CN104749273B - Method for detecting azide ions or cyanide ions in water - Google Patents
Method for detecting azide ions or cyanide ions in water Download PDFInfo
- Publication number
- CN104749273B CN104749273B CN201510108903.6A CN201510108903A CN104749273B CN 104749273 B CN104749273 B CN 104749273B CN 201510108903 A CN201510108903 A CN 201510108903A CN 104749273 B CN104749273 B CN 104749273B
- Authority
- CN
- China
- Prior art keywords
- ion
- water
- nitrine
- ions
- butanol
- 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
Links
- -1 azide ions Chemical class 0.000 title claims abstract description 58
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 title claims abstract description 43
- 238000000034 method Methods 0.000 title claims abstract description 27
- XFXPMWWXUTWYJX-UHFFFAOYSA-N Cyanide Chemical compound N#[C-] XFXPMWWXUTWYJX-UHFFFAOYSA-N 0.000 title abstract description 10
- QSDCNOAJMHCYBM-UHFFFAOYSA-N 3,5-dihydroxypentanenitrile Chemical compound OCCC(O)CC#N QSDCNOAJMHCYBM-UHFFFAOYSA-N 0.000 claims abstract description 21
- LRHPLDYGYMQRHN-UHFFFAOYSA-N Butanol Natural products CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 claims abstract description 20
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 43
- 150000002500 ions Chemical class 0.000 claims description 43
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 39
- 150000004820 halides Chemical class 0.000 claims description 36
- 108090000790 Enzymes Proteins 0.000 claims description 30
- 102000004190 Enzymes Human genes 0.000 claims description 30
- 238000001514 detection method Methods 0.000 claims description 30
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 claims description 29
- 239000000243 solution Substances 0.000 claims description 28
- 239000007788 liquid Substances 0.000 claims description 23
- RBACIKXCRWGCBB-UHFFFAOYSA-N 1,2-Epoxybutane Chemical group CCC1CO1 RBACIKXCRWGCBB-UHFFFAOYSA-N 0.000 claims description 19
- 239000007789 gas Substances 0.000 claims description 19
- 239000012071 phase Substances 0.000 claims description 13
- 239000006228 supernatant Substances 0.000 claims description 12
- MNWBNISUBARLIT-UHFFFAOYSA-N sodium cyanide Chemical compound [Na+].N#[C-] MNWBNISUBARLIT-UHFFFAOYSA-N 0.000 claims description 11
- 150000002924 oxiranes Chemical class 0.000 claims description 10
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 claims description 9
- 239000012153 distilled water Substances 0.000 claims description 9
- 238000001035 drying Methods 0.000 claims description 9
- 238000002525 ultrasonication Methods 0.000 claims description 9
- 239000012074 organic phase Substances 0.000 claims description 8
- 108090000623 proteins and genes Proteins 0.000 claims description 8
- 241000894006 Bacteria Species 0.000 claims description 7
- 239000003054 catalyst Substances 0.000 claims description 7
- 238000005215 recombination Methods 0.000 claims description 7
- 230000006798 recombination Effects 0.000 claims description 7
- 238000005695 dehalogenation reaction Methods 0.000 claims description 6
- 239000004593 Epoxy Substances 0.000 claims description 5
- KXZJHVJKXJLBKO-UHFFFAOYSA-N chembl1408157 Chemical compound N=1C2=CC=CC=C2C(C(=O)O)=CC=1C1=CC=C(O)C=C1 KXZJHVJKXJLBKO-UHFFFAOYSA-N 0.000 claims description 5
- 229910052736 halogen Inorganic materials 0.000 claims description 5
- 150000002367 halogens Chemical class 0.000 claims description 5
- 235000013844 butane Nutrition 0.000 claims description 4
- 239000012159 carrier gas Substances 0.000 claims description 4
- 239000007924 injection Substances 0.000 claims description 4
- 238000002347 injection Methods 0.000 claims description 4
- IJDNQMDRQITEOD-UHFFFAOYSA-N n-butane Chemical class CCCC IJDNQMDRQITEOD-UHFFFAOYSA-N 0.000 claims description 4
- 238000010792 warming Methods 0.000 claims description 4
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 3
- 239000001963 growth medium Substances 0.000 claims description 3
- 239000001307 helium Substances 0.000 claims description 3
- 229910052734 helium Inorganic materials 0.000 claims description 3
- SWQJXJOGLNCZEY-UHFFFAOYSA-N helium atom Chemical compound [He] SWQJXJOGLNCZEY-UHFFFAOYSA-N 0.000 claims description 3
- 230000006698 induction Effects 0.000 claims description 3
- 239000008363 phosphate buffer Substances 0.000 claims description 3
- 238000002360 preparation method Methods 0.000 claims description 3
- KWKAKUADMBZCLK-UHFFFAOYSA-N 1-octene Chemical group CCCCCCC=C KWKAKUADMBZCLK-UHFFFAOYSA-N 0.000 claims description 2
- 239000012752 auxiliary agent Substances 0.000 claims description 2
- 230000001939 inductive effect Effects 0.000 claims description 2
- 238000007689 inspection Methods 0.000 claims description 2
- 239000002994 raw material Substances 0.000 claims description 2
- WTNMMKJWWSEFNO-UHFFFAOYSA-N CC(C)[S] Chemical compound CC(C)[S] WTNMMKJWWSEFNO-UHFFFAOYSA-N 0.000 claims 1
- 241000228143 Penicillium Species 0.000 claims 1
- 229910021529 ammonia Inorganic materials 0.000 claims 1
- QGZKDVFQNNGYKY-UHFFFAOYSA-N ammonia Natural products N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 claims 1
- 150000008195 galaktosides Chemical class 0.000 claims 1
- 239000002351 wastewater Substances 0.000 abstract description 3
- 239000003651 drinking water Substances 0.000 abstract description 2
- 235000020188 drinking water Nutrition 0.000 abstract description 2
- 108010013164 halohydrin dehalogenase Proteins 0.000 abstract description 2
- 230000035945 sensitivity Effects 0.000 abstract description 2
- 150000002118 epoxides Chemical class 0.000 abstract 1
- 238000004817 gas chromatography Methods 0.000 abstract 1
- 239000010842 industrial wastewater Substances 0.000 abstract 1
- 238000006243 chemical reaction Methods 0.000 description 18
- 239000000523 sample Substances 0.000 description 10
- 238000006555 catalytic reaction Methods 0.000 description 9
- HEDRZPFGACZZDS-MICDWDOJSA-N Trichloro(2H)methane Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 8
- 230000000694 effects Effects 0.000 description 7
- 238000005119 centrifugation Methods 0.000 description 6
- 239000003153 chemical reaction reagent Substances 0.000 description 6
- 239000011734 sodium Substances 0.000 description 6
- 229910052708 sodium Inorganic materials 0.000 description 6
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 5
- CPELXLSAUQHCOX-UHFFFAOYSA-M Bromide Chemical compound [Br-] CPELXLSAUQHCOX-UHFFFAOYSA-M 0.000 description 5
