CN104090014A - Method and device for detection of water acute toxicity - Google Patents
Method and device for detection of water acute toxicity Download PDFInfo
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- CN104090014A CN104090014A CN201310110981.0A CN201310110981A CN104090014A CN 104090014 A CN104090014 A CN 104090014A CN 201310110981 A CN201310110981 A CN 201310110981A CN 104090014 A CN104090014 A CN 104090014A
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- ammonium ion
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- acute toxicity
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- 238000001514 detection method Methods 0.000 title claims abstract description 53
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 title claims abstract description 20
- 230000007059 acute toxicity Effects 0.000 title claims abstract description 18
- 231100000403 acute toxicity Toxicity 0.000 title claims abstract description 18
- 238000000034 method Methods 0.000 title abstract description 15
- QGZKDVFQNNGYKY-UHFFFAOYSA-O Ammonium Chemical compound [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 claims abstract description 49
- 241000894006 Bacteria Species 0.000 claims abstract description 44
- IOVCWXUNBOPUCH-UHFFFAOYSA-M Nitrite anion Chemical compound [O-]N=O IOVCWXUNBOPUCH-UHFFFAOYSA-M 0.000 claims abstract description 26
- 238000005206 flow analysis Methods 0.000 claims abstract description 17
- 239000007853 buffer solution Substances 0.000 claims abstract description 14
- 230000000694 effects Effects 0.000 claims abstract description 14
- 230000003647 oxidation Effects 0.000 claims abstract description 7
- 238000007254 oxidation reaction Methods 0.000 claims abstract description 7
- 239000011159 matrix material Substances 0.000 claims abstract description 6
- 239000012528 membrane Substances 0.000 claims description 32
- 238000012546 transfer Methods 0.000 claims description 31
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 claims description 22
- 238000012360 testing method Methods 0.000 claims description 18
- 230000001580 bacterial effect Effects 0.000 claims description 17
- 230000002572 peristaltic effect Effects 0.000 claims description 17
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 claims description 14
- 239000000243 solution Substances 0.000 claims description 14
- 229910021607 Silver chloride Inorganic materials 0.000 claims description 11
- 235000019270 ammonium chloride Nutrition 0.000 claims description 11
- 239000007788 liquid Substances 0.000 claims description 11
- 239000004800 polyvinyl chloride Substances 0.000 claims description 11
- 229920000915 polyvinyl chloride Polymers 0.000 claims description 11
- HKZLPVFGJNLROG-UHFFFAOYSA-M silver monochloride Chemical compound [Cl-].[Ag+] HKZLPVFGJNLROG-UHFFFAOYSA-M 0.000 claims description 11
- 230000002401 inhibitory effect Effects 0.000 claims description 10
- 239000002351 wastewater Substances 0.000 claims description 10
- RMIXHJPMNBXMBU-UHFFFAOYSA-N Nonactin Natural products CC1C(=O)OC(C)CC(O2)CCC2C(C)C(=O)OC(C)CC(O2)CCC2C(C)C(=O)OC(C)CC(O2)CCC2C(C)C(=O)OC(C)CC2CCC1O2 RMIXHJPMNBXMBU-UHFFFAOYSA-N 0.000 claims description 9
- 229920000642 polymer Polymers 0.000 claims description 7
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 claims description 7
- 241000605122 Nitrosomonas Species 0.000 claims description 6
- RMIXHJPMNBXMBU-QIIXEHPYSA-N Nonactin Chemical group C[C@H]([C@H]1CC[C@H](O1)C[C@@H](OC(=O)[C@@H](C)[C@@H]1CC[C@@H](O1)C[C@@H](C)OC(=O)[C@H](C)[C@H]1CC[C@H](O1)C[C@H](C)OC(=O)[C@H]1C)C)C(=O)O[C@H](C)C[C@H]2CC[C@@H]1O2 RMIXHJPMNBXMBU-QIIXEHPYSA-N 0.000 claims description 5
- YPUPRVWRYDPGCW-UHFFFAOYSA-N Monactin Natural products CC1C(=O)OC(C)CC(O2)CCC2C(C)C(=O)OC(C)CC(O2)CCC2C(C)C(=O)OC(CC)CC(O2)CCC2C(C)C(=O)OC(C)CC2CCC1O2 YPUPRVWRYDPGCW-UHFFFAOYSA-N 0.000 claims description 4
- 239000002555 ionophore Substances 0.000 claims description 4
- 230000000236 ionophoric effect Effects 0.000 claims description 4
