CN105441489B - A method of synthesizing selenium and bismuth selenide nano material using lysine bacillus - Google Patents
A method of synthesizing selenium and bismuth selenide nano material using lysine bacillus Download PDFInfo
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- 239000002086 nanomaterial Substances 0.000 title claims abstract description 34
- FBGGJHZVZAAUKJ-UHFFFAOYSA-N bismuth selenide Chemical compound [Se-2].[Se-2].[Se-2].[Bi+3].[Bi+3] FBGGJHZVZAAUKJ-UHFFFAOYSA-N 0.000 title claims abstract description 22
- 238000000034 method Methods 0.000 title claims abstract description 19
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 title claims abstract description 10
- 239000004472 Lysine Substances 0.000 title claims abstract description 10
- 241000193830 Bacillus <bacterium> Species 0.000 title claims abstract description 9
- 230000002194 synthesizing effect Effects 0.000 title claims abstract description 6
- BUGBHKTXTAQXES-UHFFFAOYSA-N Selenium Chemical compound [Se] BUGBHKTXTAQXES-UHFFFAOYSA-N 0.000 title abstract description 62
- 239000011669 selenium Substances 0.000 title abstract description 58
- 229910052711 selenium Inorganic materials 0.000 title abstract description 56
- BVTBRVFYZUCAKH-UHFFFAOYSA-L disodium selenite Chemical class [Na+].[Na+].[O-][Se]([O-])=O BVTBRVFYZUCAKH-UHFFFAOYSA-L 0.000 claims abstract description 29
- 235000015921 sodium selenite Nutrition 0.000 claims abstract description 29
- 230000001954 sterilising effect Effects 0.000 claims abstract description 4
- 238000004659 sterilization and disinfection Methods 0.000 claims abstract description 4
- 239000007788 liquid Substances 0.000 claims description 20
- 229960001471 sodium selenite Drugs 0.000 claims description 20
- 239000011781 sodium selenite Substances 0.000 claims description 20
- 239000006228 supernatant Substances 0.000 claims description 11
- 241000894006 Bacteria Species 0.000 claims description 10
- 238000005119 centrifugation Methods 0.000 claims description 7
- 239000000463 material Substances 0.000 claims description 7
- 239000002054 inoculum Substances 0.000 claims description 6
- 239000002244 precipitate Substances 0.000 claims description 4
- 238000004321 preservation Methods 0.000 claims description 3
- 229910052797 bismuth Inorganic materials 0.000 claims description 2
- JCXGWMGPZLAOME-UHFFFAOYSA-N bismuth atom Chemical compound [Bi] JCXGWMGPZLAOME-UHFFFAOYSA-N 0.000 claims description 2
- 238000001035 drying Methods 0.000 claims description 2
- 238000009629 microbiological culture Methods 0.000 claims description 2
- 239000013049 sediment Substances 0.000 claims description 2
- 238000011282 treatment Methods 0.000 claims description 2
- FGIUAXJPYTZDNR-UHFFFAOYSA-N potassium nitrate Chemical compound [K+].[O-][N+]([O-])=O FGIUAXJPYTZDNR-UHFFFAOYSA-N 0.000 claims 1
- 239000000243 solution Substances 0.000 abstract description 18
- 230000015572 biosynthetic process Effects 0.000 abstract description 16
- 238000006243 chemical reaction Methods 0.000 abstract description 10
- 239000002077 nanosphere Substances 0.000 abstract description 7
- RXPAJWPEYBDXOG-UHFFFAOYSA-N hydron;methyl 4-methoxypyridine-2-carboxylate;chloride Chemical compound Cl.COC(=O)C1=CC(OC)=CC=N1 RXPAJWPEYBDXOG-UHFFFAOYSA-N 0.000 abstract description 5
- 238000002360 preparation method Methods 0.000 abstract description 4
- 238000001179 sorption measurement Methods 0.000 abstract description 3
- 239000007864 aqueous solution Substances 0.000 abstract description 2
- 229910001385 heavy metal Inorganic materials 0.000 abstract description 2
- 239000000084 colloidal system Substances 0.000 abstract 1
- 229940091258 selenium supplement Drugs 0.000 description 52
- 229940082569 selenite Drugs 0.000 description 14
- 238000003786 synthesis reaction Methods 0.000 description 13
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 12