- VEXZGXHMUGYJMC-UHFFFAOYSA-M Chloride anion Chemical compound [Cl-] VEXZGXHMUGYJMC-UHFFFAOYSA-M 0.000 description 5
- ZMZDMBWJUHKJPS-UHFFFAOYSA-M Thiocyanate anion Chemical compound [S-]C#N ZMZDMBWJUHKJPS-UHFFFAOYSA-M 0.000 description 5
- 238000004458 analytical method Methods 0.000 description 5
- 229940006460 bromide ion Drugs 0.000 description 5
- 238000010790 dilution Methods 0.000 description 5
- 239000012895 dilution Substances 0.000 description 5
- LRWZZZWJMFNZIK-UHFFFAOYSA-N 2-chloro-3-methyloxirane Chemical compound CC1OC1Cl LRWZZZWJMFNZIK-UHFFFAOYSA-N 0.000 description 4
- 208000035126 Facies Diseases 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 239000007864 aqueous solution Substances 0.000 description 4
- 150000001875 compounds Chemical class 0.000 description 4
- 239000003599 detergent Substances 0.000 description 4
- 238000000605 extraction Methods 0.000 description 4
- 229910021645 metal ion Inorganic materials 0.000 description 4
- JMANVNJQNLATNU-UHFFFAOYSA-N oxalonitrile Chemical compound N#CC#N JMANVNJQNLATNU-UHFFFAOYSA-N 0.000 description 4
- JHJLBTNAGRQEKS-UHFFFAOYSA-M sodium bromide Chemical compound [Na+].[Br-] JHJLBTNAGRQEKS-UHFFFAOYSA-M 0.000 description 4
- LPXPTNMVRIOKMN-UHFFFAOYSA-M sodium nitrite Chemical compound [Na+].[O-]N=O LPXPTNMVRIOKMN-UHFFFAOYSA-M 0.000 description 4
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 3
- 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 3
- TZRXHJWUDPFEEY-UHFFFAOYSA-N Pentaerythritol Tetranitrate Chemical compound [O-][N+](=O)OCC(CO[N+]([O-])=O)(CO[N+]([O-])=O)CO[N+]([O-])=O TZRXHJWUDPFEEY-UHFFFAOYSA-N 0.000 description 3
- 239000002253 acid Substances 0.000 description 3
- 238000004587 chromatography analysis Methods 0.000 description 3
- ZMZDMBWJUHKJPS-UHFFFAOYSA-N hydrogen thiocyanate Natural products SC#N ZMZDMBWJUHKJPS-UHFFFAOYSA-N 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- 229910052757 nitrogen Inorganic materials 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- 238000003786 synthesis reaction Methods 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 238000001644 13C nuclear magnetic resonance spectroscopy Methods 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 2
- 238000005481 NMR spectroscopy Methods 0.000 description 2
- IOVCWXUNBOPUCH-UHFFFAOYSA-M Nitrite anion Chemical compound [O-]N=O IOVCWXUNBOPUCH-UHFFFAOYSA-M 0.000 description 2
- 229920001213 Polysorbate 20 Polymers 0.000 description 2
- 150000001298 alcohols Chemical class 0.000 description 2
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 2
- 229960000723 ampicillin Drugs 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- 239000012470 diluted sample Substances 0.000 description 2
- 239000011344 liquid material Substances 0.000 description 2
- 230000014759 maintenance of location Effects 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 238000012544 monitoring process Methods 0.000 description 2
- 229940005654 nitrite ion Drugs 0.000 description 2
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 2
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 2
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 description 2
- 229920000053 polysorbate 80 Polymers 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- BDERNNFJNOPAEC-UHFFFAOYSA-N propan-1-ol Chemical compound CCCO BDERNNFJNOPAEC-UHFFFAOYSA-N 0.000 description 2
- 239000012488 sample solution Substances 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- ZVCDLGYNFYZZOK-UHFFFAOYSA-M sodium cyanate Chemical compound [Na]OC#N ZVCDLGYNFYZZOK-UHFFFAOYSA-M 0.000 description 2
- 235000010288 sodium nitrite Nutrition 0.000 description 2
- 238000003756 stirring Methods 0.000 description 2
- 238000005292 vacuum distillation Methods 0.000 description 2
- DEWLEGDTCGBNGU-UHFFFAOYSA-N 1,3-dichloropropan-2-ol Chemical compound ClCC(O)CCl DEWLEGDTCGBNGU-UHFFFAOYSA-N 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 102000001554 Hemoglobins Human genes 0.000 description 1
- 108010054147 Hemoglobins Proteins 0.000 description 1
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- XLJMAIOERFSOGZ-UHFFFAOYSA-N anhydrous cyanic acid Natural products OC#N XLJMAIOERFSOGZ-UHFFFAOYSA-N 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 238000011088 calibration curve Methods 0.000 description 1
- 230000003197 catalytic effect Effects 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- VXCSPBPIGBJXJR-UHFFFAOYSA-N cyanic acid;sodium Chemical compound [Na].OC#N VXCSPBPIGBJXJR-UHFFFAOYSA-N 0.000 description 1
- 230000002498 deadly effect Effects 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- 239000012847 fine chemical Substances 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 239000013461 intermediate chemical Substances 0.000 description 1
- 238000001819 mass spectrum Methods 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 150000002894 organic compounds Chemical class 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 231100000614 poison Toxicity 0.000 description 1
- 230000007096 poisonous effect Effects 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 229910052717 sulfur Inorganic materials 0.000 description 1
- 239000011593 sulfur Substances 0.000 description 1
- 230000032258 transport Effects 0.000 description 1
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention provides a method for detecting azide ions or cyanide ions in water. According to the method, halohydrin dehalogenase is utilized for catalyzing azide ions or cyanide ions in water to be reacted with epoxide to generate corresponding 4-azide-3-hydroxyl butanol or 4-cyano-3-hydroxyl butanol, a gas chromatography is adopted to quantitatively analyze the content of the generated 4-azide-3-hydroxyl butanol or 4-cyano-3-hydroxyl butanol according to the standard curve, and the concentration of the azide ions or the cyanide ions in the sample can be determined. According to the method, the accuracy is high, and azide ions and cyanide ions, which have the minimum concentrations up to 0.1mM and 0.3mM can be detected respectively; and besides, the method has good repeatability and high sensitivity, R2 of standard curves of the measured concentrations of the azide ions and the cyanide ions is 0.997 and 0.995 respectively; the method can be used for measuring the concentrations of azide ions or cyanide ions in various samples such as environment wastewater, industrial wastewater and drinking water.
