-3 under different temperature conditions
The experiment of degrading acrylic acid by Rhodococcus ruber JJ-3 is carried out at different temperatures, and the temperature is found to be the optimal temperature at 35 ℃, and the degradation rate is the highest at the time, and the specific implementation steps are as follows:
acrylic acid as carbon source (concentration 1000 mg. L)-1) OD was inoculated at a volume concentration of 5%6000.8 of the suspension containing the bacteria (prepared by the method of example 2) was inoculated into an acrylic acid liquid selection medium (pH 7.0, composition same as in example 2) at 25.0, 30.0, 35.0, 40.0, 45.0 ℃ respectively, and cultured with shaking at 160rpm for 16 hours to obtain a culture solution, the concentration of acrylic acid was measured by high performance liquid chromatography (same as in step 1),
as shown in FIG. 4, the environmental suitability of the strain JJ-3 to low temperature is relatively poor, and the acrylic acid degradation effect of the strain JJ-3 is the best, reaching 98.7%, when the temperature is 35.0 ℃.
3. Investigating the acrylic acid degrading performance of Rhodococcus ruber JJ-3 under different inoculation amounts
The acrylic acid degradation experiment of Rhodococcus ruber JJ-3 is carried out under different inoculation amounts, and the result shows that the optimal inoculation amount is 15% (v/v), and the specific embodiment is as follows:
acrylic acid was used as a carbon source (added at a concentration of 1000 mg. multidot.L)-1) OD was graded by 6 inoculations at different volume concentrations (3%, 5%, 8%, 10%, 15%, 20%) respectively600The bacterial suspension (prepared in example 2) containing 0.8 was inoculated into an acrylic acid liquid selection medium (composition: same as in example 2) at 35 ℃ under 160rpmCulturing for 16h by shaking to obtain a culture solution, measuring the concentration of acrylic acid by adopting a high performance liquid chromatography (same as the step 1), wherein the result is shown in figure 5, and the data show that when the inoculation amount is more than 3%, because nutrient substances in the culture medium are sufficient at the beginning, the inoculation amount directly influences the degradation of the substrate, the degradation rate slowly rises along with the increase of the inoculation amount, when the inoculation amount is 15%, the degradation rate reaches 100%, and along with the continuous increase of the inoculation amount, a competition relation to the nutrient substances is formed among the degrading bacteria, so that the reduction rate is reduced. As can be seen from the figure, the optimal inoculation amount of the strain JJ-3 is about 10-15%.
4. Investigating the acrylic acid degrading performance of Rhodococcus ruber JJ-3 under different initial acrylic acid concentrations
The degradation experiment of Rhodococcus ruber JJ-3 is carried out under different initial acrylic acid concentrations, and the result shows that the degradation experiment of Rhodococcus ruber JJ-3 is carried out under the condition that the acrylic acid concentration is 100--1The acrylic acid can be effectively degraded, and the specific embodiment is as follows:
using acrylic acid as carbon source, inoculating OD according to 15% volume concentration600The bacterial-containing suspensions (prepared in example 2) of 0.8 were inoculated respectively at an initial acrylic acid concentration of 100 mg. multidot.L-1、200mg·L-1、400mg·L-1、600mg·L-1、800mg·L-1、1000mg·L-1、2000mg·L-1The culture broth was obtained by shaking-culturing in 7 acrylic acid liquid selective media (composition same as example 2) at 35 ℃ and 160rpm for 16 hours, and the concentration of acrylic acid was measured by high performance liquid chromatography (same as step 1), with the results shown in FIG. 6: at 100--1Under the initial acrylic acid concentration, the Rhodococcus ruber JJ-3 can effectively degrade acrylic acid, and the degradation rate is 100%.
Sequence listing
<110> Zhejiang industrial university
<120> Rhodococcus ruber JJ-3 and application thereof in acrylic acid degradation
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1319
<212> DNA
<213> Rhodococcus ruber (Rhodococcus ruber)
<400> 1
tagtggcgaa cgggtgagta acacgtgggt gatctgccct gcacttcggg ataagcctgg 60
gaaactgggt ctaataccgg ataggacctc gggatgcatg ttccggggtg gaaaggtttt 120
ccggtgcagg atgggcccgc ggcctatcag cttgttggtg gggtaacggc ccaccaaggc 180
gacgacgggt agccggcctg agagggcgac cggccacact gggactgaga cacggcccag 240
actcctacgg gaggcagcag tggggaatat tgcacaatgg gcgcaagcct gatgcagcga 300
cgccgcgtga gggatgacgg ccttcgggtt gtaaacctct ttcagtaccg acgaagcgca 360
agtgacggta ggtacagaag aagcaccggc caactacgtg ccagcagccg cggtaatacg 420
tagggtgcga gcgttgtccg gaattactgg gcgtaaagag ctcgtaggcg gtttgtcgcg 480
tcgtctgtga aaacccgcag ctcaactgcg ggcttgcagg cgatacgggc agacttgagt 540
actgcagggg agactggaat tcctggtgta gcggtgaaat gcgcagatat caggaggaac 600
accggtggcg aaggcgggtc tctgggcagt aactgacgct gaggagcgaa agcgtgggta 660
gcgaacagga ttagataccc tggtagtcca cgccgtaaac ggtgggcgct aggtgtgggt 720
ttccttccac gggatccgtg ccgtagctaa cgcattaagc gccccgcctg gggagtacgg 780
ccgcaaggct aaaactcaaa ggaattgacg ggggcccgca caagcggcgg agcatgtgga 840
ttaattcgat gcaacgcgaa gaaccttacc tgggtttgac atacaccgga ccgccccaga 900
gatggggttt cccttgtggt cggtgtacag gtggtgcatg gctgtcgtca gctcgtgtcg 960
tgagatgttg ggttaagtcc cgcaacgagc gcaacccttg tcctgtgttg ccagcacgta 1020
atggtgggga ctcgcaggag actgccgggg tcaactcgga ggaaggtggg gacgacgtca 1080
agtcatcatg ccccttatgt ccagggcttc acacatgcta caatggccgg tacagagggc 1140
tgcgataccg cgaggtggag cgaatccctt aaagccggtc tcagttcgga tcggggtctg 1200
caactcgacc ccgtgaagtc ggagtcgcta gtaatcgcag atcagcaacg ctgcggtgaa 1260
tacgttcccg ggccttgtac acaccgcccg tcacgtcatg aaagtcggta acacccgaa 1319