- 239000002609 medium Substances 0.000 description 12
- 239000002105 nanoparticle Substances 0.000 description 12
- 230000001580 bacterial effect Effects 0.000 description 7
- 239000007787 solid Substances 0.000 description 7
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 238000002835 absorbance Methods 0.000 description 6
- 244000005700 microbiome Species 0.000 description 6
- 239000000047 product Substances 0.000 description 6
- MCAHWIHFGHIESP-UHFFFAOYSA-L selenite(2-) Chemical compound [O-][Se]([O-])=O MCAHWIHFGHIESP-UHFFFAOYSA-L 0.000 description 6
- 239000011780 sodium chloride Substances 0.000 description 6
- 239000000843 powder Substances 0.000 description 5
- 239000000126 substance Substances 0.000 description 5
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 4
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 4
- 239000001888 Peptone Substances 0.000 description 4
- 108010080698 Peptones Proteins 0.000 description 4
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 4
- 230000003834 intracellular effect Effects 0.000 description 4
- 239000002070 nanowire Substances 0.000 description 4
- 235000019319 peptone Nutrition 0.000 description 4
- 239000011734 sodium Substances 0.000 description 4
- 229910052979 sodium sulfide Inorganic materials 0.000 description 4
- GRVFOGOEDUUMBP-UHFFFAOYSA-N sodium sulfide (anhydrous) Chemical compound [Na+].[Na+].[S-2] GRVFOGOEDUUMBP-UHFFFAOYSA-N 0.000 description 4
- PMYDPQQPEAYXKD-UHFFFAOYSA-N 3-hydroxy-n-naphthalen-2-ylnaphthalene-2-carboxamide Chemical compound C1=CC=CC2=CC(NC(=O)C3=CC4=CC=CC=C4C=C3O)=CC=C21 PMYDPQQPEAYXKD-UHFFFAOYSA-N 0.000 description 3
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 3
- 241001032013 Lysinibacillus sp. ZYM-1 Species 0.000 description 3
- 238000010521 absorption reaction Methods 0.000 description 3
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- 239000004615 ingredient Substances 0.000 description 3
- 230000008569 process Effects 0.000 description 3
- 229960001881 sodium selenate Drugs 0.000 description 3
- 239000011655 sodium selenate Substances 0.000 description 3
- 235000018716 sodium selenate Nutrition 0.000 description 3
- 238000000870 ultraviolet spectroscopy Methods 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 2
- 241001464944 Thauera selenatis Species 0.000 description 2
- 238000001784 detoxification Methods 0.000 description 2
- 239000012634 fragment Substances 0.000 description 2
- 230000001788 irregular Effects 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- QEBDLIWRLCPLCY-UHFFFAOYSA-N selanylidenebismuth Chemical class [Bi]=[Se] QEBDLIWRLCPLCY-UHFFFAOYSA-N 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- 230000001988 toxicity Effects 0.000 description 2
- 231100000419 toxicity Toxicity 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- QGLWBTPVKHMVHM-KTKRTIGZSA-N (z)-octadec-9-en-1-amine Chemical compound CCCCCCCC\C=C/CCCCCCCCN QGLWBTPVKHMVHM-KTKRTIGZSA-N 0.000 description 1
- 108020004465 16S ribosomal RNA Proteins 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- KZBUYRJDOAKODT-UHFFFAOYSA-N Chlorine Chemical compound ClCl KZBUYRJDOAKODT-UHFFFAOYSA-N 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- 206010054949 Metaplasia Diseases 0.000 description 1
- 229940096437 Protein S Drugs 0.000 description 1
- 102000029301 Protein S Human genes 0.000 description 1
- 108010066124 Protein S Proteins 0.000 description 1
- 241001459308 Pseudomonas alcaliphila Species 0.000 description 1
- 229910018143 SeO3 Inorganic materials 0.000 description 1
- 241000017150 Shewanella sp. HN-41 Species 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 239000003125 aqueous solvent Substances 0.000 description 1
- 238000004630 atomic force microscopy Methods 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000011953 bioanalysis Methods 0.000 description 1