Description
(1) technical field
The present invention relates in water azides ion and cryanide ion assay method, more particularly to using halide alcohol dehalogenase catalytic water
In azides ion and cryanide ion and epoxide reaction, produce corresponding substituted alcohols, by taking that gas chromatographic analysiss are generated
For alcohol amount determining the content of azides ion and cryanide ion in water, inspection of the method suitable for azides ion water and cryanide ion
Survey.
(2) background technology
Azido compound and cyanide occupy important effect in organic synthesis, and they are typically used to some doctors
The synthesis of medicine intermediate and fine chemicals, wherein Hydrazoic acid,sodium salt are additionally operable to the air bag for manufacturing automobile.But azides ion
All it is deadly poisonous compound with cryanide ion, can be combined with the hemoglobin of human body, reduce the ability of its transports oxygen, cause people to stop up
Breath and it is dead.Accordingly, it would be desirable to one fast and accurately analysis method detection water in azides ion and cryanide ion.
At present, detect that the method for azides ion and cryanide ion in water has spectrophotography, chromatography of ions, gas phase-mass spectrum connection
With analytic process etc..Wherein spectrophotography receives interfered by outside larger, prolongation over time, the numerical fluctuations of reading compared with
Greatly, degree of accuracy is not high.Chromatography of ions and gas phase-mass spectrography are more accurate, but cumbersome, high to equipment requirements.
(3) content of the invention
The invention provides in a kind of indirect detection water azides ion and cryanide ion method, the method is to equipment requirements
Low, operational stability is good, at the same analysis thus time it is short, quickly multiple samples can be analyzed.
The technical scheme is that：
The present invention provides a kind of method for detecting nitrine radical ion or cryanide ion in water, and methods described is：With de- containing halogenohydrin
Supernatant of the wet thallus that the recombination engineering bacteria of halogen enzyme coding gene is obtained Jing inducing culture Jing after ultrasonication is catalysis
Agent, with epoxide as auxiliary agent, using water sample to be measured as raw material, reacts 30～60min under the conditions of 40～45 DEG C, 500rpm,
Extract reaction solution and be extracted with ethyl acetate, take upper organic phase Jing after anhydrous sodium sulfate drying using gas chromatographic detection 4- nitrine-
3- butylated hydroxies or 4- cyano-3-hydroxy butanol peak areas, according to 4- nitrine -3- butylated hydroxies or 4- cyano-3-hydroxy fourths
Alcohol peak area is vertical coordinate, the Hydrazoic acid,sodium salt standard curve that made as abscissa with Hydrazoic acid,sodium salt or sodium cyanide concentration or Cyanogran.
Standard curve, determines the concentration of nitrine radical ion or cryanide ion in water sample to be measured；The consumption of the catalyst is with de- containing halogenohydrin
Before the recombination engineering bacteria Jing ultrasonications of halogen enzyme coding gene, wet thallus weight is calculated as 5-20mg/mL water samples to be measured (preferably
10mg/mL), the consumption of epoxide is 0.05～0.2mol/L water samples to be measured (preferred 0.18mol/L).
Further, the aminoacid sequence of the halide alcohol dehalogenase is shown in SEQ ID NO.2.
Further, gas chromatographic detection condition is：Using Japanese Shimadzu gas phase GC-14, chromatographic column Astec
CHIRALDEXTMG-TA, carrier gas are helium, and split ratio is 20:1, the temperature of injection port and detector is 220 DEG C, GC programs
Retain 5min for 120 DEG C, 5 DEG C/min is warming up to 140 DEG C, retain 2min.
Further, standard curve is prepared as follows：The aqueous sodium azide of 0.2～2.0mM is prepared with distilled water,
Add the aqueous sodium azide of 500 μ l variable concentrations, 450 μ l epoxy butanes solution and 50 μ l halogenohydrins respectively in 2ml EP pipes
Dehalogenase crude enzyme liquid, at 40 DEG C, 500rpm reacts 30min, adds 1ml ethyl acetate to be extracted respectively, take in EP pipes
Layer 800 μ l of organic faciess, adopt the peak area of vapor detection 4- nitrine -3- butylated hydroxies, Jing after anhydrous sodium sulfate drying with nitrine
Change na concn be abscissa, the peak area with 4- nitrine -3- butylated hydroxies as vertical coordinate, obtain Hydrazoic acid,sodium salt standard curve；Institute
It is the supernatant after the wet thallus ultrasonication that the fermented culture of halide alcohol dehalogenase is obtained to state halide alcohol dehalogenase crude enzyme liquid, described thick
The concentration of enzyme liquid is calculated as 0.1g/ml with wet thallus weight before ultrasonication；The epoxy butane solution is with 200mM, pH 7.5
PBS prepare 200mM epoxy butane solution, the making of Cyanogran. standard curve is with Hydrazoic acid,sodium salt standard curve.
Further, epoxide is epoxy butane or octylene oxide.
Further, the consumption of epoxide is 0.1mol/L water samples.
