CN106916765B - A method of heavy metal in waste water zinc is adsorbed using penicillium janthinellum - Google Patents
A method of heavy metal in waste water zinc is adsorbed using penicillium janthinellum Download PDFInfo
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- C02F3/00—Biological treatment of water, waste water, or sewage
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- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
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- C02F2101/00—Nature of the contaminant
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Abstract
A method of heavy metal in waste water zinc being adsorbed using penicillium janthinellum, belongs to the applied technical field of microorganism fungus kind.The penicillium janthinellum (Penicillium janthinellum) BC109-2, it is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, abbreviation CGMCC, deposit number is CGMCC No.13561.Step of the present invention are as follows: one, screened from soil using dilution spread flat band method and to separate and purify to obtain heavy metal zinc resistant strain；Two, the heavy metal zinc resistant strain for obtaining screening is added in the fluid nutrient medium containing heavy metal zinc, carries out adsorption conditions optimization；Three, under optimal adsorption conditions, which is added in the waste water containing heavy metal zinc, that is, is completed；Penicillium janthinellum BC109-2 has height endurability to heavy metal zinc.It is an advantage of the invention that adsorption effect is good, easy to operate, low in cost, to environment, there is no secondary pollutions.
Description
Technical field
The present invention relates to a kind of methods using penicillium janthinellum absorption heavy metal in waste water zinc, belong to microorganism fungus kind
Applied technical field.The penicillium janthinellum that more specifically, the present invention relates to a kind of for adsorbing zinc in waste water (Penicillium janthinellum) BC109-2。
Background technique
Since the 1930s, as industrial or agricultural rapidly develops, the application of heavy metal is also more and more extensive, therewith
The pollution problem come also more highlights.Heavy metal pollution is mostly derived from the industrial wastewater and biochemistry of the industries such as mining, smelting, plating
The random discharge of sewage.Currently, since exploitation of mineral resources, sewage irrigation etc. cause China by the arable land of the heavy metal pollutions such as Zn, Cd
Nearly 20,000,000 hm2, account for about the 1/5 of total area under cultivation.Zinc is the important essential trace element of one of Animal nutrition, but it is dense
High risks can be caused to biology in the case where spending height, acceptable maximum in drinking water is suggested by the World Health Organization (WHO)
Zn2+Concentration is 5 mg/L.Zinc cannot be biodegradable, and mutually converting between various forms can only occur, and can pass through food
The enrichment of object chain, causes human health risk.In order to be repaired to the water body of heavy metal pollution, currently used restorative procedure packet
Include physics and chemical method.And the physical methods higher cost such as traditional filtering, ion exchange, there are significant limitation, because
The methods of this following chemical method, electrolysis method are used widely, but chemical method has secondary pollution, electrolysis method
Then it is only applicable to the recycling of precious metal.With the development of modern biotechnology, it is biological prosthetic as it is a kind of environmental protection, low cost,
Efficient biological technique method becomes research hotspot in recent years.
Microorganism can be applied to the reparation of heavy metal-polluted water, has numerous researchs and is directed toward microbe absorption counterweight
The repair of metal reaches the potential suction-operated of heavy metal using them by screening the microorganism of preventing from heavy metal
To the purpose of repairing heavy metal pollution.Report in recent years to Zn2+The bacterium for having suction-operated includes pseudomonad
(Pseudomonas) adsorbance be 88.23 mg/g, streptomyces (Streptomyces) adsorbance be 54 mg/g, gemma bar
Bacterium (Geobacillus thermodenitrificans) adsorbance is 48.26 mg/g etc.；Fungi includes Penicillium notatum
(Penicillium sp.) adsorbance be 2.22 mg/g and sickle-like bacteria (Fusarium sp.) adsorbance is 3.79 mg/g etc.
Deng.By the way that strain is inoculated into shaken cultivation in fluid nutrient medium, by being filtered, washed, drying etc., just become after processes can
The cumbersome and harsh condition of culture of adsorbent, operating procedure makes it be dfficult to apply to industrialized production.
Summary of the invention
Object of the present invention is to for current biological adsorption agent preparation process it is cumbersome and at high cost the problems such as, and provide one
Kind using penicillium janthinellum (Penicillium janthinellum) absorption heavy metal in waste water (Zn) method, this method is green
Colour circle is protected, and is had broad application prospects.