- 238000004166 bioassay Methods 0.000 description 1
- 230000008033 biological extinction Effects 0.000 description 1
- 238000006555 catalytic reaction Methods 0.000 description 1
- 238000012512 characterization method Methods 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 239000003638 chemical reducing agent Substances 0.000 description 1
- 239000000460 chlorine Substances 0.000 description 1
- 229910052801 chlorine Inorganic materials 0.000 description 1
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- 238000010586 diagram Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- PPNKDDZCLDMRHS-UHFFFAOYSA-N dinitrooxybismuthanyl nitrate Chemical compound [Bi+3].[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O PPNKDDZCLDMRHS-UHFFFAOYSA-N 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
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- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
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- 231100000572 poisoning Toxicity 0.000 description 1
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- 230000035484 reaction time Effects 0.000 description 1
- 238000005057 refrigeration Methods 0.000 description 1
- MCAHWIHFGHIESP-UHFFFAOYSA-N selenous acid Chemical class O[Se](O)=O MCAHWIHFGHIESP-UHFFFAOYSA-N 0.000 description 1
- 239000004065 semiconductor Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
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- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The present invention relates to a kind of methods synthesizing selenium and bismuth selenide nano material using lysine bacillus ZYM-1.0.5-25mM sodium selenites are added in the LB culture mediums through 121 DEG C of high-temperature sterilizations, is inoculated with ZYM-1, cultivates 24-48h under 25-40 DEG C, pH 5-9,50-300rpm, obtain red colloid selenium nano material；0.5-25mM sodium selenites and 0.1-5mM bismuth nitrate solutions are added in the LB culture mediums through 121 DEG C of high-temperature sterilizations, is inoculated with ZYM-1, cultivates 24-48h under 25-40 DEG C, pH 5-9,50-300rpm, obtain black bismuth selenide nano material.Reaction carries out in aqueous solution, without harsh conditions such as high temperature and pressure, is easily enlarged culture.The selenium nanosphere of preparation can be used for heavy metal adsorption removal, and the bismuth selenide nano material of preparation has no the report of biosynthesis at present.
Description
Technical field
The invention belongs to environment field of nano biotechnology, it is related to a kind of utilizing lysine bacillus synthesis selenium and selenizing
The method of bismuth nano material.
Background technology
Selenium belongs to the 6th major element, is trace element necessary to a kind of biological existence, but the selenium of high concentration has life
Object toxicity and environmental pollution can be caused.Selenium in nature mainly exists with four kinds of valence states (- 2,0 ,+4 and+6).Wherein, sub- selenium
Hydrochlorate (SeO3 2-) soluble easily in water, there is stronger bio-toxicity compared with other forms.Researcher has found to be widespread in nature
Multiple-microorganism selenite can be restored by detoxification, generate toxicity it is smaller, the poor red simple substance of aqueous solvent
Selenium nano particle.Wang GJ etc. are in document (BMC Microbiology.2014.14.204-217), Lampis S etc. in document
(Microbial cell factories.2014.13.35-39) and Oremland RS etc. are in document
(Exremophiles.2009.13.695-705) specified microorganisms are reported in be reduced to elemental selenium by sodium selenite
Nano particle.But for common microorganism, sodium selenite belongs to a kind of toxicant, and a small amount of sodium selenite may
It may result in the complete inhibition of microorganism growth.