Further, the preparation method of catalyst is：The recombination engineering bacteria of the alcohol encoding gene of dehalogenation containing halogenohydrin is inoculated with
In the LB culture medium of the ampicillin containing 50 μ g/ml of final concentration, it is placed in 37 DEG C of shaking tables and cultivates to OD600Reach 0.6～
When 0.8, final concentration 0.2mM isopropylthiogalactosides are added, in 28 DEG C of shaking table induction 12-14 hours, 9000rpm centrifugations 10
Minute, supernatant is abandoned, the wet thallus for obtaining is centrifuged and is suspended in the phosphate buffer of 100mM according to the ratio of 0.1g/L, 50% work(
Rate ultrasonication 30min, crushes mixed liquor and is centrifuged 20 minutes in 12000rpm, collect supernatant, obtain containing halide alcohol dehalogenase
Crude enzyme liquid, as catalyst.
Detection method is the content determined by azides ion and cryanide ion, as long as so ensureing that enough halogenohydrins take off
Halogen enzyme so that azides ion and cryanide ion are converted completely just can be so that the pure enzyme quality of halide alcohol dehalogenase is accounted in crude enzyme liquid of the present invention
30% or so.
The quantitative detecting method of azides ion of the present invention and cryanide ion includes：(1) using in halide alcohol dehalogenase catalyzed samples
Azides ion or cryanide ion and epoxide (preferred epoxy butane) react, azides ion or cryanide ion are converted into corresponding
4- nitrine -3- butylated hydroxies or 4- cyano-3-hydroxy butanol；(2) using gas chromatogram detection by quantitative 4- nitrine -3- butylated hydroxies
Or 4- cyano-3-hydroxy butanol contents, and then determine the content of azides ion and cryanide ion in water.
The present invention utilizes biological catalysis, and the inorganic ionss (azides ion or cryanide ion) in water are converted into available gas phase
The organic compound of analysis, its reaction principle are shown below：
In formula, HHDH represents halide alcohol dehalogenase (halohydrin dehalogenase).
By ensureing to survey the accuracy of data, during determining from Specification Curve of Increasing to solution concentration to be measured, for
The calibration curve solution of variable concentrations and solution to be measured, in its continuous mode, each step operation parameter is identical.
Surveyed solution concentration to be measured need to be in the standard curve concentration range drawn, and data measured is only accurately, if
Concentration range of the solution concentration to be measured beyond standard curve, it is likely that do not meet its linear relationship, now can be by solution to be measured
Detected after dilution or concentration, till measured concentration is in standard curve concentration range.
Effective effect of the present invention is mainly reflected in：The inventive method accuracy is good, can detect azides ion and cyanogen from
The least concentration of son is respectively 0.1mM and 0.3mM；Meanwhile, the inventive method favorable reproducibility, sensitivity are high, its nitrine for determining
The R of the standard curve of ion and cyanide ion concentration2Respectively 0.997 and 0.995, the inventive method can be used for environmental wastewater, work
The concentration mensuration of azides ion or cryanide ion in the various samples such as industry waste water and drinking water.
(4) illustrate
Fig. 1 is the standard curve of 4- nitrine -3- butylated hydroxies and Hydrazoic acid,sodium salt.
Fig. 2 is the standard curve of 4- cyano-3-hydroxies butanol and Cyanogran..
Fig. 3 is chloride ion and bromide ion to azides ion and the impact of cryanide ion detection.
Fig. 4 is Thiocyanate ion to azides ion and the impact of cryanide ion detection.
Fig. 5 is cyanic acid ion to azides ion and the impact of cryanide ion detection.
Fig. 6 is nitrite ion to azides ion and the impact of cryanide ion detection.
(5) specific embodiment
With reference to specific embodiment, the present invention is described further, but protection scope of the present invention is not limited in
This：
Embodiment 1：Prepare halide alcohol dehalogenase
Halide alcohol dehalogenase genes of SEQ ID NO.1, using technique for gene engineering, realizes which at e. coli bl21 (DE3)
Middle expression, the aminoacid sequence of the halogenohydrin dehalogenation alcohol be SEQ ID NO.2 (constructing plan is shown in RSC Adv., 2014,4,64027-
64031).The recombination engineering bacteria of halide alcohol dehalogenase gene shown in the NO.1 of ID containing SEQ is seeded in containing 50 μ g/ml of final concentration
Ampicillin LB culture medium in, be placed in 37 DEG C of shaking tables cultivate, treat OD600When reaching 0.6～0.8, add 0.2mM different
Propyl dithiocarbamate galactoside, is transferred to 28 DEG C of shaking table induction 12-14 hours.Centrifugation 9000rpm × 10 minute, abandon supernatant.Centrifugation
The wet thallus of acquisition add the phosphate buffer of 100mM according to 1/10 (wt/vol) ratio.50% power ultrasonic crushes 30min,
Centrifugation 12000rpm × 20 minute, collect supernatant, as the crude enzyme liquid containing halide alcohol dehalogenase (every milliliter of crude enzyme liquid equivalent to
Obtain after 0.1g wet thallus are broken), -4 DEG C of preservations of refrigerator are placed in, its vigor is not almost lost in the case of frozen.
Embodiment 2：Prepare 4- nitrine -3- butylated hydroxies and 4- cyano-3-hydroxy butanol
In order to determine that azides ion and the catalysis epoxy butane open loop of cryanide ion Jing halide alcohol dehalogenases can form corresponding 4- and fold
Nitrogen -3- butylated hydroxies and 4- cyano-3-hydroxy butanol, we are using halide alcohol dehalogenase catalysis preparation 4- nitrine -3- hydroxyls first
Butanol and 4- cyano-3-hydroxy butanol.