- YPUPRVWRYDPGCW-GGOMOPATSA-N monactin Chemical compound C[C@H]([C@H]1CC[C@H](O1)C[C@@H](OC(=O)[C@@H](C)[C@@H]1CC[C@@H](O1)C[C@@H](C)OC(=O)[C@H](C)[C@H]1CC[C@H](O1)C[C@H](C)OC(=O)[C@H]1C)CC)C(=O)O[C@H](C)C[C@H]2CC[C@@H]1O2 YPUPRVWRYDPGCW-GGOMOPATSA-N 0.000 claims description 4
- 239000002699 waste material Substances 0.000 claims description 4
- NKJOXAZJBOMXID-UHFFFAOYSA-N 1,1'-Oxybisoctane Chemical compound CCCCCCCCOCCCCCCCC NKJOXAZJBOMXID-UHFFFAOYSA-N 0.000 claims description 3
- LQNUZADURLCDLV-UHFFFAOYSA-N nitrobenzene Chemical group [O-][N+](=O)C1=CC=CC=C1 LQNUZADURLCDLV-UHFFFAOYSA-N 0.000 claims description 3
- -1 poly-butylacrylic acid ester Chemical class 0.000 claims description 3
- 238000003756 stirring Methods 0.000 claims description 3
- CXVOIIMJZFREMM-UHFFFAOYSA-N 1-(2-nitrophenoxy)octane Chemical compound CCCCCCCCOC1=CC=CC=C1[N+]([O-])=O CXVOIIMJZFREMM-UHFFFAOYSA-N 0.000 claims description 2
- 239000004808 2-ethylhexylester Substances 0.000 claims description 2
- SCJNXSCHPUAAGK-UHFFFAOYSA-N B(O)(O)O.ClC1=CC=C(C=C1)[K] Chemical compound B(O)(O)O.ClC1=CC=C(C=C1)[K] SCJNXSCHPUAAGK-UHFFFAOYSA-N 0.000 claims description 2
- JZLNMCGKKHXZMI-UHFFFAOYSA-N B(O)(O)O.FC(C=1C=C(C=C(C1)C(F)(F)F)[Na])(F)F Chemical compound B(O)(O)O.FC(C=1C=C(C=C(C1)C(F)(F)F)[Na])(F)F JZLNMCGKKHXZMI-UHFFFAOYSA-N 0.000 claims description 2
- PYGXAGIECVVIOZ-UHFFFAOYSA-N Dibutyl decanedioate Chemical compound CCCCOC(=O)CCCCCCCCC(=O)OCCCC PYGXAGIECVVIOZ-UHFFFAOYSA-N 0.000 claims description 2
- 239000004697 Polyetherimide Substances 0.000 claims description 2
- 125000000484 butyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- MIMDHDXOBDPUQW-UHFFFAOYSA-N dioctyl decanedioate Chemical compound CCCCCCCCOC(=O)CCCCCCCCC(=O)OCCCCCCCC MIMDHDXOBDPUQW-UHFFFAOYSA-N 0.000 claims description 2
- 229920001971 elastomer Polymers 0.000 claims description 2
- 229920000058 polyacrylate Polymers 0.000 claims description 2
- 229920001601 polyetherimide Polymers 0.000 claims description 2
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 abstract description 6
- 239000001301 oxygen Substances 0.000 abstract description 6
- 229910052760 oxygen Inorganic materials 0.000 abstract description 6
- 241001148470 aerobic bacillus Species 0.000 abstract description 4
- 238000004458 analytical method Methods 0.000 abstract description 3
- 239000000463 material Substances 0.000 abstract description 3
- 230000001473 noxious effect Effects 0.000 abstract description 3
- 230000001988 toxicity Effects 0.000 abstract description 3
- 231100000419 toxicity Toxicity 0.000 abstract description 3
- 238000005516 engineering process Methods 0.000 abstract description 2
- 238000004364 calculation method Methods 0.000 abstract 1
- 238000003912 environmental pollution Methods 0.000 abstract 1
- 230000007704 transition Effects 0.000 abstract 1
- HTKFORQRBXIQHD-UHFFFAOYSA-N allylthiourea Chemical compound NC(=S)NCC=C HTKFORQRBXIQHD-UHFFFAOYSA-N 0.000 description 9
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 8
- 229960001748 allylthiourea Drugs 0.000 description 8
- 239000012530 fluid Substances 0.000 description 8
- 239000003344 environmental pollutant Substances 0.000 description 6
- 150000002500 ions Chemical class 0.000 description 6
- 231100000719 pollutant Toxicity 0.000 description 6
- 238000005086 pumping Methods 0.000 description 5
- 241001495394 Nitrosospira Species 0.000 description 4
- 229910021529 ammonia Inorganic materials 0.000 description 4
- 230000002503 metabolic effect Effects 0.000 description 4
- 238000011197 physicochemical method Methods 0.000 description 3
- 230000007096 poisonous effect Effects 0.000 description 3
- 241000251468 Actinopterygii Species 0.000 description 2
- 241000195493 Cryptophyta Species 0.000 description 2
- 241000192147 Nitrosococcus Species 0.000 description 2
- 241000605121 Nitrosomonas europaea Species 0.000 description 2
- 238000010170 biological method Methods 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 238000004401 flow injection analysis Methods 0.000 description 2
