CN106399191B - The application of one plant of arthrobacterium B2 and its nitric wastewater of degrading - Google Patents

The application of one plant of arthrobacterium B2 and its nitric wastewater of degrading Download PDF

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CN106399191B
CN106399191B CN201610930437.4A CN201610930437A CN106399191B CN 106399191 B CN106399191 B CN 106399191B CN 201610930437 A CN201610930437 A CN 201610930437A CN 106399191 B CN106399191 B CN 106399191B
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arthrobacterium
culture
thallus
application
follows
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CN106399191A (en
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周煕城
张正波
王雪玲
蔡畅
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Langjing Environmental Protection Technology Co ltd
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Hangzhou Langjing Environmental Protection Technology Co ltd
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • C12R2001/06—Arthrobacter
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, 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
    • C12N1/20—Bacteria; Culture media therefor
    • C12N1/205—Bacterial isolates
    • C—CHEMISTRY; METALLURGY
    • C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F3/00—Biological treatment of water, waste water, or sewage
    • C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, 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
    • C12N1/20—Bacteria; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F2101/00—Nature of the contaminant
    • C02F2101/10—Inorganic compounds
    • C02F2101/16—Nitrogen compounds, e.g. ammonia

Abstract

The present invention provides the applications of one plant of arthrobacterium B2 and its nitric wastewater of degrading, arthrobacterium (Arthrobacter sp.) B2 was preserved in China typical culture collection center on September 14th, 2016, deposit number: CCTCC NO:2016485, arthrobacterium B2 denitrifying capacity with higher under aerobic condition realizes nitration reaction and anti-nitration reaction.The organic matters such as organic matter, ammonia nitrogen and the cultured water that can effectively degrade in sewage treatment oxidation pond in water body, realize the purified treatment of water body.Therefore, arthrobacterium B2 can be applied in sewage treatment and cultivation wastewater purification industry at microbial inoculum.Nitrate degradation rate is 87.6%, and nitrite degradation rate is 84.5%.