Selenium nano material has excellent unilateral conduction and good photoelectric characteristic, is widely used to rectifier and light
The manufacture of battery, photoelectric characteristic also have been used for making toner cartridge in Xeroxing.Such as Abdelouas A are in document
Pigment C3 is reported in (Chemistry of materials.2000.12.1510-1512) prepares selenium for restoring sodium selenate
Nano wire.Xia YN etc. are in document (Journal of the American Chemical
Society.2000.122.12582-12583 it is reported in) and tripartite's selenium nanowires is prepared by liquid phase ageing method.But these
Method needs the presoma using expensive reducing agent or complexity, or needs the reaction time grown very much.Therefore, exploitation reaction item
Part is mild, and the friendly new process of building-up process green has important application value.Since microorganism is to selenate and selenous acid
Salt has detoxification, can realize selenium nano material using microorganism as " biological factory " of synthesis selenium nano material
Green preparation process.Such as Debieux CM are in document (Proceedings of the National Academy of
Sciences.2011.108.13480-13485 culture bacterium T.selenatis under anaerobic is reported in),
T.selenatis can synthesize the selenium nanosphere of 150-200nm sizes, and be transported to born of the same parents after intracellular assembling by Protein S efA
Outside.Hur HG et al. are reported in document (Chemosphere.2007.68.1898-1905) under anaerobic condition
Shewanella sp.HN-41 can synthesize selenium nanosphere, and by subsequently adding the dimethyl sulfoxide (DMSO) of various concentration by selenium nanometer
Ball is converted into nano wire.Li DP are in document (Colliods and Surface B:Biointerfaces.2011.88.196-
201) using Pseudomonas alcaliphila in aerobic synthesis selenium nanosphere and selenium nanosphere/selenium nanometer rods in
Mixture.However, currently with pure bioprocess, the selenium nanowires for not adding any organic solvent synthesis pure phase do not appear in the newspapers
Road.
Bismuth selenide belongs to the inorganic semiconductor material of rhombohedron crystallographic system, and energy gap is relatively narrow, and the thermoelectricity under room temperature turns
Change it is efficient, therefore heat pump, refrigeration, biosensor etc. tool have been widely used.Qian YT etc. are in document (Solid
State Communications.2008.147.36-40) in report in the mixed solvent in oleyl amine and ethylene glycol, use selenium powder
It is the hollow Bi of Material synthesis with bismuth nitrate2Se3The diameter of nano spherical shell structure, single sphere is about 600nm, and shell thickness is only
For 40nm.However, the Bi reported at present2Se3Synthetic method be chemical method or Physical, there has been no bioanalysis synthesis
The report of bismuth selenide.
Retrieval both at home and abroad in relation to bismuth selenide synthesis in terms of document and patent results show before this invention is filed, and also
The report in terms of the biosynthesis of bismuth selenide under aerobic condition is not found.
Invention content
It is restored using lysine bacillus Lysinibacillus sp.ZYM-1 the purpose of the present invention is to provide a kind of
The method that selenite synthesizes selenium nano material and bismuth selenide nano material, selenium nano material and bismuth selenide nano material are in light
Catalysis, heavy metal adsorption and thermoelectric conversion field have significant application value.
Lysine bacillus Lysinibacillus sp.ZYM-1 are inventors from Liaoning Province Panjin City Liaodong Wan coastal waters
It is isolated in bed mud sample, it is preserved in China Microbial Culture Preservation Commission's common micro-organisms center (address：Court of Beijing
No. 3 Institute of Microorganism, Academia Sinica of institute of positive area's North Star West Road 1), the deposit date is on October 22nd, 2015, culture presevation
Number is CGMCC 1.15346, and the Genbank numbers of logging in of 16s rRNA gene orders are KT263530.
The technical scheme is that：
Lysine bacillus Lysinibacillus sp.ZYM-1, it is characterised in that：Lysine bacillus ZYM-1
There are two types of training methods：
One is the glycerine bacterium solution for the bacterial strain ZYM-1 that -80 DEG C preserve is melted at 4 DEG C, it is inoculated in 20ml and contains 0.5-
It is cultivated in the LB liquid medium of 25mM (mM) sodium selenite；The LB liquid medium formula is：Sodium chloride 10g/L,
Peptone 10g/L, yeast powder 5g/L；LB liquid medium all needs to carry out 121 DEG C of sterilization treatment 20min using preceding.
Another kind is to use LB solid mediums, and the ZYM-1 bacterium solution tablets after above-mentioned culture are coated with culture；The LB is solid
Body culture medium is that the agar powder that mass fraction is 2% is added in LB liquid medium.