Technical solution of the present invention: a kind of penicillium janthinellum (Penicillium janthinellum) BC109-2,
It is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, abbreviation CGMCC, deposit number CGMCC
No.13561。
A method of heavy metal in waste water zinc being adsorbed using the penicillium janthinellum BC109-2, after purification by 0.1mL
BC109-2 bacteria suspension, control 108Cfu/mL is added in the PDB fluid nutrient medium of 100mL, natural pH condition, and cultivation temperature is
30 DEG C, shake flask culture measures growth curve, after cultivating 10-16h, the culture bacterium solution in growth logarithmic phase is obtained, by this
Bacterium solution be inoculated into containing various concentration heavy metal zinc, difference pH PDB fluid nutrient medium in carry out adsorption conditions optimization.
Adsorbed heavy metal is zinc, and penicillium janthinellum BC109-2 has height endurability to heavy metal zinc.
The microassembly robot bacterial strain BC109-2 forms mycelium pellet in the fluid nutrient medium of PDB containing zinc, and hyphal surface is in fold
Shape.
It is 8 containing Zn in 30 DEG C, pH2+PDB fluid nutrient medium in adsorbance be 217.47 mg/g.
The present invention filters out a kind of microorganism penicillium janthinellum using dilution spread flat band method from soil
(Penicillium janthinellum), there is height endurability to heavy metal zinc, and have preferable adsorption effect to zinc.
The microassembly robot bacterial strain BC109-2 is 3 mmol/L in zinc initial concentration, is expanded in the PDB fluid nutrient medium that pH is 8
Reach logarithmic growth phase when big culture 10-12 h, is then centrifuged for collecting thallus.
The thallus being collected by centrifugation is added in the waste water containing heavy metal zinc, is stirred under conditions of temperature is 10-35 DEG C,
Adsorption time is that 10 h reach saturation, that is, is completed.The wherein mass volume ratio g/mL of thallus weight in wet base and the waste water containing heavy metal zinc
For (2 ~ 4) ︰ 1000.
In 30 DEG C, the PDB fluid nutrient medium that pH is 8,24 h of 150r/min shaking table shaken cultivation, the microassembly robot bacterial strain
BC109-2 is to Zn2+Adsorbance with ZnSO4·7H2The increase of O initial concentration and increase, and increasing degree gradually tends to be flat
It is slow, in Zn2+When initial concentration is 3 mmol/L, adsorbance reaches 217.47 mg/g(attached drawings 4).
Beneficial effects of the present invention: microassembly robot bacterial strain BC109-2 culture is easy, needs not move through and is filtered, washed, dries
It the complicated processes such as does, grind, being sieved and biological adsorption agent is made, the effect for adsorbing heavy metal zinc can achieve 217.47 mg/g, and
Thallus is easy to separate from waste water, environmentally protective, not will cause secondary pollution.
Biological material specimens preservation: the bacterial strain be penicillium janthinellum (Penicillium janthinellum) BC109-
2, full name isPenicillium janthinellumBC109-2, sequence have uploaded in Genbank database, strain sequence
Number be KX891188.Penicillium janthinellum BC109-2 has been preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms
Center, abbreviation CGMCC, address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, deposit number are CGMCC No.13561, are protected
The hiding date is on January 16th, 2017.
Detailed description of the invention
The resistance to zinc of Fig. 1 bacterial strain BC109-2 in the medium.
Fig. 2 is containing Zn2+PDB fluid nutrient medium in the microassembly robot bacterial strain BC109-2 that grows form mycelium pellet.
Fig. 3 shows BC109-2 hyphal surface fold under a scanning electron microscope.
Fig. 4 contains Zn2+Waste strength and BC109-2 are to Zn2+The relationship of adsorbance.
Specific embodiment
Below with specific example come the technical solution that the present invention will be described in detail.
Screening and culture penicillium janthinellum (Penicillium janthinellum) culture medium used in BC109-2 is such as
Under:
1) LB culture medium (1000mL): tryptone 10.0g；Yeast extract 5.0g；NaCl 10.0g；Agar powder
Agar is not added in 10.0 ~ 15.0g(fluid nutrient medium), pH 7.0.
2) PDA culture medium: 200 g of potato, 20 g of glucose, agar 15-20 g, ultrapure water 1000 mL, natural pH, 1
×10520 min of Pa sterilizing.