- 229910001385 heavy metal Inorganic materials 0.000 description 2
- 231100000614 poison Toxicity 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000006467 substitution reaction Methods 0.000 description 2
- LMSDCGXQALIMLM-UHFFFAOYSA-N 2-[2-[bis(carboxymethyl)amino]ethyl-(carboxymethyl)amino]acetic acid;iron Chemical compound [Fe].OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O LMSDCGXQALIMLM-UHFFFAOYSA-N 0.000 description 1
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 1
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 1
- 125000003903 2-propenyl group Chemical group [H]C([*])([H])C([H])=C([H])[H] 0.000 description 1
- VNFYMAPAENTMMO-UHFFFAOYSA-N 5-chloro-2-methylquinoline Chemical compound ClC1=CC=CC2=NC(C)=CC=C21 VNFYMAPAENTMMO-UHFFFAOYSA-N 0.000 description 1
- 241000238571 Cladocera Species 0.000 description 1
- 239000007995 HEPES buffer Substances 0.000 description 1
- 241000192123 Nitrosovibrio Species 0.000 description 1
- 239000004695 Polyether sulfone Substances 0.000 description 1
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 1
- 241000607598 Vibrio Species 0.000 description 1
- 239000003905 agrochemical Substances 0.000 description 1
- 229940008075 allyl sulfide Drugs 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 238000001675 atomic spectrum Methods 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
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- 239000004202 carbamide Substances 0.000 description 1
- 238000013016 damping Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
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- 231100000290 environmental risk assessment Toxicity 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
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- 244000005700 microbiome Species 0.000 description 1
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- 230000009935 nitrosation Effects 0.000 description 1
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- 235000015097 nutrients Nutrition 0.000 description 1
- 150000002894 organic compounds Chemical class 0.000 description 1
- 229920006393 polyether sulfone Polymers 0.000 description 1
- 229920006254 polymer film Polymers 0.000 description 1
- 238000004451 qualitative analysis Methods 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- HNBDQABBWNOTRU-UHFFFAOYSA-N thalline Chemical compound C1=CC=[Tl]C=C1 HNBDQABBWNOTRU-UHFFFAOYSA-N 0.000 description 1
- 231100000820 toxicity test Toxicity 0.000 description 1
- WUUHFRRPHJEEKV-UHFFFAOYSA-N tripotassium borate Chemical compound [K+].[K+].[K+].[O-]B([O-])[O-] WUUHFRRPHJEEKV-UHFFFAOYSA-N 0.000 description 1
- 238000003828 vacuum filtration Methods 0.000 description 1
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention belongs to the technical field of environmental pollution detection and relates to a method and a device for detection of water acute toxicity. The method realizes toxicity analysis according to a reduction degree of an ammonium ion oxidation rate after nitrite bacteria are inhibited by noxious materials. Bacteria and a selective electrode are connected in a flow analysis system in series, a sample is fed into the system and is acted by the bacteria, then a medium is transformed, an ammonium ion-containing buffer solution is used, and an ammonium ion oxidation rate of the bacteria can be obtained by calculation of change of potential signals detected by an ammonium ion selective electrode. The method utilizes a medium transition technology, eliminates matrix effects and avoids electrode pollution and damage caused by a sample. The method and the device utilize the ammonium ion selective electrode to realize ammoxidation rate determination so that interference produced by other aerobic bacteria in oxygen electrode-based dissolved oxygen determination is avoided.
Description
Technical field
The invention belongs to environment pollution detection technical field, relate in particular to a kind of detection method and device thereof of acute toxicity of water quality.