Prepare 4- nitrine -3- butylated hydroxies：100ml PBS (200mM, pH7.5), 1ml are added in 250ml reactors
(0.017mol) epoxy butane, 2g (0.03mol) Hydrazoic acid,sodium salt and 8g halide alcohol dehalogenase wet thallus (prepared by embodiment 1).By body
The temperature of system rises to 40 DEG C, stirs 200rpm.Gas phase monitoring reaction course is used, when epoxy butane conversion ratio is more than 99%, is stopped
Only react.Centrifugation 9000rpm × 10 minute, supernatant are extracted once with 200ml ethyl acetate, separate organic faciess Jing anhydrous slufuric acid
After sodium is dried, vacuum distillation is flowed out to no solvent, obtains yellow liquid material 4- nitrine -3- butylated hydroxies.NMR is characterized：1H
NMR(500MHz,CDCl3) δ 3.71-3.37 (dd, J=5.8,3.3Hz, 1H), 3.29 (dd, J=12.4,3.3Hz, 1H),
3.27-3.25 (dd, J=12.4,7.4Hz, 1H), 2.05 (s, 1H), 1.55-1.52 (m, 2H), 1.00-0.97 (t, J=
7.5Hz,3H)。13C NMR(126MHz,CDCl3)δ72.28,56.71,56.71,27.30,9.63。
Prepare 4- cyano-3-hydroxy butanol：100ml PBS (200mM, pH7.5), 1ml are added in 250ml reactors
(0.017mol) epoxy butane, 1.5g (0.03mol) Hydrazoic acid,sodium salt and 8g halide alcohol dehalogenase wet thallus (prepared by embodiment 1).Will
The temperature of system rises to 40 DEG C, stirs 200rpm.Gas phase monitoring reaction course is used, when epoxy butane conversion ratio is more than 99%,
Stopped reaction.Centrifugation 9000rpm × 10 minute, supernatant are extracted once with 200ml ethyl acetate, separate the anhydrous sulfur of organic faciess Jing
After sour sodium is dried, vacuum distillation is flowed out to no solvent, obtains yellow liquid material 4- cyano-3-hydroxy butanol.NMR is characterized：1H NMR(500MHz,CDCl3) δ 3.89-3.86 (dd, J=6.1,5.0Hz, 1H), 2.56-2.47 (m, 3H), 1.64-1.61
(m, 2H), 1.00-0.97 (t, J=7.5Hz, 3H).13C NMR(126MHz,CDCl3)δ117.92,69.14,29.28,
25.57,9.90。
Vapor detection epoxy butane：Agilent GC-7890A, chromatographic column HP-5, carrier gas are nitrogen, injection port and detector
Temperature be respectively 230 DEG C and 250 DEG C, GC programs are, 60 DEG C retain 4min, and 20 DEG C/min is warming up to 120 DEG C.Epoxy butane
Retention time is 2.7min.
Embodiment 3：The foundation of 4- nitrine -3- butylated hydroxies and 4- cyano-3-hydroxy butanol gas phase detection methods
4- nitrine -3- butylated hydroxies and 4- cyano-3-hydroxies butanol are using Japanese Shimadzu gas phase GC-14, chromatographic column Astec
CHIRALDEXTMG-TA, carrier gas are helium, and split ratio is 20:1, the temperature of injection port and detector is 220 DEG C, GC programs
Retain 5min for 120 DEG C, 5 DEG C/min is warming up to 140 DEG C, retain 2min.4- nitrine -3- butylated hydroxies and 4- cyano-3-hydroxies
The retention time of butanol is respectively 4.62min and 7.56min.
Embodiment 4：Draw the standard curve of azides ion and cryanide ion
The inventive method it is critical to insure that azides ion and cryanide ion in testing sample to be fully converted to correspondence
Substituted alcohols, therefore we will consider the following aspects：(1) excessive epoxy butane substrate, (2) enough halogenohydrins is needed to take off
Halogen enzyme, the optimum condition of (3) halide alcohol dehalogenase catalysis.In view of some considers that we establish following measure scheme above：
Prepare the epoxy butane solution of 200mM, prepared as solvent with 200mM (pH 7.5) PBSs.
Prepare halide alcohol dehalogenase crude enzyme liquid, the halide alcohol dehalogenase crude enzyme liquid prepared with embodiment 1.
(1) 4- nitrine -3- butylated hydroxy standard curves：With distilled water prepare 0.1mM, 0.2mM, 0.3mM, 0.4mM,
4- nitrine -3- butylated hydroxy the aqueous solutions of 0.5mM and 1.0mM, the 4- nitrine -3- butylated hydroxy aqueous solutions for taking variable concentrations are each
1ml, is added separately in different 2ml EP pipes, is subsequently separately added into 1ml ethyl acetate and is extracted.Take upper organic phase
800 μ l, Jing after anhydrous sodium sulfate drying, carry out vapor detection (testing conditions are with embodiment 3), with 4- nitrine -3- butylated hydroxies
Concentration is abscissa, and with 4- nitrine -3- butylated hydroxy peak areas as vertical coordinate, 4- nitrine -3- butylated hydroxies standard curve is y=
3375.33x-42.03(R2=0.996), see Fig. 1.
(2) Hydrazoic acid,sodium salt standard curve：0.2mM, 0.4mM, 0.6mM, 0.8mM, 1.0mM and 2.0mM are prepared with distilled water
Aqueous sodium azide as water sample to be measured, in 2ml EP pipes add the Hydrazoic acid,sodium salt of 500 μ l variable concentrations water-soluble respectively
The halide alcohol dehalogenase crude enzyme liquid of liquid, 450 μ l epoxy butanes solution and 50 μ l.It is placed in Thermomixer reactors, 40 DEG C,
500rpm, reacts 30min.The extraction of 1ml ethyl acetate is added in EP pipes respectively, 800 μ l Jing anhydrous slufuric acids of upper organic phase are taken
After sodium is dried, vapor detection 4- nitrine -3- butylated hydroxy peak areas (condition is shown in embodiment 3).With 4- nitrine -3- butylated hydroxies peak
Area is vertical coordinate, with Hydrazoic acid,sodium salt concentration as abscissa, makes the standard curve y=3306.37x-20.11 of Hydrazoic acid,sodium salt
(R2=0.997) (see Fig. 1).
(3) 4- cyano-3-hydroxies butanol standard curve：With distilled water prepare 0.3mM, 0.6mM, 0.9mM, 1.2mM,
The 4- cyano-3-hydroxy butanol aqueous solutions of 1.5mM and 2.0mM, the 4- cyano-3-hydroxy butanol aqueous solutions for taking variable concentrations are each
1ml, is added separately in different 2ml EP pipes, is subsequently separately added into 1ml ethyl acetate and is extracted.Take upper organic phase
800 μ l, Jing after anhydrous sodium sulfate drying, carry out vapor detection (see embodiment 3).Concentration with 4- cyano-3-hydroxy butanol is
Abscissa, with peak area as vertical coordinate, the standard curve of 4- cyano-3-hydroxy butanol is y=1518.46x-67.52 (R2=
0.993) (see Fig. 2).