A method of synthesizing selenium nano material using ZYM-1：ZYM-1 is inoculated in 20ml sodium selenites containing 0.5-25mM
LB liquid medium in cultivate 8-12 hours；ZYM-1 bacterium solutions after culture are transferred to new with the inoculum concentration of 1%-5%
In 100ml LB liquid mediums, it is added the sodium selenite of 0.5-25mM, 25-40 DEG C, cultivates 24- under pH 5-9,50-300rpm
ZYM-1 bacterium solutions after culture are placed at 121 DEG C and handle 20min by 48h so that and the nanometer selenium of intracellular is released from thalline,
2000-3000rpm is centrifuged, the red supernatant of separating thallus fragment and wrapping biological molecule, by red supernatant 12000-
20000rpm is centrifuged, and takes centrifugation product, red selenium nano material is obtained after dry.
A method of synthesizing bismuth selenide nano material using ZYM-1：It is sub- containing 0.5-25mM that ZYM-1 is inoculated in 20ml
It is cultivated 8-12 hours in the LB liquid medium of sodium selenate；ZYM-1 bacterium solutions after culture are transferred to newly with the inoculum concentration of 1%-5%
100ml LB liquid mediums in, while the sodium selenite of 0.5-25mM and the bismuth nitrate of 0.1-5mM is added, 25-40 DEG C, pH
24-48h is cultivated under 5-9,50-300rpm, product is collected by centrifugation in subsequent 2000-3000rpm, and removal precipitates to obtain black supernatant,
Black supernatant is centrifuged with 12000-20000rpm again and obtains sediment, black bismuth selenide nanometer material can be obtained after drying precipitate
Material.Transmission electron microscope and atomic force microscope show that bismuth selenide is three-dimensional ball shape structure, between size is about 20-40nm.
The beneficial effects of the invention are as follows：The present invention closes the larger selenite of selenium-containing wastewater Poisoning in mild condition
At selenium nano material and bismuth selenide nano material, method environment green is friendly, and reaction carries out in aqueous solution, is not necessarily to high temperature and pressure
Etc. harsh conditions, it is easy to operate, be easily enlarged culture.The appearance of nano material of preparation is controllable, and selenium nanosphere can be used for a huge sum of money
The Adsorption of category, bismuth selenide nano material have not yet to see the report of biosynthesis.
Description of the drawings
Fig. 1 is the digital photograph of ZYM-1 reduction selenite synthesis selenium nano particle under tablet coating condition of culture.
Fig. 2 is the variation tendency of selenium also commercial weight under different concentration of sodium selenite.
Fig. 3 is the variation tendency of selenium also commercial weight under different pH.
Fig. 4 is the ultraviolet full wavelength scanner figure that selenium nanoparticle system is synthesized under different concentration of sodium selenite.
Fig. 5 is the topography scan electron microscope that selenium nano particle is synthesized under different concentration of sodium selenite；
(a) it is that the linear selenium nano material of nanometer is generated after 1mM sodium selenites restore；
(b) it is the selenium nano material that cube or spherical shape are generated after 2.5mM sodium selenites restore；
(c) it is the selenium nano material that nanometer spherical is generated after 5mM sodium selenites restore；
(d) it is to generate irregular selenium nano material after 25mM sodium selenites restore.
Fig. 6 is the representative transmission electron microscope picture of ZYM-1 synthesis bismuth selenides.
Fig. 7 is the atomic force microscopy diagram spectrum of ZYM-1 synthesis bismuth selenides.
Specific implementation mode
Embodiment 1：Tablet is coated with bacterial strain ZYM-1 reduction selenite under condition of culture and synthesizes selenium nano particle
Reaction system：It is small 8-12 will to be cultivated in LB liquid mediums of the 20ml containing 0.5-25mM (mM) sodium selenite
When after ZYM-1 bacterium solutions dilution different multiples after, with apply stick be uniformly coated on the LB solid mediums containing 1mM sodium selenites
On.36h is cultivated in 30 DEG C of constant incubators.The color of bacterium colony becomes red from faint yellow, discloses reduzate selenium nanometer
The tablet photo of the generation of particle, culture is as shown in Figure 1.