Background technology
The method detecting for acute toxicity of water quality at present mainly contains physico-chemical method and biological method.Physico-chemical method can the analyzing water body of quantitative and qualitative analysis in kind and the concentration of pollutant, but cannot judge the direct relation between noxious material concentration and biological effect.In addition, physico-chemical method often need to be by instruments such as gas chromatography, high performance liquid chromatography and atomic spectrum, and analysis cost is high, monitoring velocity is slow, complex operation step.Biological method can reflect the impact of pollutant on biosome more all sidedly, for environmental risk assessment and the suitable emergency measure of selection provide foundation.At present bio-toxicity test is normal adopts fish, water flea, algae etc. to measure for biomaterial, but these methods exist the defects such as determination period length, complex operation.Nitrite bacteria is obligate aerobic bacteria, belongs to Gram-negative bacteria, ammoxidation can be generated to nitrite.Its metabolic activity is vulnerable to the impact of external environment, to pollutant sensitivities such as heavy metal, agricultural chemicals, organism, is often used in toxotest.Conventionally adopt nitrite bacteria is fixed on dissolved oxygen electrode, be exposed to the method for bacterium oxygen utilization variation before and after pollutant and evaluate the toxicity of pollutant by mensurations.But this method is vulnerable to the interference of other aerobic bacteria in sample matrices, and be difficult to avoid bacterium is fixed to the impact on bacterium and electrode generation on electrode.
Summary of the invention
The object of the invention is to provide for above-mentioned deficiency a kind of detection method and device thereof of acute toxicity of water quality.
For achieving the above object, the technical solution used in the present invention is:
A kind of detection method of acute toxicity of water quality, the filter membrane support and the ammonium ion selective electrode array that are fixed with nitrite bacteria are coupled in flow analysis system, first the system that sample passed into makes itself and bacterial action, then transfer medium, pass into the buffer solution containing ammonium ion to system, bacterial oxidation ammonium ion causes ammonium concentration to reduce, and records potential change by electrodes selective, can calculate the speed of bacterial oxidation ammonium ion according to potential difference (PD), and then the acute toxicity of definite water quality.
Detection method comprises:
(a) will contain 10
-6-10
-2the buffer solution of M ammonium chloride, by being in series with the fixing filter membrane support of nitrite bacteria and the flow analysis system of ammonium ion selective electrode, makes ammonium chloride and nitrite bacteria effect, the electric potential signal of generation signal in contrast;
(b) transfer medium, only passes into testing sample the filter membrane support of the fixing nitrite bacteria in flow analysis system, makes testing sample and bacterial action, then makes testing sample enter waste liquid pool by cross-over valve;
(c) with containing 10
-6-10
-2the buffer solution of M ammonium chloride, by being in series with the fixing filter membrane support of nitrite bacteria and the flow analysis system of ammonium ion selective electrode, makes ammonium chloride and nitrite bacteria effect, and the electric potential signal of generation is as sample signal;
(d) calculate ammoxidation speed according to control signal and sample signal, and then calculate inhibiting rate;
Inhibiting rate (%)=(contrast ammoxidation speed-sample ammoxidation speed)/contrast ammoxidation speed × 100%.
Describedly contain 10
-6-10
-2in the buffer solution of M ammonium chloride, damping fluid is for being Tris-HCl, and concentration is 0.01-1mol/L, pH7.8-8.0; Described peristaltic pump (4) is by computer control, and flow velocity is 0.1-10mL/min.
Flow analysis system comprises sample cell, transfer valve, peristaltic pump and detection cell; Wherein between sample cell and detection cell, pass through pipeline connection, pipeline between sample cell and detection cell is provided with two transfer valves of controlling flow of solution direction, and the pipeline between two transfer valves is provided with for carrying the peristaltic pump of moving flow of solution and the filter membrane support of fixing nitrite bacteria; Wherein on detection cell, be inserted with potential test device, and the other end is connected with the second wastewater disposal basin; Transfer valve before described detection cell is connected with the first wastewater disposal basin.
Described potential test device is made up of ionometer or electrochemical workstation, ammonium ion selective electrode and Ag/AgCl electrode; Wherein ionometer or electrochemical workstation connect respectively one end of ammonium ion selective electrode and Ag/AgCl electrode by wire; The other end of ammonium ion selective electrode and outer contrast electrode inserts in detection cell.