(4) Cyanogran. standard curve：0.6mM, 1.2mM, 1.8mM, 2.4mM, 3.0mM and 4.0mM are prepared with distilled water
Sodium cyanide solution, adds the sodium cyanide solution of 500 μ l variable concentrations in 2ml EP pipes, 450 μ l epoxy butanes solution and
The halide alcohol dehalogenase crude enzyme liquid of 50 μ l.Thermomixer reactors are placed in, 40 DEG C, 500rpm reacts 30min.Add in EP pipes
Enter the extraction of 1ml ethyl acetate, 800 μ l of upper organic phase are taken Jing after anhydrous sodium sulfate drying, carry out vapor detection 4- cyano group -3- hydroxyls
Base butanol peak area (see embodiment 3).With sodium cyanide concentration as abscissa, with 4- cyano-3-hydroxy butanol peak areas as vertical seat
Mark, the standard curve of Cyanogran. is y=1371.50x-40.75 (R2=0.995) (see Fig. 2).
Fig. 2 can be seen that the standard curve of Cyanogran. and the linear relationship phase of the standard curve of 4- cyano-3-hydroxy butanol
Seemingly, the standard curve for illustrating the Cyanogran. set up is accurately feasible.
Embodiment 5：Azides ion and cryanide ion are determined using halide alcohol dehalogenase
With the sodium azide solution or sodium cyanide solution 10mL of distilled water random arrangement unknown concentration, fold as to be measured
Nitrogen ion sample or cryanide ion sample.
Measure scheme：Testing sample is diluted into 2 times, 5 times and 1 times with pure water respectively, in 2ml EP pipes adds 500 μ l dilute
Release sample solution, 450 μ l epoxy butane solution (embodiment 4 is prepared) and 50 μ l halide alcohol dehalogenase crude enzyme liquids (embodiment 1).It is placed in
Thermomixer reactors, 40 DEG C, 500rpm reacts 30min.The extraction of 1ml ethyl acetate is added in EP pipes, taking upper strata has
Machine phase carries out gas phase analysis Jing after anhydrous sodium sulfate drying.4- nitrine -3- the butylated hydroxies for obtaining or 4- cyano-3-hydroxies
The gas phase peak area of butanol is compared with Cyanogran. standard curve or Hydrazoic acid,sodium salt standard curve, selects peak area accordingly marking
Extension rate in the range of directrix curve peak area, obtains the concentration of azides ion or cryanide ion in water sample to be measured, as a result such as table 1
It is shown.
The measure of 1 azides ion of table and cyanide ion concentration
[a]Actual concentrations with dilute 10 calculated by peak area, dilute 5 times and 2 times peak area exceed Hydrazoic acid,sodium salt mark
Directrix curve, computing formula (1) is：
X[actual concentrations]=((Y[peak area]+20.11)/3306.37)×2[multiple of reaction dilution]×10[multiple of diluted sample]
X in formula (1)[actual concentrations]For the actual concentrations of measurement, Y[peak area]For 4- nitrine -3- butylated hydroxy peak areas,
2[multiple of reaction dilution]Refer to 500ul water samples, reaction final volume is 1mL, that is, dilute 2 times.
[b]Actual concentrations with dilute 10 calculated by peak area, it is also possible to dilute 5 times of calculated by peak area, dilute 2 times of peak
Standard curve of the area beyond Cyanogran..Computing formula (2) is：
X[actual concentrations]=((Y[peak area]+67.52)/1515.46)×2[multiple of reaction dilution]×10[multiple of diluted sample]
X in formula (2)[actual concentrations]For the actual concentrations of measurement, Y[peak area]For 4- cyano-3-hydroxy butanol peak areas,
2[multiple of reaction dilution]Refer to 500ul water samples, reaction final volume is 1mL, that is, dilute 2 times.
Embodiment 6：The impact of metal ion and detergent to halide alcohol dehalogenase vigor
In view of can there is some metal ions and detergent such as Tween 80 and Tween 20 in detected sample, this
The bright impact for also having investigated part metals ion and detergent to halide alcohol dehalogenase catalytic reaction.Prepared with distilled water and do not allow concentration
Reagent solution (being shown in Table 1) and 50mM 1,3-, bis- chloro- 2- aqueous propanol solution, make of halide alcohol dehalogenase crude enzyme liquid (embodiment 1)
Catalytic reaction.500 μ l reagent solutions, 1,3-, the bis- chloro- 2- aqueous propanol solution of 450 μ l 50mM and 50 μ are added in 2ml EP pipes
L halide alcohol dehalogenase crude enzyme liquids.Thermomixer reactors are placed in, 40 DEG C, 500rpm reacts 10min.Control reaction is with 500 μ l
Distilled water replaces reagent solution.The relative activity under the conditions of each is calculated with the volume of production of product epoxychloropropane.Enzyme activity is defined
It is that, at 40 DEG C, under the reaction condition of pH 7.5, the enzyme amount required for 1 μm of ol epoxychloropropane synthesis of catalysis per minute is 1 enzyme
Unit living.
As shown in table 2, Fe3+、Ba2+、Al3+And Cu2+There is stronger inhibitory action to the vigor of halide alcohol dehalogenase, and EDTA
There was only slight impact to halide alcohol dehalogenase vigor.Therefore, during practical measurement, in order to ensure efficient halogenohydrin dehalogenation
Enzyme activity, can remove metal ion with edta reagent, so can ensure that the accuracy for determining reaction.Bis- chloro- 2- third of 1,3-
Gas phase process of the vapor detection of alcohol and epoxychloropropane with embodiment 2, the reservation of 1,3- bis- chloro- 2- propanol and epoxychloropropane
Time is respectively 4.04min and 6.41min.
The impact of 2. metal ion of table and detergent to halide alcohol dehalogenase catalysis activity
| Reagent | Concentration | Relative activity (%) |
| Control | - | 100.0 |
| Fe2+ | 5mM | 69.02±0.34 |
| Ni2+ | 5mM | 59.7±1.92 |
| Cu2+ | 5mM | 23.74±3.56 |
| Ca2+ | 5mM | 98.04±1.23 |
| Mn2+ | 5mM | 128.34±0.41 |
| Zn2+ | 5mM | 76.60±3.78 |
| Mg2+ | 5mM | 101.05±6.34 |
| Al3+ | 5mM | 13.56±0.98 |
| Fe3+ | 5mM | 0±0.0 |
| Ba2+ | 1mM | 0±0.0 |
| EDTA | 5mM | 79.86±0.86 |
| Tween 80 | 2% (V/V) | 92.70±0.66 |
| Tween 20 | 2% (V/V) | 70.98±0.78 |
Embodiment 7：The impact of chloride ion, bromide ion, nitrite anions, thiocyanate and cyanate radical to detecting
Chloride ion, bromide ion, nitrite anions, thiocyanate and cyanate radical and epoxy can also be catalyzed in view of halide alcohol dehalogenase
Compound is reacted, therefore, the present invention also investigated chloride ion, bromide ion, nitrite anions, thiocyanate and cyanate radical to nitrine from
Son and the impact of cryanide ion detection.