Claims (8)

1.Rhodococcus ruber (C)Rhodococcus ruber) JJ-3, preserved in China center for type culture Collection, with the preservation number: CCTCC NO: m2019546, date of deposit 2019, month 07, day 12, address: china, wuhan university, zip code: 430072.
2. use of Rhodococcus ruber JJ-3 according to claim 1 for the biodegradation of acrylic acid.
3. The use according to claim 2, characterized in that said use is: inoculating a fermentation liquid, namely a bacterium-containing suspension, obtained by carrying out fermentation culture on Rhodococcus ruber JJ-3 into an acrylic acid liquid selection culture medium, and culturing at the temperature of 20-45 ℃ and the speed of 140-160rpm by taking acrylic acid as a carbon source to realize acrylic acid degradation; the final concentration composition of the acrylic acid liquid selective medium is as follows: NH (NH)4Cl 540mg·L-1,K2HPO4 1000mg·L-1,KH2PO4 1000mg·L-1,NaCl 500mg·L-1200 mg/L yeast extract powder-1Acrylic acid 100-2000 mg. L-1,MgSO4·7H2O 16mg·L-1,CaCl2 4mg·L-1,CuSO4 4mg·L-1,FeSO4 4mg·L-1,MnSO4 4mg·L-1The solvent is water, and the pH value is 5.0-10.0.
4. Use according to claim 3, characterized in that the pH is 7.0.
5. Use according to claim 3, characterized in that the temperature is 35 ℃.
6. The use according to claim 3, wherein the OD of the suspension comprising bacterial cells is600The value is 0.7-1.0, and the volume inoculation amount of the bacterial suspension is 3-20%.
7. The use according to claim 3, wherein the initial concentration of acrylic acid is 100--1
8. Use according to claim 3, wherein the bacterial-containing suspension is prepared by the steps of:
(1) slant culture: inoculating Rhodococcus ruber JJ-3 to a slant culture medium, and culturing at 30 deg.C for 3 days to obtain a thallus slant; the final concentration of the slant culture medium is as follows: NH (NH)4Cl 540mg·L-1,K2HPO4 1000mg·L-1,KH2PO4 1000mg·L-1,NaCl 500mg·L-1200 mg/L yeast extract powder-1Peptone 400 mg. L-1Acrylic acid 1000 mg. L-1,MgSO4·7H2O 16mg·L-1,CaCl2 4mg·L-1,CuSO4 4mg·L-1,FeSO4 4mg·L-1,MnSO4 4mg·L-1The solvent is water, the pH value is 7.0, and the agar is 18-20 g.L-1
(2) Seed culture: selecting colonies from the thallus slant, inoculating the colonies to an LB culture medium, and culturing at 30 ℃ for 18h to obtain a seed solution; the LB culture medium has the following final concentration composition: NaCl 10 g.L-15 g.L of yeast extract powder-1Peptone 10 g. L-1The solvent is water, and the pH value is 7.0;
(3) fermentation culture: inoculating the seed solution to a fermentation culture medium by an inoculation amount with the volume concentration of 3-20%, and culturing at 30 ℃ to obtain a fermentation culture solution, namely a bacterium-containing suspension; the final concentration of the fermentation medium is as follows: NH (NH)4Cl 540mg·L-1,K2HPO41000mg·L-1,KH2PO4 1000mg·L-1,NaCl 500mg·L-1200 mg/L yeast extract powder-1Acrylic acid 1000 mg. L-1,MgSO4·7H2O 16mg·L-1,CaCl2 4mg·L-1,CuSO4 4mg·L-1,FeSO4 4mg·L-1,MnSO4 4mg·L-1The solvent is water, and the pH value is 7.0.
CN201911393310.3A 2019-12-30 2019-12-30 Rhodococcus ruber JJ-3 and application thereof in acrylic acid degradation Active CN111269848B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201911393310.3A CN111269848B (en) 2019-12-30 2019-12-30 Rhodococcus ruber JJ-3 and application thereof in acrylic acid degradation