Described sample cell is two, is connected in parallel in the transfer valve before peristaltic pump.
Described ammonium ion selective electrode is made up of polyvinyl chloride pipe, internal-filling liquid and the internal reference electrode that inserts in wherein, and polyvinyl chloride pipe bottom is provided with sensitive membrane.
Described polymer sensitive membrane consist of ionophore, cationite, polymeric matrix material and plastifier, its ratio of weight and number is 0.1-10:0.1-10:20-40:40-80; Membrane component is dissolved in tetrahydrofuran, stirs, under room temperature, place 12-24h, after tetrahydrofuran volatilization, obtain polymer sensitive membrane;
Described polymeric matrix material is Polyvinylchloride, poly-butylacrylic acid ester, butyl polyacrylate, polyetherimide, rubber or sol-gel film; Plastifier is o-nitrobenzene octyl ether (o-NPOE), two-2-ethylhexyl ester in the last of the ten Heavenly stems, dibutyl sebacate or di-n-octyl sebacate; Ionophore is nonactin (nonactin) or monactin (monactin); Cationite is four (4-chlorphenyl) potassium borate or four (3,5-bis-(trifluoromethyl) phenyl) sodium borate.
The special purpose device of detection method, flow analysis system comprises sample cell, transfer valve, peristaltic pump and detection cell; Wherein between sample cell and detection cell, pass through pipeline connection, pipeline between sample cell and detection cell is provided with two transfer valves of controlling flow of solution direction, and the pipeline between two transfer valves is provided with for carrying the peristaltic pump of moving flow of solution and the filter membrane support of fixing nitrosomonas; Wherein on detection cell, be inserted with potential test device, and the other end is connected with the second wastewater disposal basin; Transfer valve before described detection cell is connected with the first wastewater disposal basin.
Described potential test device is made up of ionometer or electrochemical workstation, ammonium ion selective electrode and Ag/AgCl electrode; Wherein ionometer or electrochemical workstation connect respectively one end of ammonium ion selective electrode and Ag/AgCl electrode by wire; The other end of ammonium ion selective electrode and outer contrast electrode inserts in detection cell.
Action principle: nitrite bacteria is a kind of to the more sensitive microorganism of environmental change, is vulnerable to the pollutant effects such as organic compound, heavy metal in environment.Nitrite bacteria amine-oxides ion generates nitrite anions, and the variation of ammonium concentration can be recorded by ammonium ion selective electrode.In certain hour, the variable quantity of ammonium concentration can be used for characterizing the metabolic activity of bacterium.After bacterium and poisonous substance effect, metabolic activity reduces, and the amount that consumes ammonium ion reduces, and analyzes the toxicity size of poisonous substance according to the degree reducing.
Compared with prior art, tool has the following advantages this method:
1. what the inventive method was measured is the speed of nitrite bacteria amine-oxides ion, the interference of other aerobic bacteria can avoid utilizing oxygen electrode to measure dissolved oxygen DO time.
2. when working sample of the present invention, adopt change of medium method to measure, can avoid the interference of other ion pair ammonium ion selective electrodes that exist in sample, eliminated matrix effect, and guard electrode is avoided the stained of sample.
3. the inventive method is separated to nitrite bacteria and ion-selective electrode, avoided bacterium to be fixed to electrode surface and the impact on bacterial activity and electrode response that produces, makes bacterium and electrode separately in optimum condition; The operation of the inventive method simultaneously, by flow injection instruments control, easily is automated detection.
4. the inventive method is compared with toxicity test methods such as adopting fish, algae, have advantages of consuming time short, cost is low.
Brief description of the drawings book
The pick-up unit schematic diagram (wherein 3 is transfer valve, and 4 is peristaltic pump, and 5 is fixing germy filter membrane, and 8 is detection cell, and 9 is ammonium ion selective electrode, and 10 is Ag/AgCl electrode, and 11 is ionometer or electrochemical workstation) that Fig. 1 provides for the embodiment of the present invention.
The schematic diagram of electrode in the device that Fig. 2 provides for the embodiment of the present invention (wherein 13 is internal reference electrode, and 14 is internal-filling liquid, and 15 is polymer film, and 16 is polyvinyl chloride pipe).
The device that Fig. 3 provides for the embodiment of the present invention is measured response signal and the response curve of variable concentrations allylthiourea.