Description

The application of one plant of arthrobacterium B2 and its nitric wastewater of degrading
(1) technical field
The present invention relates to one plant of heterotrophism-aerobic denitrifying bacteria, in particular to a kind of arthrobacterium B2 and its in sewage treatment Application.
(2) background technique
Biological denitrificaion is the most economical effective method that nitrate pollution is removed in sewage treatment.Traditional nitrogen removal technique It is to be realized by the different process of nitrification and denitrification the two realization conditions.In aerobic aeration area, pass through nitrifying bacteria community Effect, ammonium oxidation are converted into nitrate nitrogen；In anaerobic zone, by the effect of denitrifying bacterium, most nitrate reduction turns Nitrous oxide and nitrogen are turned to, the nutriment of bacterium itself is converted into there are also a small part, to achieve the purpose that denitrogenation. The nitrogen processing technological flow is long, system HRT longer, needs biggish aeration tank, and investment and operating cost are high；Remain higher Biological concentration and good denitrification effect, it is necessary to while carrying out sludge and nitrification liquid reflux etc..In recent years, Many researchers are found Denitrification is also able to achieve under aerobic condition.It may be implemented synchronous nitration and denitrification, i.e. nitration reaction and anti-nitration reaction can be with It is carried out simultaneously in aerobic tank, to greatly reduce system space and project cost, reduces operating cost.
(3) summary of the invention
It is an object of the present invention to provide a kind of arthrobacterium B2, with stronger heterotrophism-aerobic denitrification ability, to realize Nitration reaction is with anti-nitration reaction synchronous under aerobic condition carries out.It can be gone ammonia nitrogen with higher under conditions of being aerated aerobic Removing solid capacity realizes the biological denitrificaion of sewage.
The technical solution adopted by the present invention is that:
One plant of new strains -- arthrobacterium (Arthrobacter sp.) B2 is preserved in Chinese Typical Representative culture for present invention offer Collection, preservation date: on September 14th, 2016, deposit number: CCTCC NO:2016485, address: China, Wuhan, Wuhan University, postcode: 430072.
The present invention also provides a kind of application of arthrobacterium B2 in degradation nitric wastewater.Total nitrogen in the ammonia nitrogen waste water Higher, it is higher that ammonia nitrogen in sewage accounts for total nitrogen ratio, COD 300-600mg/L, total nitrogen 100-300mg/L, ammonia nitrogen 20-50mg/L.
Further, the application is that the wet thallus obtained with the fermented culture of arthrobacterium B2 mixes made of drying with carrier Microbial inoculum is catalyst, is 1-3m in ventilatory capacity using nitric wastewater as substrate3/ h, under conditions of 26-30 DEG C of temperature, in aeration tank Middle carry out degradation reaction realizes the fast degradation of ammonia nitrogen and total nitrogen in nitric wastewater；The carrier is diatomite or bentonite.
Further, the wet thallus and carrier quality ratio are 1:50-200, and preferably 1:100, the dosage of the microbial inoculum is to contain Nitrogen sewage volume is calculated as 1.0-5.0g/L, preferably 1.5g/L.
Further, the wet thallus is prepared as follows:
(1) arthrobacterium B2 is seeded to LB solid medium, in 37 DEG C of culture 26h, obtains thallus inclined-plane；The LB solid Culture medium final concentration composition are as follows: tryptone 10g/L, yeast extract 5g/L, sodium chloride 10g/L, agar 20g/L, solvent are Water, pH 7.0；
(2) it is seeded in LB liquid medium from one ring thallus of thallus inclined-plane picking, 37 DEG C of culture 28h, forms seed liquor； LB liquid medium final concentration composition are as follows: tryptone 10g/L, yeast extract 5g/L, sodium chloride 10g/L, solvent are water, pH 7.0；
(3) seed liquor is transferred to the fermentor equipped with fermentation medium with the inoculum concentration of volumetric concentration 3% and carries out fermentation training It supports, volume liquid amount 70%, fermentation temperature is 37 ± 1 DEG C, stirring rate 200rpm, ventilatory capacity 0.9-1.1V/vm, molten Oxygen concentration is 80-90%, culture to for 24 hours when, fermentation liquid is centrifuged, harvest wet thallus；The fermentation medium final concentration composition Are as follows: tryptone 10g/L, yeast extract 5g/L, sodium chloride 10g/L, solvent are water, pH 7.0.