3) PDA of agar PDB culture medium: is not added.
Embodiment 1: the screening and identification of microassembly robot bacterial strain
1) screening of microassembly robot bacterial strain BC109-2
Soil sample is acquired near discarded Tong Xin electroplate factory, sampling depth is 15 ~ 20cm.Pedotheque 5g is weighed, is added to
In conical flask containing 45mL sterile water, shaking table room temperature shakes 10min, and 1mL sludge suspension is then taken to be added to sterile water containing 9mL
Test tube in, mix well, be diluted to 10 step by step-7Sludge suspension.10 are taken respectively-2、10-3、10-4、10-5、10-6、10-7It is dilute
It releases liquid 1mL and is coated on Zn on superclean bench2+Concentration is on the LB solid medium of 30mg/L, 30 in constant incubator
48h is cultivated under conditions of DEG C.After bacterium colony is grown, the visibly different bacterial strain of colony characteristics is picked them separately, is crossed using trilinear method
Transposing is to containing Zn2+LB plate on continue to cultivate, fungi is then in transposing to PDA plate, obtains after bacterial strain by separating for several times
Purifying, finally obtains one plant in the Zn containing 2100mg/L2+PDA plate on normal growth bacterial strain, number BC109-2.
2) identification of BC109-2
(a) extracting genome DNA
It is operated according to the Ezup pillar fungal genomic DNA extraction agent box (SK8259) of Shanghai Sangon Biotech Company.
(b) PCR amplification
A primer 1:5 ' TCCGTAGGTG AACCTGCGG mono- 3 ',
Primer 2: 5 ' one TCCTCCGCTT ATTGATATGC mono- 3 ',
Reaction system:
PCR cycle condition: 94 DEG C of the first stage, 4min；94 DEG C of second stage (30 circulations), 45s, 55 DEG C, 45s, 72
℃,1min；72 DEG C of phase III, 10min.
DNA purpose band, PCR product PCR primer direct Sequencing needed for PCR product electrophoretic band is cut.DNA sequencing committee
Shanghai Sangon Biotech Company is ask to carry out.
Obtained ITS sequence overall length 552bp is sequenced, sequence is as shown in SEQ ID NO:1.
The analysis of Blast Homology search is carried out in Genbank database, it is found that the bacterium reaches with microassembly robot homology
100%, combining form identification, the bacterial strain be penicillium janthinellum (Penicillium janthinellum), full name isPenicillium janthinellumBC109-2, sequence have uploaded in Genbank database, and strain sequence number is
KX891188。
Embodiment 2: the influence that zinc components grow penicillium janthinellum BC109-2
Bacterial strain is picked them separately in fresh PDA liquid medium, 30 DEG C, 150 r/min shaken cultivations, to bacterium solution OD600
When reaching 0.2 ~ 0.4, takes each 2mL of bacterium solution to be placed in 250mL triangular flask respectively, add PDA liquid medium to 50mL.In 30 DEG C,
150r/min continues shaken cultivation, to OD600Value reaches 0.2 or so, and addition final concentration is respectively 1,2,5 mmol/L
ZnSO4·7H2O solution, while control experiment is done, three parallel repetitions are arranged in each concentration.Continue will above 12 triangular flasks in
30 DEG C, cultivate 2d in the shaken cultivation case of 150 r/min, (0,6,12,18,24,30,36,42,48 h) takes at regular intervals
L mL bacterium solution measures OD600Value.Three repetition summations of each sample are averaged the OD as the sample600Value.
Thallus is free of Zn in the medium2+When growing state it is best, and culture for 24 hours left and right biomass reach highest.With
Zn in culture medium2+The increase of concentration, thalli growth obviously slow down, and biomass reduces, and illustrate Zn2+Have one to the growth of bacterial strain
Fixed inhibiting effect, but the bacterial strain is in Zn2+It remains to grow in the culture medium that concentration is 3 mmol/L, shows that bacterial strain BC109-2 has
Certain resistance to zinc, specific visible Figure of description 1.
30 DEG C, pH be 8 contain Zn2+PDB fluid nutrient medium in the microassembly robot bacterial strain BC109-2 that grows form mycelium pellet
(Fig. 2), and hyphal surface fold (Fig. 3) is shown under a scanning electron microscope.