Embodiment
After the present invention utilizes nitrite bacteria to be subject to noxious material to suppress, degree that the speed of amine-oxides ion reduces is carried out oxicity analysis.Bacterium and electrodes selective are series in flow analysis system, first sample are passed into system and bacterial action a period of time, then transfer medium, passes into the buffer solution containing ammonium ion, and the speed of bacterial oxidation ammonium ion can be recorded by ammonium ion selective electrode.
Embodiment
Taking allylthiourea in this electrode test water as example.
As shown in Figure 1, flow analysis system comprises sample cell 2, transfer valve 3, peristaltic pump 4 and detection cell 8; Wherein between sample cell 2 and detection cell 8, pass through pipeline connection, pipeline between sample cell 2 and detection cell 8 is provided with two pipelines between 3, the first and second two transfer valves 3 of the first and second transfer valves of controlling flow of solution directions and is provided with for carrying the peristaltic pump 4 of moving flow of solution and a filter membrane support 5 for fixing nitrosomonas; Wherein on detection cell 8, be inserted with potential test device, and the other end is connected with the second wastewater disposal basin 12; The second transfer valve 3 before described detection cell 8 is connected with the first wastewater disposal basin 7.Described product sample cell 2 is two, is connected in parallel in the first transfer valve 3 before peristaltic pump 4.
Described potential test device forms by controlling potentiometer 11, ammonium ion selective electrode 9 and Ag/AgCl electrode 10; Wherein control potentiometer 11 connects respectively ammonium ion selective electrode 9 and Ag/AgCl electrode 10 one end by wire; The other end of ammonium ion selective electrode 9 and outer contrast electrode 10 inserts in detection cell 8.
The internal reference electrode of described ammonium ion selective electrode by polyvinyl chloride pipe, internal-filling liquid and among inserting in forms as shown in Figure 2, and polyvinyl chloride pipe bottom is provided with sensitive membrane.
Instrument: high-pressure sterilizing pot, aseptic operating platform, shaking table, vacuum pump, flow injection analyzer, ionometer.
Bacterial classification used: Nitrosomonas europaea (Nitrosomonas europaea, NBRC14298).
Liquid Culture based formulas: 18.90mM (NH
4)
2sO
4, 3.67mM KH
2pO
4, 5.95mM NaHCO
3, 400 μ M MgSO
47H
2o, 30 μ M CaCl
22H
2o, 180 μ M Fe-EDTA, and50mM HEPES.pH7.8-8.0.
Described nitrite bacteria can be replaced by bacterial strains such as Nitromonas (Nitrosomona), Nitrosospira (Nitrosospira), Nitrosolobus (Nitrosolobus), nitrosation vibrio (Nitrosovibrio) or Nitrosococcus (Nitrosococcus).Cultivate bacterial strain according to existing condition of culture simultaneously.
1. implementation step:
1) cultivation of bacterial classification: European nitrite bacteria original bacteria liquid is inoculated into fluid nutrient medium, and 28 DEG C, cultivate 3 days 120rpm dark place;
2) immobilization of bacterium: by the vacuum filtration of nitrite bacteria bacterium liquid to the polyethersulfone millipore filter of aperture 0.2 μ m, diameter 25mm, on thalline, cover again the miillpore filter that one deck is identical, bacterium is fixed in the middle of two-layer filter membrane, more fixing germy filter membrane is fixed on the filter membrane support of diameter 25mm;
3) buffer solution pond (6) are contained to 10
-4tris-HCl (0.05M, the pH8.0) buffer solution of M ammonium chloride passes through filter membrane support (5), the second cross-over valve (3) and the detection cell (8) of the first cross-over valve (3) in flow analysis system, peristaltic pump (4), fixing nitrosomonas successively with the flow velocity of 1.62mL/min, flow into waste liquid (12), turn on pump after termination of pumping 30min, the electric potential signal that records termination of pumping front and back, this is control signal;
4) by the sample that contains allylthiourea (1) successively by filter membrane support (5) and the second cross-over valve (3) of the first cross-over valve (3) in flow analysis system, peristaltic pump (4), fixing nitrosomonas, inflow waste liquid (7), then termination of pumping; After 30min, turn on pump, rotates the first transfer valve (3), contains 10 with buffer solution pond (6)
-4the buffer solution of M ammonium chloride rinses above-mentioned pipeline, rotates the second transfer valve (3), makes buffer solution enter detection cell (8), and turn on pump after termination of pumping 30min records the electric potential signal of termination of pumping front and back, and this is sample signal.
5) calculate inhibiting rate according to control signal and sample signal.