Compared with prior art, beneficial effect of the present invention is mainly reflected in: arthrobacterium B2 has heterotrophism-aerobic denitrification function Can, it not only can use the organic matter in sewage, nitrogen source is grown, but also organic matter, ammonia nitrogen and nitrous in the sewage that can degrade Sour nitrogen pollutant.Nitrate degradation rate is 87.6%, and nitrite degradation rate is 84.5%.
(3) Detailed description of the invention
Fig. 1 is bacterial strain B2 phylogenetic tree.
(4) specific embodiment
The present invention is described further combined with specific embodiments below, but protection scope of the present invention is not limited in This:
The screening and identification of 1 bacterial strain B2 of embodiment
(1) bacterial strain B2 is screened
The activated sludge being derived from Lishui of Zhejiang sewage treatment plant aeration tank is sample, is carried out using heterotrophism method aerobic anti- The screening technique of nitrifier is (referring in particular to the screening of high-efficiency aerobic denitrifying bacteria and the research of nitrogen removal characteristics, Cai Changfeng, beam Environmental science and technology of heap of stone, 2011,34 (1): 41-44.), isolated bacterial strain B2；Its colonial morphology are as follows: bacterium colony milky, Opaque, bacterium colony surface wettability, intermediate projections, colony edge is smooth.
(2) bacterial strain B2 is identified
Extract bacterial strain B2 genome, measurement 16sRNA (reference: the separation identification and denitrogenation of the poor nutrition aerobic denitrifying bacteria of strain Characteristic, Wei Wei, Huang Tinglin, Su Junfeng etc.；Ecological environment journal, 2010,19 (9): 2166-2171.), 16sRNA nucleotides sequence It is classified as shown in SEQ ID NO.1.It compares and analyzes by 16sRNA, the homology that bacterial strain B2 and Arthrobacter sp. belongs to reaches 99%, and the bacterial strain is identified (Fig. 1, Tables 1 and 2) using Biolog (GEN III) microbial identification system, and with report Bacterial strain highest similarity degree is 0.368, not high with known bacterial strain similarity degree, is new discovery bacterial strain.Therefore it names are as follows: arthrobacterium (Arthrobacter sp.)B2.Biolog system provides 36h qualification result, as shown in Table 1 and Table 2.
Table 1, bacterial strain B2 are to the Utilization abilities of 71 kinds of carbon sources on III plate of Biolog GEN
Notes:+,positive；-,negative；B,borderline
Table 2, bacterial strain B2 are to the chemosensitivities of 23 kinds of chemical substances on III plate of Biolog GEN
Notes:+,positive；-,negative；B,borderline
Embodiment 2
(1) microbial inoculum prepares
Arthrobacterium B2 is seeded to LB solid medium, in 37 DEG C of culture 26h, obtains thallus inclined-plane；The LB solid training Supporting base final concentration composition are as follows: tryptone 10g/L, yeast extract 5g/L, sodium chloride 10g/L, agar 20g/L, solvent are water, pH 7.0。
It is seeded in LB liquid medium from one ring thallus of thallus inclined-plane picking, 37 DEG C of culture 28h, forms seed liquor, Concentration is 1.6 × 108cfu/mL.LB liquid medium final concentration composition are as follows: tryptone 10g/L, yeast extract 5g/L, chlorine Change sodium 10g/L, solvent is water, pH 7.0.
Seed liquor is transferred to the 30L seeding tank equipped with fermentation medium with the inoculum concentration of 3% (volume ratio) and carries out fermentation training It supports, liquid amount 70%, fermentation temperature is 37 ± 1 DEG C, stirring rate 200rpm, ventilatory capacity 0.9-1.1V/vm, and dissolved oxygen is dense Degree is 80-90%, and every primary light absorption value for surveying 600nm of 3h sampling from after being inoculated with determines that the logarithmic growth phase of strain is 9-16h. When culture to for 24 hours when, fermentation liquid is centrifuged, harvest wet thallus 670g.The fermentation medium final concentration composition are as follows: tryptone 10g/L, yeast extract 5g/L, sodium chloride 10g/L, solvent are water, pH 7.0.
100g wet thallus is uniformly mixed with diatomite with mass ratio 1:100,8-10 DEG C of low temperature air-dry microbial inoculum is made 1080g。
(2) measurement of aerobic denitrification intensity