Embodiment 3: bacterial strain is to Zn2+Absorption
100 mL PDA liquid mediums are added in 250 mL triangular flasks, bacterial strain BC109-2 is inoculated into and is contained respectively
The ZnSO of 0,1,2 and 3mmol/L4·7H2In the culture medium of O, the pH value 1 mol/L HCl and 1 mol/L NaOH of culture medium
8 are adjusted to, 30 DEG C is subsequently placed in, cultivates in 150 r/min shaking tables.Three groups of repetitions of experimental setup.After cultivating 24 h, 8000 g from
It is washed three times after 15 min of the heart with 0.9% NaCl aqueous solution.Weighing will be dried under 80 DEG C of constant temperature of cell, then use nitric acid and height
Zn is measured after chloric acid (7 ︰ 3, V/V) micro-wave digestion2+Content.
It is 8 in pH value of solution, and under conditions of reaching saturated adsorption time, thallus is to Zn2+Adsorbance with ZnSO4·
7H2The increase of O initial concentration and increase, and increasing degree gradually tends towards stability, in Zn2+When initial concentration is 3 mmol/L, inhale
Attached amount reaches 217.47 mg/g(Fig. 4).
Sequence table
<210>SEQ ID NO:1
<211> 552
<212> DNA
<213>penicillium janthinellum BC109-2
<400>1
gcttaagttc agcgggtatc cctacctgat ccgaggtcaa cctgagaaag attgaggggg 60
gtcgccggcg ggcgccggcc gggcctacag agcgggtgac gaagccccat acgctcgagg 120
accggacgcg gtgccgccgc tgcctttcgg gcccgccccc cgggagccgg ggggcggggg 180
cccaacacac aagccgtgct tgagggcagc aatgacgctc ggacaggcat gccccccgga 240
ataccagggg gcgcaatgtg cgttcaaaga ctcgatgatt cactgaattc tgcaattcac 300
attacttatc gcatttcgct gcgttcttca tcgatgccgg aaccaagaga tccgttgttg 360
aaagttttaa ctgatttagc taatcgctca gacagcaatc ttcagacagc gttcaggggg 420
ggcttcggcg ggcgcgggcc cgggggcggg tgccccccgg cggccatgac ggcgggcccg 480
ccgaagcaac taggtatgat aaacacgggt gggaggttgg acccagaggg ccctcactcg 540
gtaatgatcc tt 552
Claims (3)
1. it is a kind of using penicillium janthinellum (Penicillium janthinellum) BC109-2 absorption heavy metal in waste water zinc
Method, it is characterised in that:
The penicillium janthinellum (Penicillium janthinellum) BC109-2, it has been preserved in Chinese microorganism strain guarantor
Administration committee's common micro-organisms center, abbreviation CGMCC are hidden, deposit number is CGMCC No.13561；
Step are as follows: by 0.1mL bacterial concentration be after purification 108The BC109-2 bacteria suspension of cfu/mL is added to the PDB liquid of 100mL
In body culture medium, natural pH condition, cultivation temperature is 30 DEG C, and shake flask culture measures growth curve, after cultivating 10-16h,
The culture bacterium solution in growth logarithmic phase is obtained, which is inoculated into the PDB liquid containing various concentration heavy metal zinc, difference pH
It is adsorbed in body culture medium.
2. the method according to claim 1 using penicillium janthinellum BC109-2 absorption heavy metal in waste water zinc, feature
It is that the microassembly robot bacterial strain BC109-2 forms mycelium pellet in the fluid nutrient medium of PDB containing zinc, hyphal surface is in accordion.
3. the method according to claim 1 using penicillium janthinellum BC109-2 absorption heavy metal in waste water zinc, feature
It is that in 30 DEG C, pH be 8 containing Zn2+PDB fluid nutrient medium in thallus to Zn2+Adsorbance with ZnSO4·7H2O is initial
The increase of concentration and increase, in Zn2+When initial concentration is 3 mmol/L, adsorbance is 217.47 mg/g.
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| CN109967040A (en) * | 2017-12-28 | 2019-07-05 | 营口富里泥炭科技有限公司 | A kind of preparation method of novel sludge compound adsorbent |
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| CN115011488B (en) * | 2022-05-31 | 2023-03-24 | 贵州省生物研究所 | Penicillium purpureum C1-GP and application thereof in cultivation of blueberry mycorrhizal seedlings |
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