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201911393310.3A CN111269848B (en) 2019-12-30 2019-12-30 Rhodococcus ruber JJ-3 and application thereof in acrylic acid degradation

Publications (2)

Publication Number Publication Date
CN111269848A CN111269848A (en) 2020-06-12
CN111269848B true CN111269848B (en) 2022-03-18

Family

ID=71001534

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201911393310.3A Active CN111269848B (en) 2019-12-30 2019-12-30 Rhodococcus ruber JJ-3 and application thereof in acrylic acid degradation

Country Status (1)

Country Link
CN (1) CN111269848B (en)

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN113046267B (en) * 2021-03-22 2022-11-15 哈尔滨师范大学 Rhodococcus ruber and application thereof
CN114045238B (en) * 2021-11-08 2023-09-29 浙江树人学院(浙江树人大学) Rhodococcus ruber HJM-8 capable of efficiently degrading dimethylacetamide and application thereof

Citations (21)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1998058072A1 (en) * 1997-06-19 1998-12-23 Ciba Speciality Chemicals Water Treatments Limited Flocculation of biological material from organic acid-containing systems
WO2006007957A1 (en) * 2004-07-19 2006-01-26 Ciba Specialty Chemicals Water Treatments Limited Process for preparing monomers and polymers thereof
TW200726840A (en) * 2006-01-10 2007-07-16 Lg Chemical Ltd Rhodococcus erythropolis LG12 having acrylic acid degrading activity and method for removing acrylic acid using the same
CN101033454A (en) * 2006-04-29 2007-09-12 浙江工业大学 Erythro micrococcus ZJB-064 and application thereof
CN101268196A (en) * 2005-09-19 2008-09-17 西巴特殊化学品控股有限公司 Biocatalytic manufacturing of (meth)acrylic esters
CN102010826A (en) * 2009-07-24 2011-04-13 大野绿水株式会社 Preserving method of microbe and suspension of microbe
CN102965309A (en) * 2012-11-14 2013-03-13 浙江工业大学 Rhodococcus sp. and application thereof to micro-biologically degrading 4-fluorocinnamic acid
EP2744906A1 (en) * 2011-08-19 2014-06-25 Genomatica, Inc. Microorganisms and methods for producing 2,4-pentadienoate, butadiene, propylene, 1,3-butanediol and related alcohols
CN104673710A (en) * 2014-12-31 2015-06-03 浙江至美环境科技有限公司 Rhodococcus sp. strain and application thereof
CN104726364A (en) * 2015-01-09 2015-06-24 江苏大学 Rhodococcus and application of rhodococcus in degradation of lignin
CN104745645A (en) * 2015-02-11 2015-07-01 江南大学 Method for synthesizing acrylic acid through biotransformation of acraldehyde
WO2016050819A1 (en) * 2014-09-30 2016-04-07 Basf Se Method for preparing an acrylamide solution having a low acrylic acid concentration
CN106715702A (en) * 2014-09-30 2017-05-24 巴斯夫欧洲公司 Method for preparing aqueous acrylamide solution having low acrylic acid concentration
EP3194604A1 (en) * 2014-09-18 2017-07-26 Genomatica, Inc. Non-natural microbial organisms with improved energetic efficiency
CN107075494A (en) * 2014-09-30 2017-08-18 巴斯夫欧洲公司 Prepare the means and method of the amide compound with less acrylic acid
EP3225693A1 (en) * 2016-03-29 2017-10-04 Basf Se Method for preparing aqueous acrylamide solutions
CN108410756A (en) * 2018-02-11 2018-08-17 浙江工业大学 Thermophilic pyridine Rhodococcus sp and its application in degradable organic pollutant
CN108410771A (en) * 2018-04-08 2018-08-17 宁夏大学 The application of Rhodococcus sp YC915 and its absorption microbial inoculum in soil phthalic acid ester of degrading
CN109576197A (en) * 2019-01-31 2019-04-05 平湖石化有限责任公司 A kind of acrylic acid degrading bacterium and its application
CN110499274A (en) * 2019-09-18 2019-11-26 清华大学 A kind of genetic engineering Rhodococcus sp and its construction method and application
CN113046267A (en) * 2021-03-22 2021-06-29 哈尔滨师范大学 Rhodococcus ruber and application thereof