Embodiment 2：Reducing degrees of the bacterial strain ZYM-1 to selenite under different concentration of sodium selenite
0.5,1,2.5,5,10,25mM sodium selenite, ZYM-1 bacterium solutions is added in the LB liquid medium of 20mL respectively
Inoculum concentration is 1%, and cultivation temperature is 30 DEG C, and pH 5.5,100rpm cultivate 48h；The LB liquid medium ingredient is：Chlorine
Change sodium 10g/L, peptone 10g/L, yeast powder 5g/L.The elemental selenium nano particle Na of generation2S is aoxidized, and is calculated selenium simple substance and is contained
Amount, the specific method is as follows：
1. preparing the Na of 1M2S solution is added suitable NaOH solids and inhibits Na2S's hydrolyzes certainly.The NaCl for configuring 1M is molten
Liquid.It is separately added into different amounts of commercialization selenium powder in 10ml test tubes, 5mL Na are added2S solution fully vibrates dissolving so that mark
The concentration of selenium is respectively 0.5,1,2,3,4,5mM in quasi- liquid.Using ultraviolet-visible spectrophotometer at 500nm bioassay standard
Absorbance value (the OD of solution500nm), draw standard curve.Wherein ordinate is absorbance value OD500nm, abscissa is corresponding
Selenium concentration of standard solution.The corresponding relationship of the two is obtained using linear fit equation.
2. the sample containing thalline and intracellular selenium nano particle is centrifuged 20min by 12000rpm, with separation of the supernatant.
Using 1M NaCl washing precipitations, selenite radical remaining in solution is removed, then, by remaining solids with 5mL 1M's
Na2S fully vibrates dissolving, measures the absorbance value of solution at 500nm using ultraviolet-visible spectrophotometer.Utilize step 1
The standard curve linear fit obtained is as a result, the OD that can be surveyed from sample500nmCalculate the content of corresponding selenium element.When
Measured absorbance OD500nmIt when the absorbance upper limit beyond standard curve, needs sample being diluted, ensures the extinction measured
Degree is in the range of standard curve is included.Experimental result is as shown in Figure 2.When starting selenite radical content be 0.5,1,
When 2.5 and 5mM, conversion ratios of the bacterial strain ZYM-1 in 48h can reach 100%, and when starting selenite radical concentration rises to
When 10mM, conversion ratio is reduced to 53%, when a concentration of 25mM of starting selenite radical, conversion ratio 31.7%.
Embodiment 3：Different pH restore bacterial strain the influence of selenite radical.
It is utilized respectively HCl or NaOH and the pH of LB liquid medium is adjusted to 4,5,6,7,8,9,10,11, and ensure to cultivate
System total volume is 20mL.Bacterium solution ZYM-1 inoculum concentrations are 1%, a concentration of 5mM of selenite radical, and cultivation temperature is 30 DEG C, 200rpm
Cultivate 36h；The LB liquid medium ingredient is：Sodium chloride 10g/L, peptone 10g/L, yeast powder 5g/L.Utilize Na2S oxygen
Metaplasia at elemental selenium nano particle, specific measurement method is as follows：
Sample containing thalline and intracellular selenium nano particle is centrifuged into 10min by 10000rpm, with separation of the supernatant.Profit
It is rinsed 2 times with 1M NaCl solutions, selenite radical remaining in solution is removed, then, by remaining solids with 5mL 1M's
Na2S fully vibrates dissolving, measures the absorbance value of solution at 500nm using ultraviolet-visible spectrophotometer.According to standard
Curve calculates the amount of the elemental selenium in solution.Experimental result is as shown in Figure 3.Bacterial strain ZYM-1 can be right in the range of pH is 5-9
The selenite radical of 5mM is effectively restored, and reduction rate is respectively 88%, 98%, 99%, 95% and 93%.
Embodiment 4：Influence of the difference starting concentration of sodium selenite to synthesis selenium appearance of nano material.