The microbial inoculum of step (1) preparation is measured to the denitrification intensity of the bacterium with Giltay method, moral Han Shi is small within the 2nd day Gas is produced in pipe, and culture medium color becomes blue from green；After 48h, nitrate degradation rate is 87.6%, nitrite degradation rate It is 84.5%.
Embodiment 3
Mr. Yu's sewage plant water inlet takes sanitary sewage 20L, in laboratory simulation aeration tank, ventilatory capacity 1m3/h；Inoculation Microbial inoculum amount is 30g；Handling the time is for 24 hours.Sewage before handling after sewage, addition microbial inoculum processing for 24 hours, and be not added at microbial inoculum Sewage after managing for 24 hours (is compareed) COD and is measured using potassium dichromate method, and TN (total nitrogen) uses potassium persulfate oxidation/ultraviolet spectrometry Photometry measurement；NH3- N (ammonia nitrogen) is measured using Nessler's reagent photometer.The results are shown in Table 3.
3 microbial inoculum wastewater treatment efficiency of table
Remarks: sanitary sewage ----refer to the sanitary sewage biochemical indicator before processing；Microbial inoculum --- refer at sanitary sewage microbial inoculum Biochemical indicator after reason；Control --- refer to that sanitary sewage is not added with microbial inoculum biochemistry of (aeration) under the processing of same physical condition and refers to Mark.
Embodiment 4
Factory is cultivated in Wenling white shrimp, takes breeding wastewater 20L, in laboratory simulation aeration tank, ventilatory capacity are as follows: 1m3/h； It is inoculated with microbial inoculum 30g；Handling the time is for 24 hours.Sewage before handling after sewage, addition microbial inoculum processing for 24 hours, and be not added at microbial inoculum Sewage after managing for 24 hours (is compareed) COD and is measured using potassium dichromate method, NO2-The measurement of-N spectrophotometry；NH3-N (ammonia nitrogen): it receives The measurement of family name's reagent photometry.The results are shown in Table 4.
4 microbial inoculum wastewater treatment efficiency of table
Remarks: breeding wastewater ----refer to the sanitary sewage biochemical indicator before processing；Microbial inoculum --- refer at breeding wastewater microbial inoculum Biochemical indicator after reason；Control --- refer to that breeding wastewater is not added with microbial inoculum biochemistry of (aeration) under the processing of same physical condition and refers to Mark.
SEQUENCE LISTING
<110>Zhejiang Polytechnical University
The application of<120>one plants of arthrobacterium B2 and its nitric wastewater of degrading
<130>
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 1523
<212> DNA
<213> Arthrobacter nicotianae
<400> 1
agagtttgat cctggctcag gatgaacgct ggcggcgtgc ttaacacatg caagtcgaac 60
gatgaagccg aagcttgctt tggtggatta gtggcgaacg ggtgagtaac acgtgagtaa 120
cctgcccctg actctgggat aagcctggga aactgggtct aataccggat atgactgcgg 180
aacgcatgtt ctgtggtgga aagatttatc ggttggggat ggactcgcgg cctatcagct 240
tgttggtgag gtaatggctc accaaggcga cgacgggtag ccggcctgag agggtgaccg 300
gccacactgg gactgagaca cggcccagac tcctacggga ggcagcagtg gggaatattg 360
cacaatgggc gaaagcccga tgcagcgacg ccgcgtgagg gatgacggcc ttcgggttgt 420
aaacctcttt cagtagggaa gaagcgagag tgacggtacc tgcagaagaa gcgccggcta 480
actacgtgcc agcagccgcg gtaatacgta gggcgcaagc gttatccgga tttattgggc 540
gtaaagagct cgtaggcggt ttgtcgcgtc tgccgtgaaa gtccgaggct caacctcgga 600
tctgcggtgg gtacgggcag actagagtga tgtaggggag actggaattc ctggtgtagc 660
ggtgaaatgc gcagatatca ggaggaacac cgatggcgaa ggcaggtctc tgggcattta 720
ctgacgctga ggagcgaaag catggggagc gaacaggatt agataccctg gtagtccatg 780
ccgtaaacgt tgggcactag gtgtggggga cattccacgt tttccgcgcc gtagctaacg 840
cattaagtgc cccgcctggg gagtacggcc gcaaggctaa aactcaaagg aattgacggg 900
ggcccgcaca agcggcggag catgcggatt aattcgatgc aacgcgaaga accttaccaa 960
ggcttgacat gttccagacc gggctagaga tagtccttcc cttcggggct ggttcacagg 1020
tggtgcatgg ttgtcgtcag ctcgtgtcgt gagatgttgg gttaagtccc gcaacgagcg 1080
caaccctcgt tccatgttgc cagcacgtag tggtggggac tcatgggaga ctgccggggt 1140
caactcggag gaaggtgggg atgacgtcaa atcatcatgc cccttatgtc ttgggcttca 1200
cgcatgctac aatggccggt acaatgggtt gcgatactgt gaggtggagc taatccctaa 1260
aagccggtct ccgttcggat tggggtctgc aactcgaccc catgaagtcg gagtcgctag 1320
taatcgcaga tcagcaacgc tgcggtgaat acgttcccgg gccttgtaca caccgcccgt 1380
caagtcacga aagttggtaa cacccgaagc cgatggccta accaccttgt gtggggggag 1440
tcgtcgaagg tgggactggc gattgggact aagtcgtaac aaggtagccg taccggaagg 1500
tgcggctgga tcacctcctt tct 1523