The preparation of ammonium ion selective electrode:
Take respectively 1.26mg nonactin, 0.31mg tetra-(4-chlorphenyl) potassium borate, 40.24mg Polyvinylchloride, 82.28mg o-nitrobenzene octyl ether, dissolve in 2mL tetrahydrofuran solution, stir, under room temperature, in flat small beaker (diameter 2.0cm), place 12h, volatilize completely after dry and obtain resilient polymer sensitive membrane until tetrahydrofuran.
The sensitive membrane that is 0.6cm with card punch cut-off footpath by described polymer sensitive membrane.The polymer sensitive membrane of preparation attaches to ion-selective electrode polyvinyl chloride pipe bottom by tetrahydrofuran.Electrode interior injects 10
-2the NH of M
4cl solution is as internal-filling liquid; Need be 10 before the first use of electrode
-2the NH of M
4in Cl solution, activate 12h.
2. the calculating of inhibiting rate:
Inhibiting rate adds the variation of allylthiourea front and back ammoxidation speed (AOR) to calculate by contrast.
Inhibiting rate (%)=(AOR
contrast-AOR
sample)/AOR
contrast× 100% (formula 1)
FA
0, FA
1and FA
ibe respectively the concentration of ammonia after concentration, bacterium and the carrier fluid effect a period of time (t) of original ammonia in carrier fluid and bacterial exposure after allyl with carrier fluid effect a period of time (t) after the concentration of ammonia.
Can be obtained by formula (1) (2) (3)
Free state ammonia is provided by ammonium ion hydrolysis:
The concentration of ammonium ion can calculate by Nernst equation.
Control signal:
Potential value before bacterium and carrier fluid effect:
Potential value after bacterium and carrier fluid effect 30min:
The potential difference (PD) of control experiment:
Bacterial exposure after allylthiourea, then with carrier fluid effect 30min after potential value:
Potential difference (PD) after bacterium is affected by allylthiourea:
Formula (5) substitution formula (9) (10)
Obtained by (formula 11) (formula 12):
FA
1=FA
010
-F Δ E/2.303RT(formula 13)
(formula 13) (formula 14) substitution (formula 4) is obtained:
In formula, F, R and T are respectively Faraday constant, gas law constant and absolute temperature.
Conventionally characterize its poisonous effect by the test-compound concentration (IC50,503nhibiting concentration) of anti-bacteria 50% metabolic activity.
Measuring concentration be the inhibiting effect of the allylthiourea of 0.05,0.1,0.2,0.3,0.5 μ M, the results are shown in Figure 3 and table 1.Be 0.17 μ M by the 503nhibiting concentration of Fig. 3 and the known allylthiourea of table 1, detect and be limited to 0.02 μ M.
Table 1
| Allyl sulfide urea concentration (μ M) | Potential change Δ E (mV) | Relative activity (%) | Inhibiting rate (%) |
| 0.00 | 17 | 100 | 0 |
| 0.05 | 13 | 81.1 | 18.9 |
| 0.10 | 10 | 67.2 | 32.8 |
| 0.20 | 7 | 44.9 | 55.1 |
| 0.30 | 4 | 30.1 | 69.9 |
| 0.50 | 2 | 13.5 | 86.5 |
Claims (9)
1. the detection method of an acute toxicity of water quality, it is characterized in that: the filter membrane support and the ammonium ion selective electrode array that are fixed with nitrite bacteria are coupled in flow analysis system, first the system that sample passed into makes itself and bacterial action, then transfer medium, pass into the buffer solution containing ammonium ion to system, bacterial oxidation ammonium ion causes ammonium concentration to reduce, record potential change by electrodes selective, can calculate the speed of bacterial oxidation ammonium ion according to potential difference (PD), and then the acute toxicity of definite water quality.
2. by the detection method of acute toxicity of water quality claimed in claim 1, it is characterized in that:
(a) will contain 10
-6-10
-2the buffer solution of M ammonium chloride, by being in series with the fixing filter membrane support of nitrite bacteria and the flow analysis system of ammonium ion selective electrode, makes ammonium chloride and nitrite bacteria effect, the electric potential signal of generation signal in contrast;
(b) transfer medium, only passes into testing sample the filter membrane support of the fixing nitrite bacteria in flow analysis system, makes testing sample and bacterial action, then makes testing sample enter waste liquid pool by cross-over valve;
(c) with containing 10
-6-10
-2the buffer solution of M ammonium chloride, by being in series with the fixing filter membrane support of nitrite bacteria and the flow analysis system of ammonium ion selective electrode, makes ammonium chloride and nitrite bacteria effect, and the electric potential signal of generation is as sample signal;
(d) calculate ammoxidation speed according to control signal and sample signal, and then calculate inhibiting rate;
Inhibiting rate (%)=(contrast ammoxidation speed-sample ammoxidation speed)/contrast ammoxidation speed × 100%.