Sodium Chloride, sodium bromide, sodium nitrite, sodium sulfocynanate and the cyanic acid sodium water solution for preparing 20mM respectively is molten as sample
Liquid, adds above-mentioned each sample solution of 500 μ l, 450 μ l epoxy butane solution in 2ml EP pipes (200mM, embodiment 4 are prepared)
With the halide alcohol dehalogenase crude enzyme liquid (embodiment 1) of 50 μ l.Thermomixer reactors are placed in, 40 DEG C, 500rpm reacts
30min.The extraction of 1ml ethyl acetate is added in EP pipes, upper organic phase Jing anhydrous sodium sulfate drying is taken.To in organic faciess respectively
Add a certain amount of 4- nitrine -3- butylated hydroxies or 4- cyano-3-hydroxy butanol (addition it is how many on this experiment without affecting,
Main purpose is to judge whether the compound that other nucleopilic reagents are formed affects the compound that azides ion or cryanide ion are formed
Detection, the present embodiment addition be about 500 μ l ethyl acetate in add 1 μ l 4- nitrine -3- butylated hydroxies or 4- cyano group -3-
Butylated hydroxy), carry out gas phase analysis.
Whether the product of observation Sodium Chloride, sodium bromide, sodium nitrite, sodium sulfocynanate and Sodium cyanate (NaOCN) and epoxy butane reaction is right
4- nitrine -3- butylated hydroxies and the detection of 4- cyano-3-hydroxies butanol have an impact.Wherein chloride ion and bromide ion to azides ion and
The detection of cryanide ion does not affect (Fig. 3), and detection of the Thiocyanate ion on azides ion and cryanide ion does not affect (Fig. 4), cyanogen
The detection of acid ion azides ion and cryanide ion does not affect (Fig. 5), and nitrite ion does not affect the detection of azides ion
But affect the detection (Fig. 6) of cryanide ion.
Claims (4)
1. in a kind of detection water nitrine radical ion or cryanide ion method, it is characterised in that methods described is：With dehalogenation containing halogenohydrin
Supernatant of the recombination engineering bacteria of enzyme coding gene Jing after the wet thallus ultrasonication that inducing culture is obtained is catalyst, with
Epoxide is auxiliary agent, using water sample to be measured as raw material, 30～60min is reacted under the conditions of 40～45 DEG C, 500rpm, and negating should
Liquid is extracted with ethyl acetate, and takes upper organic phase and gas chromatographic detection 4- nitrine -3- hydroxyls are adopted Jing after anhydrous sodium sulfate drying
Butanol or 4- cyano-3-hydroxy butanol peak areas, according to 4- nitrine -3- butylated hydroxies or 4- cyano-3-hydroxy butanol peaks face
Accumulate as vertical coordinate, the Hydrazoic acid,sodium salt standard curve made as abscissa with Hydrazoic acid,sodium salt or sodium cyanide concentration or Cyanogran. standard song
Line, determines the concentration of nitrine radical ion or cryanide ion in water sample to be measured；The consumption of the catalyst is with containing halide alcohol dehalogenase volume
Before the recombination engineering bacteria Jing ultrasonications of code gene, wet thallus weight is calculated as 5～20mg/mL water samples to be measured, epoxide
Consumption is 0.05～0.2mol/L water samples to be measured；The epoxide is epoxy butane or octylene oxide；The gas chromatogram inspection
Survey condition is：Using Japanese Shimadzu gas phase GC-14, chromatographic column Astec CHIRALDEXTMG-TA, carrier gas is helium, split ratio
For 20:1, the temperature of injection port and detector is 220 DEG C, and GC programs are 120 DEG C of reservation 5min, and 5 DEG C/min is warming up to 140
DEG C, retain 2min；The standard curve is prepared as follows：The Hydrazoic acid,sodium salt that 0.2～2.0mM is prepared with distilled water is water-soluble
Liquid, adds the aqueous sodium azide of 500 μ l variable concentrations, 450 μ l epoxy butanes solution and 50 μ l halogen respectively in 2mlEP pipes
Alcohol dehalogenase crude enzyme liquid, at 40 DEG C, 500rpm reacts 30min, adds 1ml ethyl acetate to be extracted respectively, take in EP pipes
800 μ l of upper organic phase, adopt the peak area of vapor detection 4- nitrine -3- butylated hydroxies Jing after anhydrous sodium sulfate drying, with folded
Nitridation na concn be abscissa, the peak area with 4- nitrine -3- butylated hydroxies as vertical coordinate, obtain Hydrazoic acid,sodium salt standard curve；
The halide alcohol dehalogenase crude enzyme liquid is the supernatant after the wet thallus ultrasonication that the fermented culture of halide alcohol dehalogenase is obtained, described
The concentration of crude enzyme liquid is calculated as 0.1g/ml with wet thallus weight before ultrasonication；The epoxy butane solution is with 200mM, pH
The 200mM epoxy butane solution that 7.5 PBS is prepared, the making of Cyanogran. standard curve is with Hydrazoic acid,sodium salt standard curve.
2. the method for detecting nitrine radical ion or cryanide ion in water as claimed in claim 1, it is characterised in that the halogenohydrin dehalogenation
The aminoacid sequence of enzyme is shown in SEQ ID NO.2.
3. the method for detecting nitrine radical ion or cryanide ion in water as claimed in claim 1, it is characterised in that the use of epoxide
Measure as 0.1mol/L water samples.