Claims (6)

1. arthrobacterium (Arthrobacter sp.) B2, is preserved in China typical culture collection center, preservation date: 2016 September 14 days, deposit number: CCTCC NO:2016485, address: China, Wuhan, Wuhan University, postcode: 430072.
2. a kind of application of the arthrobacterium B2 described in claim 1 in degradation nitric wastewater.
3. application as claimed in claim 2, it is characterised in that COD is 300-600mg/L, total nitrogen 100- in the nitric wastewater 300mg/L, ammonia nitrogen 20-50mg/L.
4. application as claimed in claim 3, it is characterised in that the application is with the wet of the fermented culture acquisition of arthrobacterium B2 It is catalyst that thallus, which mixes microbial inoculum made of drying with carrier, is 1-3m in ventilatory capacity using nitric wastewater as substrate3/ h, temperature Under conditions of 26-30 DEG C, degradation reaction is carried out in aeration tank, realizes the fast degradation of ammonia nitrogen and total nitrogen in nitric wastewater；Institute Stating carrier is diatomite or bentonite.
5. application as claimed in claim 4, it is characterised in that the wet thallus is 1:50-200, the bacterium with carrier quality ratio The dosage of agent is calculated as 1.0-5.0g/L with nitric wastewater volume.
6. application as claimed in claim 4, it is characterised in that the wet thallus is prepared as follows:
(1) arthrobacterium B2 is seeded to LB solid medium, in 37 DEG C of culture 26h, obtains thallus inclined-plane；The LB solid culture Base final concentration composition are as follows: tryptone 10g/L, yeast extract 5g/L, sodium chloride 10g/L, agar 20g/L, solvent are water, pH 7.0；
(2) it is seeded in LB liquid medium from one ring thallus of thallus inclined-plane picking, 37 DEG C of culture 28h, forms seed liquor；LB liquid Body culture medium final concentration composition are as follows: tryptone 10g/L, yeast extract 5g/L, sodium chloride 10g/L, solvent are water, pH 7.0；
(3) seed liquor is transferred to the fermentor equipped with fermentation medium with the inoculum concentration of volumetric concentration 3% to cultivate, volume dress Liquid measure 70%, fermentation temperature are 37 ± 1 DEG C, stirring rate 200rpm, ventilatory capacity 0.9-1.1V/vm, and oxyty is 80-90%, culture to for 24 hours when, fermentation liquid is centrifuged, harvest wet thallus；The fermentation medium final concentration composition are as follows: tryptose Peptone 10g/L, yeast extract 5g/L, sodium chloride 10g/L, solvent are water, pH 7.0.
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CN107245463B (en) * 2017-06-22 2020-09-04 温州大学 Heterotrophic nitrification-aerobic denitrification arthrobacter and application thereof
CN111099743B (en) * 2018-10-26 2022-06-07 中国石油化工股份有限公司 Composition for enhancing denitrification and denitrification of wastewater and denitrification method thereof
CN110195027B (en) * 2019-03-08 2022-03-01 大连理工大学 Tobacco arthrobacter ZL-1 and preparation method and application of compost low-temperature fermentation microbial inoculum thereof
CN110184223B (en) * 2019-06-17 2020-11-24 华中农业大学 Arthrobacter calsium capable of removing ammonia nitrogen in culture sewage and application thereof
CN112760260A (en) * 2021-01-26 2021-05-07 武汉市农业科学院 Sporobacillus partialis and application thereof
CN113979548B (en) * 2021-11-09 2023-09-08 上海水源地建设发展有限公司 Use of Arthrobacter TY2 for water treatment and microbial inoculum for water treatment

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