3. by the detection method of acute toxicity of water quality claimed in claim 2, it is characterized in that: flow analysis system comprises sample cell (2), transfer valve (3), peristaltic pump (4) and detection cell (8); Wherein between sample cell (2) and detection cell (8), pass through pipeline connection, pipeline between sample cell (2) and detection cell (8) is provided with two transfer valves (3) of controlling flow of solution direction, and the pipeline between two transfer valves (3) is provided with for carrying the peristaltic pump (4) of moving flow of solution and the filter membrane support (5) of fixing nitrite bacteria; Wherein on detection cell (8), be inserted with potential test device, and the other end is connected with the second wastewater disposal basin (12); The front transfer valve (3) of described detection cell (8) is connected with the first wastewater disposal basin (7).
4. by the detection method of acute toxicity of water quality claimed in claim 3, it is characterized in that: described potential test device is made up of ionometer or electrochemical workstation (11), ammonium ion selective electrode (9) and Ag/AgCl electrode (10); Wherein ionometer or electrochemical workstation (11) connect respectively one end of ammonium ion selective electrode (9) and Ag/AgCl electrode (10) by wire; The other end of ammonium ion selective electrode (9) and outer contrast electrode (10) inserts in detection cell (8).
5. by the detection method of acute toxicity of water quality claimed in claim 3, it is characterized in that: described sample cell (2) is two, be connected in parallel in the front transfer valve (3) of peristaltic pump (4).
6. by the detection method of acute toxicity of water quality claimed in claim 3, it is characterized in that: described ammonium ion selective electrode is made up of polyvinyl chloride pipe, internal-filling liquid and the internal reference electrode that inserts in wherein, and polyvinyl chloride pipe bottom is provided with sensitive membrane.
7. by the detection method of acute toxicity of water quality claimed in claim 6, it is characterized in that: described polymer sensitive membrane consist of ionophore, cationite, polymeric matrix material and plastifier, its ratio of weight and number is 0.1-10:0.1-10:20-40:40-80; Membrane component is dissolved in tetrahydrofuran, stirs, under room temperature, place 12-24h, after tetrahydrofuran volatilization, obtain polymer sensitive membrane; Described polymeric matrix material is Polyvinylchloride, poly-butylacrylic acid ester, butyl polyacrylate, polyetherimide, rubber or sol-gel film; Plastifier is o-nitrobenzene octyl ether (o-NPOE), two-2-ethylhexyl ester in the last of the ten Heavenly stems, dibutyl sebacate or di-n-octyl sebacate; Ionophore is nonactin (nonactin) or monactin (monactin); Cationite is four (4-chlorphenyl) potassium borate or four (3,5-bis-(trifluoromethyl) phenyl) sodium borate.
8. a special purpose device for the detection method of acute toxicity of water quality claimed in claim 1, is characterized in that: flow analysis system comprises sample cell (2), transfer valve (3), peristaltic pump (4) and detection cell (8); Wherein between sample cell (2) and detection cell (8), pass through pipeline connection, pipeline between sample cell (2) and detection cell (8) is provided with two transfer valves (3) of controlling flow of solution direction, and the pipeline between two transfer valves (3) is provided with for carrying the peristaltic pump (4) of moving flow of solution and the filter membrane support (5) of fixing nitrosomonas; Wherein on detection cell (8), be inserted with potential test device, and the other end is connected with the second wastewater disposal basin (12); The front transfer valve (3) of described detection cell (8) is connected with the first wastewater disposal basin (7).
9. by the special purpose device of the detection method of acute toxicity of water quality claimed in claim 8, it is characterized in that: described potential test device is made up of ionometer or electrochemical workstation (11), ammonium ion selective electrode (9) and Ag/AgCl electrode (10); Wherein ionometer or electrochemical workstation (11) connect respectively one end of ammonium ion selective electrode (9) and Ag/AgCl electrode (10) by wire; The other end of ammonium ion selective electrode (9) and outer contrast electrode (10) inserts in detection cell (8).
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