The thalline comprising selenium nano particle that the reaction of embodiment 2 is obtained passes through 121 DEG C of damp and hot high-temperature process 20min, place
Mixture after reason centrifuges 5min under conditions of 3000rpm, the red supernatant of separating thallus fragment and wrapping biological molecule,
It then takes supernatant to centrifuge 20min at 12000rpm, obtains centrifugation product, product carries out after washing is dry being the selenium obtained
Nano material.The selenium nano particle of acquisition is resuspended in distilled water, visible region full wavelength scanner is carried out, as a result such as Fig. 4 institutes
Show.Fig. 4 show 0.5-25mM sodium selenite system reaction products the sections 450-800nm absorption spectrogram.It can be with from figure
Find out that the reacted product of 1mM and 5mM has maximum absorption peak at 580nm and 592nm respectively.The reaction of other concentration is produced
There is no apparent absorption peaks in the region for object.
Scanning electron microscope characterization is carried out to the reduzate comprising 1mM, 2.5mM, 5mM, the sodium selenite of 25mM, as a result such as
Shown in Fig. 5.As can be seen from the figure apparent pattern evolutionary process.Fig. 5 a show to be generated after the sodium selenite of 1mM restores
Selenium nano material, pattern is presented as uniform nanometer line morphology, the length of 5-10 μm, width 50-100nm.When dense
When degree continues to increase to 2.5mM, the pattern of selenium nano material is presented as based on cube, is partly the state of ball-type, such as Fig. 5 b
It is shown.When concentration increases to 5mM, selenium nano material substantially completely evolves into nanosphere, and diameter is in 300nm or so, such as Fig. 5 c institutes
Show.When concentration continues to increase to 25mM, selenium nano material evolves into irregular pattern, as fig 5d.
Embodiment 5：Bacterial strain ZYM-1 is used for the synthesis of bismuth selenide nano material
0.5,1,2.5,5,10,25mM sodium selenite and corresponding Asia is added in the LB liquid medium of 20mL respectively
The bismuth nitrate of sodium selenate substance withdrawl syndrome 1/10.If the bismuth nitrate of 0.05mM is added in the sodium selenite system of 0.5mM, 1mM's
The bismuth nitrate of 0.1mM is added in sodium selenite system, and other systems are similar.Inoculum concentration is 1%, and cultivation temperature is 30 DEG C, and pH is
5.5,300rpm cultures are for 24 hours；The LB liquid medium ingredient is：Sodium chloride 10g/L, peptone 10g/L, yeast powder 5g/L.
3000rpm centrifuges 5min and collects the supernatant after centrifugation, then obtains product with 12000rpm centrifugations 20min, utilizes ultrapure washing
It washs 3 times, can be obtained bismuth selenide nano material after dry 6h in 80 DEG C of baking ovens.Representative bismuth selenide transmission electron microscope and original
Sub- force microscope figure such as Fig. 6, shown in Fig. 7.It can be seen from the figure that the bismuth selenide of synthesis is mainly ball-type, size is about 20nm.
The above, only this patent preferably specific implementation mode, but protection scope of the present invention is not limited to that,
Even if any one skilled in the art in the range of present disclosure, technique according to the invention method and its
Inventive concept is subject to equivalent substitution or change, should all be included within the scope of the present invention.
Claims (1)
1. a kind of method synthesizing bismuth selenide nano material using lysine bacillus ZYM-1, it is characterised in that：By ZYM-1
It is inoculated in the LB liquid medium of the 20ml sodium selenites of mM containing 0.5-25 8-12 hours；By the ZYM-1 bacterium solutions after culture with
The inoculum concentration of 1%-5% is transferred in 100ml LB liquid mediums, and the sodium selenite of 0.5-25 mM and the nitre of 0.1-5 mM is added
Sour bismuth, cultivates 24-48h under pH 5-9,50-300 rpm by 25-40 DEG C；2000-3000 rpm centrifugations, removal precipitates black
Black supernatant 12000-20000 rpm centrifugations are obtained sediment, bismuth selenide nanometer material are obtained after drying precipitate by color supernatant
Material；The LB liquid medium is using preceding needing 121 DEG C of sterilization treatments 20 minutes；The lysine bacillusLysinibacillusSp.ZYM-1 is preserved in China Microbial Culture Preservation Commission's common micro-organisms center, preservation date
For on October 22nd, 2015, culture presevation number was CGMCC 1.15346.
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