Patent Citations (22)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1998058072A1 (en) * 1997-06-19 1998-12-23 Ciba Speciality Chemicals Water Treatments Limited Flocculation of biological material from organic acid-containing systems
WO2006007957A1 (en) * 2004-07-19 2006-01-26 Ciba Specialty Chemicals Water Treatments Limited Process for preparing monomers and polymers thereof
CN1989251A (en) * 2004-07-19 2007-06-27 西巴特殊化学水处理有限公司 Process for preparing monomers and polymers thereof
CN101268196A (en) * 2005-09-19 2008-09-17 西巴特殊化学品控股有限公司 Biocatalytic manufacturing of (meth)acrylic esters
TW200726840A (en) * 2006-01-10 2007-07-16 Lg Chemical Ltd Rhodococcus erythropolis LG12 having acrylic acid degrading activity and method for removing acrylic acid using the same
CN101033454A (en) * 2006-04-29 2007-09-12 浙江工业大学 Erythro micrococcus ZJB-064 and application thereof
CN102010826A (en) * 2009-07-24 2011-04-13 大野绿水株式会社 Preserving method of microbe and suspension of microbe
EP2744906A1 (en) * 2011-08-19 2014-06-25 Genomatica, Inc. Microorganisms and methods for producing 2,4-pentadienoate, butadiene, propylene, 1,3-butanediol and related alcohols
CN102965309A (en) * 2012-11-14 2013-03-13 浙江工业大学 Rhodococcus sp. and application thereof to micro-biologically degrading 4-fluorocinnamic acid
EP3194604A1 (en) * 2014-09-18 2017-07-26 Genomatica, Inc. Non-natural microbial organisms with improved energetic efficiency
CN107075494A (en) * 2014-09-30 2017-08-18 巴斯夫欧洲公司 Prepare the means and method of the amide compound with less acrylic acid
WO2016050819A1 (en) * 2014-09-30 2016-04-07 Basf Se Method for preparing an acrylamide solution having a low acrylic acid concentration
CN106715702A (en) * 2014-09-30 2017-05-24 巴斯夫欧洲公司 Method for preparing aqueous acrylamide solution having low acrylic acid concentration
CN104673710A (en) * 2014-12-31 2015-06-03 浙江至美环境科技有限公司 Rhodococcus sp. strain and application thereof
CN104726364A (en) * 2015-01-09 2015-06-24 江苏大学 Rhodococcus and application of rhodococcus in degradation of lignin
CN104745645A (en) * 2015-02-11 2015-07-01 江南大学 Method for synthesizing acrylic acid through biotransformation of acraldehyde
EP3225693A1 (en) * 2016-03-29 2017-10-04 Basf Se Method for preparing aqueous acrylamide solutions
CN108410756A (en) * 2018-02-11 2018-08-17 浙江工业大学 Thermophilic pyridine Rhodococcus sp and its application in degradable organic pollutant
CN108410771A (en) * 2018-04-08 2018-08-17 宁夏大学 The application of Rhodococcus sp YC915 and its absorption microbial inoculum in soil phthalic acid ester of degrading
CN109576197A (en) * 2019-01-31 2019-04-05 平湖石化有限责任公司 A kind of acrylic acid degrading bacterium and its application
CN110499274A (en) * 2019-09-18 2019-11-26 清华大学 A kind of genetic engineering Rhodococcus sp and its construction method and application
CN113046267A (en) * 2021-03-22 2021-06-29 哈尔滨师范大学 Rhodococcus ruber and application thereof