4. the method for detecting nitrine radical ion or cryanide ion in water as claimed in claim 1, it is characterised in that the preparation of catalyst
Method is：The recombination engineering bacteria of the alcohol encoding gene of dehalogenation containing halogenohydrin is seeded in into the ammonia benzyl penicillium sp containing 50 μ g/ml of final concentration
In the LB culture medium of element, it is placed in 37 DEG C of shaking tables and cultivates to OD600When reaching 0.6～0.8, final concentration 0.2mM isopropyl sulfur is added
For galactoside, in 28 DEG C of shaking table induction 12-14 hours, 9000rpm is centrifuged 10 minutes, abandons supernatant, the wet thallus for obtaining are centrifuged
Ratio according to 0.1g/L is suspended in the phosphate buffer of 100mM, and 50% power ultrasonic crushes 30min, and broken mixed liquor exists
12000rpm is centrifuged 20 minutes, collects supernatant, obtains the crude enzyme liquid containing halide alcohol dehalogenase, as catalyst.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510108903.6A CN104749273B (en) | 2015-03-12 | 2015-03-12 | Method for detecting azide ions or cyanide ions in water |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510108903.6A CN104749273B (en) | 2015-03-12 | 2015-03-12 | Method for detecting azide ions or cyanide ions in water |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104749273A CN104749273A (en) | 2015-07-01 |
| CN104749273B true CN104749273B (en) | 2017-04-12 |
Family
ID=53589267
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510108903.6A Active CN104749273B (en) | 2015-03-12 | 2015-03-12 | Method for detecting azide ions or cyanide ions in water |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104749273B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108918443A (en) * | 2018-07-14 | 2018-11-30 | 浙江迪特西科技有限公司 | A kind of integrated portable water-quality COD detection device |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1158054A1 (en) * | 2000-05-25 | 2001-11-28 | Rijksuniversiteit te Groningen | Enzymatic conversion of epoxides |
| CN104263713A (en) * | 2014-08-29 | 2015-01-07 | 浙江工业大学 | Tistrella mobilis, halohydrin dehalogenase, gene, vector, recombinant strain and application of halohydrin dehalogenase |
| CN104357468A (en) * | 2014-10-17 | 2015-02-18 | 浙江工业大学 | Parvibaculum lavamentivorans ZJB 14001, halohydrin dehalogenase enzyme gene, enzyme, engineered bacterium and application |
-
2015
- 2015-03-12 CN CN201510108903.6A patent/CN104749273B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1158054A1 (en) * | 2000-05-25 | 2001-11-28 | Rijksuniversiteit te Groningen | Enzymatic conversion of epoxides |
| CN104263713A (en) * | 2014-08-29 | 2015-01-07 | 浙江工业大学 | Tistrella mobilis, halohydrin dehalogenase, gene, vector, recombinant strain and application of halohydrin dehalogenase |
| CN104357468A (en) * | 2014-10-17 | 2015-02-18 | 浙江工业大学 | Parvibaculum lavamentivorans ZJB 14001, halohydrin dehalogenase enzyme gene, enzyme, engineered bacterium and application |
Non-Patent Citations (3)
| Title |
|---|
| Enantioselective Ring Opening of Epoxides with Cyanide Catalysed Non-Racemic b-Hydroxy Nitriles;Maja Majeric´Elenkov et al.;《Adv.Synth.Catal.》;20061231;第348卷;全文 * |
| Highly Enantioselective and Regioselective Biocatalytic Azidolysis of Aromatic Epoxides;Jeffrey H.Lutje Spelberg et al.;《ORGANIC LETTERS》;20011213;第3卷(第1期);全文 * |
| 多功能生物催化剂———卤醇脱卤酶的研究进展;郑楷 等;《化工学报》;20081231;第59卷(第12期);全文 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104749273A (en) | 2015-07-01 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN110455896B (en) | Preparation method of metal organic framework composite ratio electrochemical miR3123 aptamer sensor | |
| Zeng et al. | Determination of melamine by flow injection analysis based on chemiluminescence system | |
| da Silva et al. | Direct nitrate sensing in water using an array of copper-microelectrodes from flat flexible cables | |
| del Pozo et al. | Cucurbit [8] uril-based electrochemical sensors as detectors in flow injection analysis. Application to dopamine determination in serum samples | |
| Cao et al. | Electrochemical sensing of melamine with 3, 4-dihydroxyphenylacetic acid as recognition element | |
| Cinková et al. | Electrochemical method for point-of-care determination of ciprofloxacin using boron-doped diamond electrode | |
| CN105385753A (en) | Electrochemical sensor for detecting isocarbophos based on nucleic acid aptamer and preparation method of electrochemical sensor | |
| CN104749273B (en) | Method for detecting azide ions or cyanide ions in water | |
| CN104020199B (en) | A kind of method based on fit recognition reaction electrochemical gaging dopamine | |
| Ye et al. | An electrochemiluminescence sensor for adrenaline assay based on the tyrosinase/SiC/chitosan modified electrode | |
| CN110308140B (en) | Method for detecting chemical substances by using potassium ferrate | |
| CN105403612A (en) | Method for rapidly detecting pesticide residue based on plant esterase | |
| CN109459478B (en) | miRNA-21 electrochemical sensor based on copper-hexahydroxy triphenylene and detection method | |
| Wang et al. | Amperometric Flow‐Biosensor for Cyanide Based on an Inhibitory Effect upon Bioelectrocatalytic Reduction of Oxygen by Peroxidase‐Modified Carbon‐Felt | |
| Qu et al. | A homogeneous electrochemical aptasensor based on DNA assembly for zearalenone detection | |
| CN112098381B (en) | Method for detecting chromium by combining copper-doped carbon dot mimic enzyme with fluorescent probe | |
| CN109021000A (en) | A kind of fluorescence probe, synthetic method and application detecting hydrogen peroxide | |
| Zhao et al. | Analyte-induced formation of partial duplexes for the preparation of a label-free electrochemiluminescent aptasensor | |
| CN108226493A (en) | A kind of industrial wastewater bio-toxicity detection method based on genetic recombination photogen | |
| CN107167443A (en) | A kind of method that utilization ultraviolet spectrometer detects PCB77 | |
| Wu et al. | Electrochemical impedance sensor based on nano-cobalt-oxide-modified graphenic electrode for total phosphorus determinations in water | |
| CN102692409B (en) | Method and kit for detecting concentration of chiral amino acid | |
| Chen et al. | Flow-injection determination of iron (III) in soil by biamperometry using two independent redox couples | |
| CN110632139B (en) | Method for cathode photoelectrochemical detection of zearalenone | |
| CN105717099B (en) | A kind of preparation method and application of electrogenerated chemiluminescence furazolidone biology sensor |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |