CN114252416B - Fluorescence measurement method for nitrite - Google Patents
Fluorescence measurement method for nitrite Download PDFInfo
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- CN114252416B CN114252416B CN202010992779.5A CN202010992779A CN114252416B CN 114252416 B CN114252416 B CN 114252416B CN 202010992779 A CN202010992779 A CN 202010992779A CN 114252416 B CN114252416 B CN 114252416B
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- IOVCWXUNBOPUCH-UHFFFAOYSA-M Nitrite anion Chemical compound [O-]N=O IOVCWXUNBOPUCH-UHFFFAOYSA-M 0.000 title claims abstract description 101
- 238000000691 measurement method Methods 0.000 title abstract description 7
- 229920002873 Polyethylenimine Polymers 0.000 claims abstract description 105
- 229920000642 polymer Polymers 0.000 claims abstract description 104
- 239000007850 fluorescent dye Substances 0.000 claims abstract description 99
- 238000000034 method Methods 0.000 claims abstract description 29
- RWSXRVCMGQZWBV-WDSKDSINSA-N glutathione Chemical compound OC(=O)[C@@H](N)CCC(=O)N[C@@H](CS)C(=O)NCC(O)=O RWSXRVCMGQZWBV-WDSKDSINSA-N 0.000 claims abstract description 22
- 239000010842 industrial wastewater Substances 0.000 claims abstract description 20
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 19
- 229960003180 glutathione Drugs 0.000 claims abstract description 11
- 108010024636 Glutathione Proteins 0.000 claims abstract description 10
- 239000008399 tap water Substances 0.000 claims abstract description 9
- 235000020679 tap water Nutrition 0.000 claims abstract description 9
- 239000000243 solution Substances 0.000 claims description 46
- 239000007853 buffer solution Substances 0.000 claims description 23
- 230000005284 excitation Effects 0.000 claims description 17
- 239000000523 sample Substances 0.000 claims description 15
- 238000007865 diluting Methods 0.000 claims description 10
- 238000005259 measurement Methods 0.000 claims description 10
- 238000002156 mixing Methods 0.000 claims description 10
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 8
- 229910052799 carbon Inorganic materials 0.000 claims description 8
- 239000000706 filtrate Substances 0.000 claims description 8
- 239000012154 double-distilled water Substances 0.000 claims description 7
- 238000001816 cooling Methods 0.000 claims description 6
- -1 nitrite ions Chemical class 0.000 claims description 6
- 239000012488 sample solution Substances 0.000 claims description 6
- 238000002189 fluorescence spectrum Methods 0.000 claims description 5
- 238000002360 preparation method Methods 0.000 claims description 5
- 238000009210 therapy by ultrasound Methods 0.000 claims description 5
- 239000012153 distilled water Substances 0.000 claims description 4
- 238000001914 filtration Methods 0.000 claims description 4
- 239000012528 membrane Substances 0.000 claims description 4
- 239000002245 particle Substances 0.000 claims description 4
- 238000000967 suction filtration Methods 0.000 claims description 4
- 238000001471 micro-filtration Methods 0.000 claims description 3
- 229910021642 ultra pure water Inorganic materials 0.000 claims description 3
- 239000012498 ultrapure water Substances 0.000 claims description 3
- 238000010008 shearing Methods 0.000 claims description 2
- 238000001132 ultrasonic dispersion Methods 0.000 claims description 2
- 238000010791 quenching Methods 0.000 abstract description 12
- 230000000171 quenching effect Effects 0.000 abstract description 12
- 238000006243 chemical reaction Methods 0.000 abstract description 9
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- 230000008859 change Effects 0.000 abstract description 3
- 230000035945 sensitivity Effects 0.000 abstract description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 18
- 230000000694 effects Effects 0.000 description 12
- 238000001514 detection method Methods 0.000 description 10
- LYGJENNIWJXYER-UHFFFAOYSA-N nitromethane Chemical compound C[N+]([O-])=O LYGJENNIWJXYER-UHFFFAOYSA-N 0.000 description 8
- 239000011734 sodium Substances 0.000 description 8
- 239000011780 sodium chloride Substances 0.000 description 8
- LPXPTNMVRIOKMN-UHFFFAOYSA-M sodium nitrite Chemical compound [Na+].[O-]N=O LPXPTNMVRIOKMN-UHFFFAOYSA-M 0.000 description 8
- 239000007864 aqueous solution Substances 0.000 description 7
- 230000000873 masking effect Effects 0.000 description 6
- QGZKDVFQNNGYKY-UHFFFAOYSA-N ammonia Natural products N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 5
- 238000010438 heat treatment Methods 0.000 description 5
- 229910021645 metal ion Inorganic materials 0.000 description 5
- 239000000126 substance Substances 0.000 description 5
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 4
- 235000011114 ammonium hydroxide Nutrition 0.000 description 4
- 238000002474 experimental method Methods 0.000 description 4
- 238000005375 photometry Methods 0.000 description 4
- 235000010288 sodium nitrite Nutrition 0.000 description 4
- 239000012086 standard solution Substances 0.000 description 4
- VTLYFUHAOXGGBS-UHFFFAOYSA-N Fe3+ Chemical compound [Fe+3] VTLYFUHAOXGGBS-UHFFFAOYSA-N 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 3
- 230000002378 acidificating effect Effects 0.000 description 3
- 230000003197 catalytic effect Effects 0.000 description 3
- 238000011549 displacement method Methods 0.000 description 3
- 229910001447 ferric ion Inorganic materials 0.000 description 3
- 238000011068 loading method Methods 0.000 description 3
- 230000007246 mechanism Effects 0.000 description 3
- 229910052760 oxygen Inorganic materials 0.000 description 3
- 229910052938 sodium sulfate Inorganic materials 0.000 description 3
- 238000002798 spectrophotometry method Methods 0.000 description 3
- 241000251468 Actinopterygii Species 0.000 description 2
- 241000143060 Americamysis bahia Species 0.000 description 2
- 206010010356 Congenital anomaly Diseases 0.000 description 2
- 108010054147 Hemoglobins Proteins 0.000 description 2
- 102000001554 Hemoglobins Human genes 0.000 description 2
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 2
- 108010061951 Methemoglobin Proteins 0.000 description 2
- 239000007832 Na2SO4 Substances 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 2
- 239000012964 benzotriazole Substances 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000004737 colorimetric analysis Methods 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- 239000000975 dye Substances 0.000 description 2
- 238000000295 emission spectrum Methods 0.000 description 2
- 238000000695 excitation spectrum Methods 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 230000003993 interaction Effects 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- STZCRXQWRGQSJD-GEEYTBSJSA-M methyl orange Chemical compound [Na+].C1=CC(N(C)C)=CC=C1\N=N\C1=CC=C(S([O-])(=O)=O)C=C1 STZCRXQWRGQSJD-GEEYTBSJSA-M 0.000 description 2
- 229940012189 methyl orange Drugs 0.000 description 2
- 238000001000 micrograph Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
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- 239000001301 oxygen Substances 0.000 description 2
- 238000006479 redox reaction Methods 0.000 description 2
- 239000002351 wastewater Substances 0.000 description 2
- GEYOCULIXLDCMW-UHFFFAOYSA-N 1,2-phenylenediamine Chemical compound NC1=CC=CC=C1N GEYOCULIXLDCMW-UHFFFAOYSA-N 0.000 description 1
- 241000193155 Clostridium botulinum Species 0.000 description 1
- 208000032170 Congenital Abnormalities Diseases 0.000 description 1
- 208000000461 Esophageal Neoplasms Diseases 0.000 description 1
- CWYNVVGOOAEACU-UHFFFAOYSA-N Fe2+ Chemical compound [Fe+2] CWYNVVGOOAEACU-UHFFFAOYSA-N 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 208000005718 Stomach Neoplasms Diseases 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 229910021529 ammonia Inorganic materials 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- NEDCLUPMYCVKFO-UHFFFAOYSA-N azanium;4-aminobenzenesulfonate Chemical compound [NH4+].NC1=CC=C(S([O-])(=O)=O)C=C1 NEDCLUPMYCVKFO-UHFFFAOYSA-N 0.000 description 1
- 238000006149 azo coupling reaction Methods 0.000 description 1
- 235000013405 beer Nutrition 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 239000012472 biological sample Substances 0.000 description 1
- 230000007698 birth defect Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 230000000711 cancerogenic effect Effects 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 231100000315 carcinogenic Toxicity 0.000 description 1
- 238000006555 catalytic reaction Methods 0.000 description 1
- 210000003169 central nervous system Anatomy 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
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- 239000003344 environmental pollutant Substances 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 201000004101 esophageal cancer Diseases 0.000 description 1
- LECSHJWIACEDPZ-UHFFFAOYSA-N ethane-1,2-diamine naphthalene hydrochloride Chemical compound C(CN)N.C1=CC=CC2=CC=CC=C12.Cl LECSHJWIACEDPZ-UHFFFAOYSA-N 0.000 description 1
- 238000010812 external standard method Methods 0.000 description 1
- 229910001448 ferrous ion Inorganic materials 0.000 description 1
- 238000001917 fluorescence detection Methods 0.000 description 1
- 235000013373 food additive Nutrition 0.000 description 1
- 239000002778 food additive Substances 0.000 description 1
- 239000005452 food preservative Substances 0.000 description 1
- 235000019249 food preservative Nutrition 0.000 description 1
- 206010017758 gastric cancer Diseases 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 230000005802 health problem Effects 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 239000008235 industrial water Substances 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 230000036244 malformation Effects 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 235000013622 meat product Nutrition 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 150000004005 nitrosamines Chemical class 0.000 description 1
- 239000007800 oxidant agent Substances 0.000 description 1
- 230000001590 oxidative effect Effects 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- 231100000719 pollutant Toxicity 0.000 description 1
- VKJKEPKFPUWCAS-UHFFFAOYSA-M potassium chlorate Chemical compound [K+].[O-]Cl(=O)=O VKJKEPKFPUWCAS-UHFFFAOYSA-M 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 230000035484 reaction time Effects 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 125000000467 secondary amino group Chemical class [H]N([*:1])[*:2] 0.000 description 1
- 235000015170 shellfish Nutrition 0.000 description 1
- 235000011152 sodium sulphate Nutrition 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 201000011549 stomach cancer Diseases 0.000 description 1
- 150000003512 tertiary amines Chemical class 0.000 description 1
- 125000001302 tertiary amino group Chemical group 0.000 description 1
- 125000003396 thiol group Chemical group [H]S* 0.000 description 1
- 238000002211 ultraviolet spectrum Methods 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
- 230000003313 weakening effect Effects 0.000 description 1
Classifications
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
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- C—CHEMISTRY; METALLURGY
- C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
- C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
- C09K11/00—Luminescent, e.g. electroluminescent, chemiluminescent materials
- C09K11/06—Luminescent, e.g. electroluminescent, chemiluminescent materials containing organic luminescent materials
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
- G01N2021/6432—Quenching
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- Chemical & Material Sciences (AREA)
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- General Physics & Mathematics (AREA)
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- Life Sciences & Earth Sciences (AREA)
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- Pathology (AREA)
- Engineering & Computer Science (AREA)
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- Organic Chemistry (AREA)
- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
Abstract
The invention belongs to the technical field of fluorescence sensing, and particularly relates to a fluorescence measurement method of nitrite. The polymer point fluorescent probe is prepared by using the reaction of glutathione and polyethyleneimine, the fluorescence intensity of the polymer point fluorescent probe is in linear quenching relation with the change of nitrite concentration at the emission wavelength of 462nm, so that a standard working straight line is established, and the nitrite concentration in tap water and industrial wastewater can be measured. The method has the advantages of convenient operation, good specific selectivity, high sensitivity, quick response time and good accuracy, and can be used for measuring the concentration of nitrite in complex water industrial wastewater.
Description
Technical Field
The invention belongs to the technical field of fluorescence sensing, and particularly relates to a fluorescence measurement method of nitrite, in particular to a method for measuring nitrite by using a polymer dot fluorescent probe.
Background
In recent years, food safety problems and waste water pollutant discharge exceeding problems have attracted great attention, people are increasingly conscious of physical health, and the nitrite content has become very sensitive. Nitrite is a commonly used food additive in life, most commonly in various pickled vegetables and pickled meats, for color development and color protection. Nitrite can effectively inhibit clostridium botulinum and can be used as food preservative. However, the human body is greatly harmed by taking nitrite to a certain extent, and researches show that excessive nitrite in the human body can oxidize ferrous ions in blood into ferric ions to influence an oxygen transport system in the human body, so that normal hemoglobin is converted into methemoglobin, and the hemoglobin is irreversibly converted into methemoglobin in blood flow, thereby causing methemoglobia. In addition, after nitrite is taken in human body, the nitrite can react with secondary amine, tertiary amine and amino compounds to form strong cancerogenic substance nitrosamine compounds, which not only easily cause gastric cancer and esophagus cancer, but also are unfavorable for the treatment and recovery of cancer. Excessive nitrite can also cause health problems such as congenital malformations, central nervous system birth defects, etc. in infants. In addition, the excessive nitrite content in the water body can cause liver mutation of fishes and shrimps, and induce death of fish, shrimps and shellfish which are fulminant diseases of animals, so that the serious environmental pollution is caused. It is therefore important to establish an efficient, rapid and accurate method for detecting nitrite content in food, water samples and biological samples.
Wang Dan adopts ultraviolet spectrophotometry to measure the nitrite content in meat products, o-phenylenediamine is used as a color reagent, and forms a benzotriazole compound with nitrite in an acidic medium, the benzotriazole compound has a strong absorption peak at the position of 280nm in the ultraviolet region, the method is simple and quick to operate, is suitable for mass sample detection, but has a narrow detection range, and the mass concentration of nitrite is 5-250 mug/mL and obeys the beer law. The detection method commonly adopted by the national standard is a Gris reagent colorimetric method, which is essentially a visible spectrophotometry. Under the acidic condition, nitrite and ammonia sulfanilic acid are diazotized, coupled with naphthalene ethylenediamine hydrochloride, and the total content of nitrite is measured by an external standard method. However, this method has not only the influence of the pigment of the sample itself, but also the Cu 2+、Fe3+ plasma interferes with the measurement, which is inconvenient. The catalytic photometry is a dynamic photometry analysis method, has simpler instrument and equipment, higher sensitivity, wide application in early stage and quick development. However, the method has no fundamental breakthrough point so far, and most of the methods adopt an oxidizing agent such as dissolved oxygen, hydrogen peroxide, potassium chlorate and the like to oxidize the organic dye in an acidic medium, so that the nitrite is measured according to the decoloring speed of the organic dye. Kuang Chun et al, mixing NO 2 - with KIO 3 and methyl orange at room temperature in a strong acid medium, found that methyl orange was rapidly oxidized and discolored and that the rate of catalytic reaction was linearly related to the concentration of NO 2 - over a range of concentrations. Electrochemical methods for determining nitrite are not yet mature and are yet to be further investigated. The existing ultraviolet spectrophotometry, diazo coupling colorimetry (Griess method), catalytic photometry and the like have low nitrite detection range, are suitable for detecting actual samples with low nitrite content, and are more required to be detected in industrial wastewater with high nitrite content and environmental wastewater. In recent years, the fluorescence photometry is based on the catalytic action of nitrite on oxidation-reduction reaction, so that the fluorescence intensity of an indicator substance is weakened or quenched, the weakening or quenching of the intensity is in linear relation with the concentration of nitrite in a certain range, and the nitrite content is measured by measuring the change of the fluorescence intensity. In the fluorescence detection method, the selection of an appropriate fluorescent probe to detect nitrite is a key factor.
Disclosure of Invention
In order to solve the technical problem of detecting nitrite in the prior art, the invention provides a fluorescence measurement method of nitrite, and particularly prepares a polymer point fluorescence probe, and uses the polymer point fluorescence probe for measuring nitrite.
The technical scheme of the invention is as follows:
Dissolving Glutathione (GSH) and Polyethyleneimine (PEI) in water, uniformly mixing, heating for reaction, and cooling to obtain a polymer dot G-PEI PDs fluorescent probe solution; and adding the polymer point G-PEI PDS fluorescent probe solution into a sample to be detected to measure the fluorescence intensity, and obtaining the concentration of nitrite in the sample to be detected according to the relationship between the concentration of nitrite and a fluorescence spectrum.
In particular the number of the elements to be processed,
The water is preferably double distilled water, triple distilled water, ultrapure water or the like, which does not affect nitrite measurement.
The mixing mode is selected from any one of ultrasonic dispersion, a mechanical stirrer, a high-speed shearing dispersion machine or a homogenizer.
Further, the method comprises the steps of,
The mass ratio of the glutathione to the polyethyleneimine is 0.3 (0.8-1.2);
The heating reaction temperature is 200-250 ℃, and the reaction time is 3-5h.
Further, the method comprises the steps of,
The fluorescence intensity value of the polymer point G-PEI PDS fluorescent probe at the emission wavelength of 462nm is F 0, the fluorescence intensity value after nitrite is added is F, the fluorescence intensity ratio I F＝F0/F, the concentration of nitrite in a sample to be detected is obtained according to the linear quenching relation between the I F value and the concentration of nitrite, and the used excitation wavelength is 340nm.
Further, the method comprises the steps of,
The concentration of nitrite is in the range of 0.02-2.2mg/mL, the I F value and the concentration of nitrite are in linear quenching relation, and the detection limit of nitrite is 0.0032mg/mL.
Further, the method comprises the steps of,
According to the fluorescence measurement method, through the linear relation between the change of a system fluorescence signal and the concentration of nitrite ions, whether nitrite is contained in a sample or the concentration of nitrite is quantitatively detected can be rapidly and qualitatively judged by a tap water sample or industrial wastewater (such as industrial wastewater produced by a nitrite displacement method).
Further, the method comprises the steps of,
The fluorescence measurement method of nitrite comprises the following steps:
(1) Preparation of a Polymer Point G-PEI PDS fluorescent probe:
Dissolving glutathione and polyethylenimine in water, uniformly mixing, reacting for 3-5 hours at 200-250 ℃, and naturally cooling to room temperature to obtain a polymer dot G-PEI PDS fluorescent probe solution;
(2) Drawing a standard working straight line:
taking a proper amount of the polymer point G-PEI PDS fluorescent probe solution prepared in the step (1), dissolving the polymer point G-PEI PDS fluorescent probe solution in a buffer solution with pH=2-10, and detecting a fluorescence intensity value F 0 of the G-PEI-PD polymer point fluorescent probe under the excitation of 340nm at the emission wavelength of 462nm after constant volume;
Taking a proper amount of the polymer point G-PEI PDS fluorescent probe solution prepared in the step (1), dissolving the polymer point G-PEI PDS fluorescent probe solution in a buffer solution with pH=2-10, adding nitrite ions into the solution, after constant volume, enabling the concentration of nitrite to be 0.02-2.2mg/mL, detecting a fluorescence intensity value F at an emission wavelength of 462nm under excitation of 340nm, establishing a standard working straight line according to the relation between the concentration of nitrite and a fluorescence spectrum, and taking a regression equation of the ratio I F of the fluorescence intensity and the concentration c of nitrite: i F = 2.75338c +0.89781, linear correlation coefficient R = 0.99013 (n = 10), detection limit D = 0.0032mg/mL, (see example 4);
(3) Determination of nitrite in a sample to be tested:
And (3) dissolving a proper amount of the polymer dot G-PEI PDS fluorescent probe solution prepared in the step (1) in a buffer solution with pH=2-10, adding a sample solution to be detected into the solution, detecting the fluorescence intensity value of the sample solution to be detected under the excitation of 340nm and the emission wavelength of 462nm after the volume is fixed, and calculating the concentration of nitrite in the sample solution to be detected according to the regression equation described in the step (2).
Further, the buffer solution preferably has a ph=2 to 6.5.
Further, the method comprises the steps of,
Step (3) pretreatment is carried out before the measurement of the sample to be measured,
The pretreatment comprises the following steps of:
A. tap water is directly measured without pretreatment;
B. Decolorizing industrial wastewater by using active carbon, performing ultrasonic treatment, standing, performing suction filtration, removing active carbon particles, filtering the filtrate by using a microfiltration membrane, and properly diluting the filtrate for later use.
Further, the method comprises the steps of,
In the steps (2) and (3), 1mL of the polymer spot G-PEI PDS fluorescent probe solution prepared in the step (1) is diluted 10 times, and 200 mu L of the polymer spot G-PEI PDS fluorescent probe solution is taken;
The buffer solution is B-R buffer solution, the concentration is 40mmol/L, the pH=6.0, and 625 mu L of buffer solution is taken;
the volume is fixed to 5.0mL in the steps (2) and (3).
The invention also provides application of the polymer dot G-PEI PDS fluorescent probe in nitrite determination. In examples 6,7 and 8, nitrite in tap water or industrial water can be determined using the polymer dot G-PEI PDS fluorescent probe prepared in accordance with the present invention.
Advantageous effects
Compared with the prior art, the invention has the beneficial effects that:
(1) The polymer dot fluorescent probe is simple to synthesize;
(2) The method for measuring nitrite by adopting the polymer dot fluorescent probe has the advantages of low cost, simple detection method, good specific selectivity, high sensitivity, quick response time and wider linear range;
(3) The method for measuring the polymer dot fluorescent probe can be used for measuring nitrite in tap water, even complex water industrial wastewater with higher nitrite content.
Additional features and advantages of the invention will be set forth in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. The objectives and other advantages of the invention will be realized and attained by the structure particularly pointed out in the written description and claims hereof as well as the appended drawings.
The technical scheme of the invention is further described in detail through the drawings and the embodiments.
Drawings
In order to more clearly illustrate the embodiments of the invention or the technical solutions in the prior art, the drawings that are required in the embodiments or the description of the prior art will be briefly described, it being obvious that the drawings in the following description are only some embodiments of the invention, and that other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a scanning electron microscope image of a polymer dot G-PEI PDS fluorescent probe;
FIG. 2 is an infrared spectrogram of a polymer dot G-PEI PDS fluorescent probe;
FIG. 3 is a graph of fluorescence excitation and emission spectra of a polymer dot G-PEI PDS fluorescent probe;
FIG. 4 is an ultraviolet spectrum of a polymer dot G-PEI PDS fluorescent probe;
FIG. 5 is a graph of fluorescence intensity of a polymer spot G-PEI PDS fluorescent probe at different pH values;
FIG. 6 is a Stern-Volmer plot of the interaction of nitrite with a polymer site G-PEI PDS fluorescent probe at various concentrations;
FIG. 7 is a standard operating line graph of fluorescence intensity ratio F 0/F versus nitrite concentration, where c is nitrite concentration and R is a linear correlation coefficient;
FIG. 8 is a graph showing fluorescence emission spectra of polymer spot G-PEI PDS fluorescent probes at nitrite concentrations of 0,0.02,0.03,0.05,0.08,0.3,0.5,0.8,1.2,1.5,1.8,2.0,2.2 mg/mL;
FIG. 9 is a graph showing the response of various metal ions and nitrite in water to a polymer point G-PEI PDS fluorescent probe;
FIG. 10 is a graph showing the effect of masking Hg 2+ with aqueous ammonia on nitrite measurement;
FIG. 11 is a graph showing the effect of different mass percentages Na 2SO4 on the fluorescence intensity of a polymer dot G-PEI PDS fluorescent probe;
FIG. 12 is a graph showing the effect of different volume percentages of CH 3 OH on the fluorescence intensity of a polymer dot G-PEI PDS fluorescent probe;
FIG. 13 is a graph showing the effect of different mass percentages of NaCl on the fluorescence intensity of a polymer dot G-PEI PDS fluorescent probe;
FIG. 14 is a graph of fluorescence intensity of a polymer spot G-PEI PDS fluorescent probe with the addition of various interferents in the presence of quencher nitrite.
Wherein,
6,7,9, 10, 11, 12 And 13, wherein F 0 is the fluorescence intensity value of the polymer dot G-PEI PDS fluorescent probe, and F is the fluorescence intensity value of the polymer dot G-PEI PDS fluorescent probe after different substances are added;
FIG. 14F 0' is the fluorescence intensity value of sodium nitrite after adding to the polymer site G-PEI PDS fluorescent probe;
f' is the fluorescence intensity value of the sodium nitrite added to the polymer point G-PEI PDS fluorescent probe and then added with different substances.
Detailed Description
Other advantages and effects of the present invention will become apparent to those skilled in the art from the following disclosure, which is to be read by way of example only. The invention may be practiced or carried out in other embodiments that depart from the specific details, and the details of the present description may be modified or varied from the spirit and scope of the present invention.
Before the embodiments of the invention are explained in further detail, it is to be understood that the invention is not limited in its scope to the particular embodiments described below; it is also to be understood that the terminology used in the examples of the invention is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the invention.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In addition to the specific methods, devices, materials used in the embodiments, any methods, devices, and materials of the prior art similar or equivalent to those described in the embodiments of the present invention may be used to practice the present invention according to the knowledge of one skilled in the art and the description of the present invention.
Example 1:
Preparation of a Polymer Point G-PEI PDS fluorescent probe:
Taking 1mL (0.123G/mL) of glutathione aqueous solution and 1mL (0.4G/mL) of polyethyleneimine aqueous solution, diluting to 40mL by ultrapure water, carrying out ultrasonic treatment for 5min, loading into a reaction kettle, heating at 200 ℃ for reaction for 4 hours, and naturally cooling to room temperature to obtain a light yellow polymer dot G-PEI PDS fluorescent probe solution.
As can be seen from a scanning electron microscope image (shown in figure 1) of the polymer dot G-PEI PDs fluorescent probe, the polymer dot G-PEI PDs are in a hexagonal sheet shape and uniformly dispersed in an aqueous solution.
As can be seen from an infrared spectrogram (shown in fig. 2) of the polymer point G-PEI PDS fluorescent probe, the polymer point G-PEI PDS has obvious peaks at 1465cm- 1,1575cm-1,3123cm-1,3411cm-1, and represents the stretching vibration of C=O, N-H and O-H respectively, which indicates that the surface of the polymer point G-PEI PDS fluorescent probe possibly contains carboxyl, hydroxyl and amino. Since G-PEI PDs have sulfhydryl groups in glutathione and are reducing, while nitrite has oxidizing, the presumed reaction mechanism is that G-PEI PDs and nitrite undergo oxidation-reduction reaction, thereby causing fluorescence quenching of G-PEI PDs, and thus the concentration of nitrite can be detected by using G-PEI PDs as fluorescent probes.
As can be seen from the fluorescence excitation and emission spectra of the polymer spot G-PEI PDs fluorescent probe (as shown in FIG. 3), the fluorescence excitation wavelength of the G-PEI PDs is 340nm, and the emission wavelength is 462nm.
In the ultraviolet spectrogram (as shown in FIG. 4) of the polymer dot G-PEI PDS fluorescent probe, the maximum ultraviolet absorption wavelength of the G-PEI PDS is 340nm.
In the fluorescence intensity diagram (shown in fig. 5) of the G-PEI PDs under different pH values, the fluorescence intensity of the polymer points is gradually increased when the pH value is between 2.0 and 6.0, and the fluorescence intensity of the polymer points is basically unchanged after the decrease when the pH value is between 6.0 and 11.0.
Example 2:
quenching mechanism experiment:
To explore the effect of temperature on the quenching process, the Stern-Volmer curve (F 0/F) was studied with different concentrations of nitrite interacting with the polymer site G-PEI PDS fluorescent probe for 5min at different temperatures in B-R buffer at pH 6.0, 293.15K,313.15K, 333.15K. As shown in fig. 6, results F 0/F＝1+KSV c were obtained by the stem-Volmer equation, F0 and F represent the fluorescence intensity of the polymer spot G-PEI PDs before and after addition of nitrite as quencher, respectively, K SV represents the quenching constant, F 0/F has a good linear fit to nitrite at temperatures of 293.15K,313.15K and 333.15K, respectively, and K SV increases with increasing temperature, indicating that the dynamic quenching mechanism plays a major role in quenching, and the results are shown in table 1.
TABLE 1 Stern-volumer equation for interaction of Polymer Point G-PEI PDS fluorescent probes with nitrite at three different temperatures and quenching constant Ksv
Example 3:
Preparation of a Polymer Point G-PEI PDS fluorescent probe:
Taking 1mL (0.123G/mL) of glutathione aqueous solution and 0.82mL (0.4G/mL) of polyethyleneimine aqueous solution, diluting to 40mL by triple distilled water, homogenizing for 2min at 3000rpm, loading into a reaction kettle, heating at 250 ℃ for reaction for 3 hours, and naturally cooling to room temperature to obtain a light yellow polymer dot G-PEI PDS fluorescent probe solution.
Example 4:
Preparation of a Polymer Point G-PEI PDS fluorescent probe:
Taking 1mL (0.123G/mL) of glutathione aqueous solution and 1.23mL (0.4G/mL) of polyethyleneimine aqueous solution, diluting to 40mL by triple distilled water, stirring for 8min, loading into a reaction kettle, heating and reacting for 5 hours at 220 ℃, and naturally cooling to room temperature to obtain a light yellow polymer dot G-PEI PDS fluorescent probe solution.
Example 5:
Drawing a nitrite standard working line:
Taking 1mL of the polymer spot G-PEI PDS fluorescent probe solution prepared in the embodiment 1, diluting by 10 times, taking 200 mu L of the polymer spot G-PEI PDS fluorescent probe solution, mixing with 625 mu L of B-R buffer solution with the concentration of 40mmol/L and the pH=6.0 at room temperature, fixing the volume to 5.0mL, and detecting the fluorescence intensity value F 0 of the G-PEI-PD polymer spot fluorescent probe with the emission wavelength of 462nm under excitation of 340 nm;
Taking 1mL of the polymer spot G-PEI PDS fluorescent probe solution prepared in the example 1, diluting the solution by 10 times, taking 200 mu L of the solution and 625 mu L of B-R buffer solution with the concentration of 40mmol/L and pH=6.0, mixing the solution at room temperature, then adding 0 mu L,10 mu L,15 mu L,25 mu L,40 mu L,150 mu L,250 mu L,400 mu L,600 mu L,750 mu L,900 mu L,1000 mu L and 1100 mu L of sodium nitrite standard solution respectively, fixing the volume to 5.0mL by double distilled water, detecting the fluorescence intensity value F of all standard solutions with the emission wavelength of 462nm by using a fluorescence spectrophotometer under excitation of 340nm, taking the ratio of the fluorescence intensity I F and the concentration c of nitrite as a regression equation;
IF＝2.75338c+0.89781，
wherein I F is the ratio of fluorescence intensity, c is the concentration of nitrite, and the linear correlation coefficient r= 0.99013 (n=10), as shown in fig. 7, the correlation coefficient approaching 1 indicates that the linear relationship of the regression equation is better, the detection limit d=0.0032 mg/mL, and the linear range of nitrite measurement is 0.02-2.2mg/mL.
FIG. 8 is a graph showing fluorescence emission spectra of polymer spot G-PEI PDS fluorescent probes at nitrite standard solution concentrations of 0,0.02,0.03,0.05,0.08,0.3,0.5,0.8,1.2,1.5,1.8,2.0,2.2mg/mL, respectively.
Example 6:
determination of tap water:
1mL of the polymer spot G-PEI PDS fluorescent probe solution prepared in example 1 is diluted 10 times, 200 mu L of the polymer spot G-PEI PDS fluorescent probe solution is mixed with 625 mu L of B-R buffer solution with the concentration of 40mmol/L and the pH value of 6.0 at room temperature, 1mL of tap water is added to be fixed to 5.0mL by double distilled water, the fluorescence intensity value F at the emission wavelength of 462nm is detected by a fluorescence spectrophotometer under the excitation of 340nm, and the concentration of nitrite in the tap water is calculated according to the standard working straight line drawn in example 4, so that the nitrite concentration is too low or absent and is not detected in the detection range.
Example 7:
Determination of nitromethane industrial wastewater No. 1 containing nitrite:
Adding 3.0g of active carbon into No. 1 industrial wastewater produced by a nitrite displacement method for decoloring, standing for 5 hours in ultrasonic for 15min, removing active carbon large particles by a suction filtration bottle, filtering the filtrate by a 0.45 mu m microfiltration membrane, and diluting the filtrate for 6 times for later use;
1mL of the polymer spot G-PEI PDS fluorescent probe solution prepared in example 1 is diluted 10 times, 200 mu L of the polymer spot G-PEI PDS fluorescent probe solution is mixed with 625 mu L of B-R buffer solution with the concentration of 40mmol/L and the pH of 6.0 at room temperature, 50 mu L of industrial wastewater No. 1 which is subjected to pretreatment dilution 6 times is added, the volume is fixed to 5.0mL by double distilled water, after ultrasonic treatment is carried out for 10min, under the excitation of 340nm, the fluorescence intensity value F at the emission wavelength of 462nm is detected by a fluorescence spectrophotometer, and the concentration of nitrite in the industrial wastewater No. 1 is calculated to be 82.8mg/mL according to the standard working straight line drawn in example 4.
Example 8:
determination of nitromethane industrial wastewater No. 2 containing nitrite:
Adding 3.0g of active carbon into No.2 industrial wastewater produced by a nitrite displacement method for decoloring, standing for 5 hours in ultrasonic for 20min, removing active carbon large particles by a suction filtration bottle, filtering the filtrate by a 0.45 mu m filter membrane, and diluting the filtrate by 5 times for later use;
1mL of the polymer spot G-PEI PDS fluorescent probe solution prepared in example 1 is diluted 10 times, 200 mu L of the polymer spot G-PEI PDS fluorescent probe solution is mixed with 625 mu L of B-R buffer solution with the concentration of 40mmol/L and the pH of 6.0 at room temperature, 50 mu L of industrial wastewater No.2 which is diluted 5 times through pretreatment is added, the volume is fixed to 5.0mL by double distilled water, after ultrasonic treatment is carried out for 10min, under the excitation of 340nm, the fluorescence intensity value F at the emission wavelength of 462nm is detected by a fluorescence spectrophotometer, and the concentration of nitrite in industrial wastewater No.2 is calculated to be 57.5mg/mL according to a standard working straight line drawn in example 4.
Example 9:
response experiment of various metal ions and nitrite to polymer point G-PEI PDS fluorescent probes:
Taking 1mL of the polymer spot G-PEI PDS fluorescent probe solution prepared in the embodiment 1, diluting by 10 times, taking 200 mu L of the polymer spot G-PEI PDS fluorescent probe solution, mixing with 625 mu L of B-R buffer solution with the concentration of 40mmol/L and the pH=6.0 at room temperature, fixing the volume to 5.0mL, and detecting the fluorescence intensity value F 0 of the polymer spot fluorescent probe of the G-PEI PDS under the excitation of 340nm at the emission wavelength of 462 nm;
1mL of the polymer spot G-PEI PDS fluorescent probe solution prepared in example 1 was diluted 10 times, 200. Mu.L of the polymer spot G-PEI PDS fluorescent probe solution was mixed with 625. Mu.L of a B-R buffer solution having a concentration of 40mmol/L and a pH=6.0 at room temperature, and then Ag+,Pb2+,Ba2+,Mg2+,Co2+,Fe3+,Mn2+,Ca2+,Cr3+,Cr6+,Hg2+,NO2 - was added to the mixture to fix the volume to 5.0mL with double distilled water, wherein the concentrations of metal ions were 40ng/mL and the concentration of NO 2 - was 2.1mg/mL, respectively. The fluorescence intensity values F of all standard solutions at the emission wavelength of 462nm were detected with a fluorescence spectrophotometer under 340nm excitation. As shown in FIG. 9, blank is used as a Blank contrast, only contains a polymer dot G-PEI PDS fluorescent probe, NO 2 - has obvious response to the polymer dot G-PEI PDS fluorescent probe in the presence of a buffer solution, and most metal ions do not respond to the polymer dot G-PEI PDS fluorescent probe in the presence of the buffer solution, so that the stability of the polymer dot G-PEI PDS fluorescent probe of various metal ions in the presence of the buffer solution is good. Hg 2+ suppresses fluorescence to some extent, and through further experimentation we have found that fluorescence quenched by Hg 2+ can be eliminated by the addition of a masking agent, such as ammonia.
FIG. 10 is a graph showing the effect of masking Hg 2+ with aqueous ammonia on nitrite measurement; wherein,
The concentration of sodium nitrite is 0.3mg/mL, the concentration of Hg 2+ is 40ng/mL, the concentration of ammonia water is 1.1mmol/L,
Polymer Point G-PEI PDS fluorescent Probe 1mL of the polymer Point G-PEI PDS fluorescent probe solution prepared in example 1 was diluted 10 times, and 200. Mu.L was further prepared.
Wherein,
1 Is the experimental result of a fluorescent probe containing only polymer spots G-PEI PDs;
2 is the experimental result of adding NaNO 2 containing Hg 2+ into the polymer dot G-PEI PDS fluorescent probe;
3 is the experimental result of measuring NaNO 2 by NH 4 OH masking Hg 2+.
As can be seen from fig. 10, the masking of Hg 2+ with aqueous ammonia did not affect the determination of nitrite.
In performing industrial sample measurements, samples containing Hg 2+ can be measured after masking with NH 4 OH.
Example 10:
interference experiments of Na 2SO4,CH3 OH, naCl and the like:
The Blank in FIGS. 11, 12 and 13 is compared with the fluorescent probe of example 9 containing only polymer spots G-PEI PDs,
Different mass percentages of Na 2SO4 (e.g., 1%: 0.01G Na 2SO4 in 1G water) were added to the polymer point G-PEI PDS fluorescent probe solution, as shown in FIG. 11, the different mass percentages of Na 2SO4 had little effect on the fluorescence intensity of the polymer point G-PEI PDS fluorescent probe.
Different volume percentages of CH 3 OH (e.g., 1% to 1mL of water containing 0.01mL of CH 3 OH) were added to the polymer dot G-PEI PDS fluorescent probe solution, as shown in FIG. 12, a small volume percentage of CH 3 OH had little effect on the fluorescent intensity of the polymer dot G-PEI PDS fluorescent probe.
Different mass percentages of NaCl in the water (e.g., 5% to 1G of water containing 0.05G of NaCl) were added to the polymer spot G-PEI PDS fluorescent probe solution, as shown in FIG. 13, with little effect on the fluorescent intensity of the polymer spot G-PEI PDS fluorescent probe.
The industrial wastewater produced by the nitrite replacement method contains Na 2SO4,CH3 OH, naCl and a small amount of nitromethane (CH 3NO2), does not contain Hg 2+, and can not consider quenching of Hg 2+ on fluorescence. The small amount of nitromethane has little influence on the fluorescence intensity of the polymer dot G-PEI PDS fluorescent probe. For the industrial wastewater of nitromethane, when nitrite is measured by an actual sample, the industrial wastewater of nitromethane is diluted to reach the detection range of 0.02-2.2mg/mL of nitrite detected by a polymer point G-PEI PDS fluorescent probe, at the moment, the concentration of the diluted nitromethane is far less than 0.02mg/mL, and the effect of the diluted nitromethane on the polymer point G-PEI PDS fluorescent probe is negligible, so that the measurement of nitrite is not influenced. The dilution concentration of other substances is correspondingly reduced, so that the measurement of nitrite ions is not interfered.
Example 11:
Mixing interference experiment:
When the quencher nitrite exists, sodium sulfate, sodium chloride and various interferents of methanol are respectively added to measure the influence of nitrite on the polymer point G-PEI PDs fluorescent probe, as shown in figure 14, the influence of various interferents on the G-PEI PDs fluorescent intensity is small, which indicates that the polymer point G-PEI PDs fluorescent probe has good stability.
Wherein,
Blank 0 was used as a Blank for comparison, and the experimental result of 1mg/mL NaNO 2 was added to the Blank of example 9;
NaCl is the experimental result of Blank 0 +0.01g NaCl;
Na 2SO4 is the experimental result of Blank 0+0.0125g Na2SO4;
CH 3 OH is the experimental result of Blank 0+0.003mL CH3 OH;
NaCl is mixed to be the experimental result of Blank 0+0.01g NaCl+0.003mL CH3 OH;
Na 2SO4 was mixed as the experimental result of Blank 0+0.0125g Na2SO4+0.003mL CH3 OH.
It will be apparent to those skilled in the art that various modifications and variations can be made to the present invention without departing from the spirit or scope of the invention. Thus, it is intended that the present invention also include such modifications and alterations insofar as they come within the scope of the appended claims or the equivalents thereof.
Claims (6)
1. A method for fluorometric determination of nitrite comprising the steps of:
(1) Preparation of Polymer Point G-PEIPDs fluorescent Probe:
Dissolving glutathione and polyethylenimine in water, uniformly mixing, reacting for 3-5 hours at 200-250 ℃, and naturally cooling to room temperature to obtain a polymer point G-PEIPDs fluorescent probe solution;
The mass ratio of the glutathione to the polyethyleneimine is 0.3 (0.8-1.2);
(2) Drawing a standard working straight line:
Taking a proper amount of the polymer spot G-PEIPDs fluorescent probe solution prepared in the step (1), dissolving the polymer spot G-PEIPDs fluorescent probe solution in a buffer solution with pH=2-10, and detecting a fluorescence intensity value F 0 of the polymer spot G-PEI-PD fluorescent probe with an emission wavelength of 462nm under excitation of 340nm after constant volume;
Taking a proper amount of the polymer spot G-PEI PDS fluorescent probe solution prepared in the step (1), dissolving the polymer spot G-PEI PDS fluorescent probe solution in a buffer solution with pH=2-10, adding nitrite ions into the solution, after constant volume, enabling the concentration of nitrite to be 0.02-2.2mg/mL, detecting a fluorescence intensity value F at an emission wavelength of 462nm under excitation of 340nm, establishing a standard working straight line according to the relation between the concentration of nitrite and a fluorescence spectrum, and taking a regression equation of the fluorescence intensity ratio I F and the concentration c of nitrite;
(3) Determination of nitrite in a sample to be tested:
And (3) dissolving a proper amount of the polymer dot G-PEI PDS fluorescent probe solution prepared in the step (1) in a buffer solution with pH=2-10, adding a sample solution to be detected into the solution, detecting the fluorescence intensity value of the sample solution to be detected under the excitation of 340nm and the emission wavelength of 462nm after the volume is fixed, and calculating the concentration of nitrite in the sample solution to be detected according to the regression equation described in the step (2).
2. The fluorometric method of claim 1, wherein the water is selected from any one of double distilled water, triple distilled water, and ultra pure water.
3. The method according to claim 2, wherein the mixing means is selected from any one of ultrasonic dispersion, mechanical stirrer, high-speed shearing disperser and homogenizer.
4. The method according to claim 1, wherein the step (3) is performed with pretreatment before the measurement of the sample to be measured, and the pretreatment is selected from the following steps, respectively, depending on the samples:
A, directly measuring tap water without pretreatment;
And B, decolorizing industrial wastewater by using active carbon, performing ultrasonic treatment, standing, performing suction filtration, removing active carbon particles, filtering the filtrate by using a microfiltration membrane, and properly diluting the filtrate for later use.
5. A fluorometric method according to claim 4, characterized in that,
In the steps (2) and (3), 1mL of the polymer spot G-PEI PDS fluorescent probe solution prepared in the step (1) is diluted 10 times, and 200 mu L of the polymer spot G-PEI PDS fluorescent probe solution is taken;
The buffer solution is B-R buffer solution, the concentration is 40mmol/L, the pH=6.0, and 625 mu L of buffer solution is taken; the volume is fixed to 5.0mL in the steps (2) and (3).
6. The fluorometric method of claim 1, wherein the fluorometric method is suitable for the determination of nitrite in complex water industrial wastewater.
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