Non-Patent Citations (10)

* Cited by examiner, † Cited by third party
Title
Achromobacter sp.TB一氧化氮还原酶基因的克隆表达及还原性能研究;王雨;《中国优秀硕士学位论文全文数据库(电子期刊)工程科技I辑》;20200315;全文 *
PATHWAY AND ENZYMOLOGY OF BY MYCOBACTERIUM VACCAE JOBsCYCLIC ETHER BIODEGRADATION;Renny Lan;《North Carolina State Universitv》;20070322;全文 *
Rapid and Complete Biodegradation of Acrylic Acid by a Novel Strain Rhodococcus ruber JJ-3: Kinetics, Carbon Balance, and Degradation Pathways;Jinjia He 等;《Biotechnology and Bioprocess Engineering》;20200831;第589-598页 *
一株环己酮高效降解菌株的筛选及鉴定;何倩 等;《吉首人学学报(自然科学版)》;20080325;第92-95页 *
一株石油降解赤红球菌(Rhodococcus rubber)特性及处理含油废水研究;刘猛 等;《环境科学学报》;20160318;第3651-3657页 *
印染废水强化处理的高效菌剂研发与试验研究;王冬;《中国优秀硕士学位论文全文数据库(电子期刊)》;20161015;全文 *
可降解三乙胺的赤红球菌S6-的筛选与鉴定及降解特性;孙磊 等;《环境科学研究》;20161231;第1882-1886页 *
石油烃类污染对青藏高原北麓河地区冻区土壤微生物多样性的影响;王宝山等;《环境工程学报》;20181005(第10期);全文 *
醋酸杆菌生物催化丙烯醛合成丙烯酸;王旭昌等;《应用与环境生物学报》;20150825(第04期);全文 *
金属离子对赤红球菌( Rhodococcus ruber L9)降解芘的影响及其作用机制;阮珍 等;《环境化学》;20191209;第2649-2656页 *

Also Published As

Publication number Publication date
CN111269848A (en) 2020-06-12

Similar Documents

Publication Publication Date Title
US10350546B2 (en) Fungi-bacteria composite microecologics and methods for preparing and using the same
CN110643534B (en) Phellinus deltoidea capable of degrading triphenyl phosphate
CN111269848B (en) Rhodococcus ruber JJ-3 and application thereof in acrylic acid degradation
CN110656066A (en) Acinetobacter strain for shortcut nitrification and denitrification and application thereof
CN110699291B (en) Achromobacter xylosoxidans with sulfide degradation performance and application thereof
CN113462622B (en) Pseudomonas for efficiently degrading various aromatic pollutants and application thereof
CN113234625B (en) Acinetobacter pittericus and application thereof
CN114045238A (en) Rhodococcus ruber HJM-8 capable of efficiently degrading dimethylacetamide and application thereof
CN113980851A (en) Paracoccus YBH-X with dimethylacetamide degradation capability and application thereof
CN109576197B (en) Acrylic acid degrading bacterium and application thereof
CN110819553B (en) Bacillus aryabhattai and application thereof in acrylic acid degradation
CN114231451B (en) Salt-tolerant bacillus capable of efficiently degrading bisphenol A and application thereof
CN104745515B (en) A kind of acinetobacter calcoaceticus of degrading polycyclic aromatic hydrocarbons and its application
CN103011423A (en) Application of Bacillus cereus DS1 in degradation of organic pollutants in saponin waste water
CN106399200B (en) Alcaligenes and application thereof in high-salt high-polymer wastewater
CN105670965B (en) Strain with iron reduction capacity and application thereof
CN103013875B (en) Bacillus cereus DS1
CN100386428C (en) Chloride ion resistant strain S616 and its screening method
CN103013874B (en) Bacillus subtilis ds3
CN112940995A (en) Arthrobacter capable of degrading n-eicosane and application thereof
CN115010269B (en) Application of halophila campaigns in uranium-containing wastewater treatment
CN114350544B (en) Pseudomonas stutzeri WX3-1 for efficiently degrading trioctyl phosphate and application thereof
CN114292771B (en) Pseudomonas stutzeri WZY-3-1 for efficiently degrading anthrone and application thereof
CN116925932B (en) Trichoderma Guizhou fungus and application thereof
CN114990027B (en) Pseudomonas W52 for degrading halogenated acid and application thereof

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant