CN110026562B - Synthesis method and application of near-infrared fluorescent probe iron nanocluster - Google Patents
Synthesis method and application of near-infrared fluorescent probe iron nanocluster Download PDFInfo
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- CN110026562B CN110026562B CN201810028871.2A CN201810028871A CN110026562B CN 110026562 B CN110026562 B CN 110026562B CN 201810028871 A CN201810028871 A CN 201810028871A CN 110026562 B CN110026562 B CN 110026562B
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- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 title claims abstract description 157
- 229910052742 iron Inorganic materials 0.000 title claims abstract description 78
- 239000007850 fluorescent dye Substances 0.000 title claims abstract description 15
- 238000001308 synthesis method Methods 0.000 title abstract description 4
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims abstract description 30
- 229940098773 bovine serum albumin Drugs 0.000 claims abstract description 30
- NWZSZGALRFJKBT-KNIFDHDWSA-N (2s)-2,6-diaminohexanoic acid;(2s)-2-hydroxybutanedioic acid Chemical compound OC(=O)[C@@H](O)CC(O)=O.NCCCC[C@H](N)C(O)=O NWZSZGALRFJKBT-KNIFDHDWSA-N 0.000 claims abstract description 19
- IKDUDTNKRLTJSI-UHFFFAOYSA-N hydrazine monohydrate Substances O.NN IKDUDTNKRLTJSI-UHFFFAOYSA-N 0.000 claims abstract description 19
- 229910021578 Iron(III) chloride Inorganic materials 0.000 claims abstract description 15
- RBTARNINKXHZNM-UHFFFAOYSA-K iron trichloride Chemical compound Cl[Fe](Cl)Cl RBTARNINKXHZNM-UHFFFAOYSA-K 0.000 claims abstract description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 11
- 230000008859 change Effects 0.000 claims abstract description 4
- 238000006243 chemical reaction Methods 0.000 claims abstract description 4
- JPVYNHNXODAKFH-UHFFFAOYSA-N Cu2+ Chemical compound [Cu+2] JPVYNHNXODAKFH-UHFFFAOYSA-N 0.000 claims description 46
- 238000001514 detection method Methods 0.000 claims description 41
- 229910001431 copper ion Inorganic materials 0.000 claims description 36
- 239000012498 ultrapure water Substances 0.000 claims description 30
- 239000000243 solution Substances 0.000 claims description 24
- 239000000523 sample Substances 0.000 claims description 22
- 239000006185 dispersion Substances 0.000 claims description 13
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- 239000000017 hydrogel Substances 0.000 claims description 9
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- 238000002189 fluorescence spectrum Methods 0.000 claims description 3
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- 230000035945 sensitivity Effects 0.000 abstract description 4
- 239000010949 copper Substances 0.000 description 26
- 241000282414 Homo sapiens Species 0.000 description 9
- 239000012086 standard solution Substances 0.000 description 8
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 7
- 229910052802 copper Inorganic materials 0.000 description 7
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 description 5
- 238000000502 dialysis Methods 0.000 description 4
- 229910052751 metal Inorganic materials 0.000 description 4
- 239000002184 metal Substances 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- 238000010791 quenching Methods 0.000 description 4
- 229910052737 gold Inorganic materials 0.000 description 3
- 239000010931 gold Substances 0.000 description 3
- 230000036541 health Effects 0.000 description 3
- 238000002372 labelling Methods 0.000 description 3
- 229910052697 platinum Inorganic materials 0.000 description 3
- 229910052709 silver Inorganic materials 0.000 description 3
- 229910021591 Copper(I) chloride Inorganic materials 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 206010067125 Liver injury Diseases 0.000 description 2
- BQCADISMDOOEFD-UHFFFAOYSA-N Silver Chemical compound [Ag] BQCADISMDOOEFD-UHFFFAOYSA-N 0.000 description 2
- 238000003917 TEM image Methods 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- OXBLHERUFWYNTN-UHFFFAOYSA-M copper(I) chloride Chemical compound [Cu]Cl OXBLHERUFWYNTN-UHFFFAOYSA-M 0.000 description 2
- ORTQZVOHEJQUHG-UHFFFAOYSA-L copper(II) chloride Chemical compound Cl[Cu]Cl ORTQZVOHEJQUHG-UHFFFAOYSA-L 0.000 description 2
- 230000006378 damage Effects 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 239000003651 drinking water Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000003344 environmental pollutant Substances 0.000 description 2
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 2
- 229910001385 heavy metal Inorganic materials 0.000 description 2
- 231100000234 hepatic damage Toxicity 0.000 description 2
- 210000003734 kidney Anatomy 0.000 description 2
- 230000008818 liver damage Effects 0.000 description 2
- 239000011259 mixed solution Substances 0.000 description 2
- 230000004770 neurodegeneration Effects 0.000 description 2
- 208000015122 neurodegenerative disease Diseases 0.000 description 2
- 231100000252 nontoxic Toxicity 0.000 description 2
- 230000003000 nontoxic effect Effects 0.000 description 2
- 230000000171 quenching effect Effects 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- 239000004332 silver Substances 0.000 description 2
- 208000024827 Alzheimer disease Diseases 0.000 description 1
- 229910021592 Copper(II) chloride Inorganic materials 0.000 description 1
- 102000000634 Cytochrome c oxidase subunit IV Human genes 0.000 description 1
- 108090000365 Cytochrome-c oxidases Proteins 0.000 description 1
- 208000002972 Hepatolenticular Degeneration Diseases 0.000 description 1
- WTDRDQBEARUVNC-LURJTMIESA-N L-DOPA Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C(O)=C1 WTDRDQBEARUVNC-LURJTMIESA-N 0.000 description 1
- 102000019197 Superoxide Dismutase Human genes 0.000 description 1
- 108010012715 Superoxide dismutase Proteins 0.000 description 1
- 102000003425 Tyrosinase Human genes 0.000 description 1
- 108060008724 Tyrosinase Proteins 0.000 description 1
- 208000018839 Wilson disease Diseases 0.000 description 1
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 1
- 230000002159 abnormal effect Effects 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 125000003277 amino group Chemical group 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000031018 biological processes and functions Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 235000020188 drinking water Nutrition 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000000295 emission spectrum Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 230000037406 food intake Effects 0.000 description 1
- 125000000524 functional group Chemical group 0.000 description 1
- XLYOFNOQVPJJNP-ZSJDYOACSA-N heavy water Substances [2H]O[2H] XLYOFNOQVPJJNP-ZSJDYOACSA-N 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 238000011065 in-situ storage Methods 0.000 description 1
- 239000010842 industrial wastewater Substances 0.000 description 1
- FBAFATDZDUQKNH-UHFFFAOYSA-M iron chloride Chemical compound [Cl-].[Fe] FBAFATDZDUQKNH-UHFFFAOYSA-M 0.000 description 1
- -1 iron ions Chemical class 0.000 description 1
- 229910021645 metal ion Inorganic materials 0.000 description 1
- 150000002739 metals Chemical class 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 239000010413 mother solution Substances 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 230000036542 oxidative stress Effects 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 238000011946 reduction process Methods 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 239000004065 semiconductor Substances 0.000 description 1
- 239000010944 silver (metal) Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000008399 tap water Substances 0.000 description 1
- 235000020679 tap water Nutrition 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
- 239000002351 wastewater Substances 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
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- B22F9/00—Making metallic powder or suspensions thereof
- B22F9/16—Making metallic powder or suspensions thereof using chemical processes
- B22F9/18—Making metallic powder or suspensions thereof using chemical processes with reduction of metal compounds
- B22F9/24—Making metallic powder or suspensions thereof using chemical processes with reduction of metal compounds starting from liquid metal compounds, e.g. solutions
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- C09K11/60—Luminescent, e.g. electroluminescent, chemiluminescent materials containing inorganic luminescent materials containing iron, cobalt or nickel
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- 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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Abstract
The invention discloses a synthesis method and application of near-infrared fluorescent probe iron nanoclusters, which comprises the steps of synthesizing fluorescent iron nanoclusters under the condition of room-temperature water phase by using bovine serum albumin as a template and a protective agent, using ferric chloride as an iron source and hydrazine hydrate as a reducing agent, then using the fluorescent iron nanoclusters as near-infrared fluorescent probes, and utilizing fluorescence intensity change to carry out fluorescence intensity reaction on Cu in a solution at room temperature2+The content of (A) is detected simply, rapidly, with high sensitivity and high selectivity.
Description
Technical Field
The invention belongs to the technical field of biological analysis and detection, and relates to a green synthesis method of near-infrared fluorescent probe iron nanoclusters and a method for detecting Cu in actual samples by using the prepared iron nanoclusters2+The use of (1).
Background
The metal nanoclusters are photoluminescent semiconductor nanoclusters, and mainly comprise gold nanoclusters, silver nanoclusters, platinum nanoclusters, copper nanoclusters and the like. The metal nanoclusters are fluorescent water-soluble molecular aggregates consisting of several to dozens of atoms of metals such as Au, Ag, Pt and the like. Compared with gold nanoclusters, silver nanoclusters and platinum nanoclusters, iron nanoclusters have the advantages of being rich in resources, low in cost, capable of being widely applied to enterprises and the like. The diameter of the iron nanocluster is about 2nm, and since the iron nanocluster is discovered, the iron nanocluster has unique fluorescence property, is non-toxic, small and uniform in size and good in water solubility, and is widely applied to detection of various negative ions, heavy metal ions and small biological molecules. However, iron nanoclusters are of less interest in analytical detection.
Following zinc and iron, copper is the third most abundant metal element in the human body and plays a vital role in a wide range of biological processes. Copper is used as a cofactor of several important biological enzymes in human bodies, such as cytochrome c oxidase, superoxide dismutase, dopa beta-hydroxylase, tyrosinase and the like, and plays an important role in regulating the health of human beings. However, if the concentration of copper ions in human body is too high, it will inhibit the activity of some essential enzymes in human body, and make the relevant biological oxidation or reduction process abnormal, thereby causing liver damage, kidney damage and neurodegenerative diseases, such as Wilson's disease, Alzheimer's disease and oxidative stress related diseases, which pose a great threat to human health. The main way for human body to absorb copper element is Cu in food and drinking water2+Therefore, the Cu content of these substances must be strictly controlled2+In an amount to ensure food and water safety, the U.S. national Environmental Protection Agency (EPA) sets a safety limit for copper ions in drinking water of 1.3ppm (about 20. mu.M). However, with the rapid development of modern industry, industrial wastewater and waste contain a large amount of heavy metal ions Cu2+If the emission is improper, the ecological environment can be seriously affected, and then the human health is threatened. Therefore, the development of quantitative detection of Cu in water2+Various techniques for ions are extremely necessary.
Among the many metal ions, the problem of environmental pollution caused by copper ions is particularly prominent, and the ingestion of excessive copper into the human body causes serious side effects such as liver damage, kidney damage and neurodegenerative diseases. The copper-containing wastewater can generate great toxicity to aquatic organisms after being discharged into a water body. The project aims to prepare the iron nanocluster with high sensitivity, low detection limit and good selectivity, and the iron nanocluster is used for detecting and analyzing environmental pollutants, namely copper ions.
Disclosure of Invention
Objects of the inventionAims to overcome the defects of the prior art, provides a method for synthesizing a fluorescent iron nano-cluster which is water-soluble, non-toxic, synthesized at room temperature, easy to store and capable of emitting near infrared, and utilizes the change of fluorescence intensity to Cu in a solution at room temperature2+The content of (A) is detected simply, rapidly, with high sensitivity and high selectivity.
The technical purpose of the invention is realized by the following technical scheme: using Bovine Serum Albumin (BSA) as a template and a protective agent, using ferric chloride as an iron source, using hydrazine hydrate as a reducing agent, synthesizing fluorescent iron nano-clusters (BSA-Fe NCs) under the condition of a room-temperature water phase, and using the fluorescent iron nano-clusters as probes to react with Cu in a solution by using fluorescence intensity change2+The content of (b) is detected.
A near-infrared fluorescence probe-based divalent copper ion detection method for iron nanoclusters includes the steps of forming a divalent copper ion detection system by high-purity water, an iron nanocluster dispersion system and a sample to be detected, detecting fluorescence intensity changes before and after the sample to be detected is added, comparing standard curves to obtain the content of divalent copper ions in the sample to be detected, wherein a linear equation is F0/F＝1.00+100.15C，F0The fluorescence intensity detection value is the fluorescence intensity detection value when no bivalent copper ions are added, F is the fluorescence intensity detection value after the bivalent copper ions are added, and C is the bivalent copper ion concentration, namely the concentration of the bivalent copper ions in the sample to be detected.
In the above detection method, the linear range of detection is 1.0 × 10-4－1.0×10-2M, detection limit of 7.23 × 10- 5M。
In the detection method, 2.8mL of high-purity water, 1.0mL of iron nano-cluster dispersion system and 0.2mL of sample to be detected form 4.0mL of divalent copper ion detection system.
In the detection method, after a sample to be detected is added into the high-purity water and iron nano-cluster dispersion system, the reaction is carried out for 50-70 s, and then the fluorescence intensity is detected.
The iron nano-cluster dispersion system, namely the near-infrared fluorescent probe iron nano-cluster, is prepared by synthesizing a fluorescent iron nano-cluster under the condition of room temperature water phase by taking bovine serum albumin as a template and a protective agent, taking ferric chloride as an iron source and hydrazine hydrate as a reducing agent according to the following steps:
step 1, weighing 60-120 mg of bovine serum albumin, and uniformly dispersing the bovine serum albumin in a beaker filled with 8-10 mL of high-purity water;
in step 1, 80-100 mg of bovine serum albumin was weighed and uniformly dispersed in a beaker containing 10mL of high purity water.
Step 2, weighing FeCl3·6H2O is uniformly dispersed in high-purity water to form 0.1M ferric chloride aqueous solution;
step 3, dripping 80-100 mu L of the ferric chloride aqueous solution synthesized in the step 2 into the bovine serum albumin aqueous solution prepared in the step 1 at the room temperature of 20-25 ℃, and changing the solution into a hydrogel state from clarification; and dropwise adding 300-1000 mu L of hydrazine hydrate into the hydrogel, gradually dissolving the hydrogel into a yellow transparent clear solution, adding pure water until the whole system is 40-50 mL, and continuously stirring at room temperature for reacting for 2-5h to obtain the iron nano-cluster dispersion system.
In step 3, the hydrazine hydrate is an aqueous solution of hydrazine hydrate, and the mass percentage of the hydrazine hydrate is 80%.
In the step 3, the dropping speed of the hydrazine hydrate is 0.05-0.1mL/min, and the adding speed of the high-purity water is 0.5-1 mL/s. The purification condition is that high-purity water is changed every 4h, dialysis is carried out for 24h, and the purified iron nanoclusters are dried at 35 ℃ under the vacuum condition after dialysis is finished.
After preparation, purifying the iron nanoclusters prepared in the step 3 by using a dialysis bag with the molecular weight cutoff of 6000-8000 and the diameter of 25mm, changing high-purity water once every 3-5h, dialyzing for 20-30h, and drying the purified iron nanoclusters at 30-40 ℃ under a vacuum condition after dialysis is finished to obtain the near-infrared fluorescent probe iron nanoclusters, wherein the maximum excitation wavelength is 284nm, and the maximum emission wavelength is 664 nm.
The particle size of the prepared iron nanoclusters is concentrated to be 1.8-2 nm, and the particle size of the synthesized copper nanoclusters is uniform and is distributed uniformly; the iron nanoclusters are distributed on a bovine serum albumin matrix, namely iron ions are coordinated with functional groups such as amino groups and hydroxyl groups on the surface of protein, and in-situ reduction is realized under the action of hydrazine hydrate to form the iron nanoclusters.
The application of the iron nanocluster dispersion system prepared by the invention in detection of divalent copper ions, Cu2+After the iron nano-cluster fully reacts with the iron nano-cluster, the fluorescence of the system is quenched and the fluorescence emission spectrum is detected, and the Cu nano-cluster dispersion system can be used for detecting the Cu through the change value of the fluorescence emission spectrum intensity2+Detection of (3).
In the detection, the copper chloride-related solution is prepared as follows:
(1) preparing a copper chloride mother solution: 1.705g of CuCl were weighed2·2H2O, dissolving in 100mL volumetric flask with high-purity water to prepare 0.1M CuCl2And (4) storing the high-grade solution for later use.
(2) Preparing a copper ion standard solution:
removing a series of CuCl with different volumes2Diluting the high-standard solution in a 10mL colorimetric tube to prepare the high-standard solution with the concentration of 1.0 multiplied by 10 in sequence-4M、5.0×10-4M、1.0×10-3M、2.0×10-3M、6.0×10-3M、8.0×10-3M、1.0×10-2M of copper ion standard solution.
Compared with the prior art, the method utilizes Bovine Serum Albumin (BSA) as a template and a protective agent, ferric chloride as an iron source and hydrazine hydrate as a reducing agent to synthesize the fluorescent iron nanoclusters (BSA-Fe NCs) under the condition of room-temperature water phase, and the fluorescent iron nanoclusters are used as probes to react with Cu in a solution by utilizing fluorescence intensity change2+The content of (A) is detected simply, rapidly, with high sensitivity and high selectivity.
Drawings
Fig. 1 is a Transmission Electron Micrograph (TEM) of iron nanoclusters based on bovine serum albumin as a template.
FIG. 2 is a fluorescent emission spectrum of iron nanoclusters based on bovine serum albumin as a template.
FIG. 3 is a diagram of the detection of Cu in iron nanoclusters based on bovine serum albumin as a template2+Linear range diagram of (c).
Detailed Description
The foregoing features and advantages will become more apparent and be readily understood from the following further description of the present invention, taken in conjunction with the accompanying specific embodiments. All the reagents used were analytically pure, and the reagents and manufacturers used were as follows: bovine serum albumin, beijing dingguoshang biotechnology limited; hydrazine hydrate, Tianjin Guangfu Fine chemical Co., Ltd; iron chloride (99%), Tianjin Guangfu Fine chemical Co., Ltd.
Example 1
0.1M FeCl3Preparing a solution: 1.3515g of FeCl were weighed3Dissolving in a proper amount of high-purity water, transferring to a 50mL volumetric flask for constant volume, and labeling for later use;
preparing bovine serum albumin solution: weighing 0.1000g of bovine serum albumin, dissolving in 10mL of high-purity water, and fully dissolving under stirring;
synthesizing iron nanoclusters: adding prepared FeCl into the bovine serum albumin solution under the stirring condition3And adding 300 mu L of hydrazine hydrate into the mixed solution after the solution is fully reacted, and reacting for 5 hours under the stirring condition to obtain the fluorescent iron nano cluster.
Example 2
0.1M FeCl3Preparing a solution: 1.3515g of FeCl were weighed3Dissolving in a proper amount of high-purity water, transferring to a 50mL volumetric flask for constant volume, and labeling for later use;
preparing bovine serum albumin solution: weighing 0.1000g of bovine serum albumin, dissolving in 10mL of high-purity water, and fully dissolving under stirring;
synthesizing iron nanoclusters: adding prepared FeCl into the bovine serum albumin solution under the stirring condition3And adding 500 mu L of hydrazine hydrate into the mixed solution after the solution is 90 mu L and fully reacts, and reacting for 5h under the stirring condition to obtain the fluorescent iron nano cluster.
Example 3
Cu2+Detection of (2): respectively taking 2 empty centrifuge tubes, numbering the centrifuge tubes, respectively transferring 2.8mL of high-purity water into the centrifuge tubes, transferring 1.0mL of iron nanocluster solution into the centrifuge tubes, uniformly mixing, continuously adding 200 mu L of high-purity water into the centrifuge tube, taking the blank as a blank control group, and adding 200 mu L of Cu into the centrifuge tube2+And (3) reacting for 1min to quench fluorescence and detect fluorescence emission intensity by using a fluorescence photometer, so that detection of the divalent copper ions can be realized.
Example 4
Cu2+Detection of (2): respectively taking 2 empty centrifuge tubes, numbering the centrifuge tubes, transferring 2.6mL of high-purity water into the centrifuge tubes, transferring 1.2mL of iron nanocluster solution into the centrifuge tubes, mixing uniformly, continuously adding 200 mu L of high-purity water into the centrifuge tube, taking the blank as a blank control group, adding 200 mu L of Cu into the centrifuge tube, and adding 200 mu L of Cu into the centrifuge tube2+And (3) reacting for 1min to quench fluorescence and detect fluorescence emission intensity by using a fluorescence photometer, so that detection of the divalent copper ions can be realized.
Example 5 determination of Linear detection Range for detection of divalent copper ions
Adding 2.8mL of high-purity water into a 4mL centrifuge tube, adding 1.0mL of iron nanocluster solution, uniformly mixing, and respectively adding 0.2mL of Cu with different concentrations2+The fluorescence intensity of the standard solution was measured after 1min of reaction and measured in triplicate. According to different concentrations of Cu2+Quenching degree of fluorescence of the iron nanocluster to obtain the iron nanocluster and Cu2+The linear relation between the two is taken as the basis for carrying out quantitative detection on the copper ions, and the linear range of the detection is 1.0 multiplied by 10-4－1.0×10-2M, detection limit of 7.23 × 10- 5M。
EXAMPLE 6 detection treatment of actual samples
(1) Treatment of the actual sample: three tap water samples with the volume of 4.0mL are taken, and a certain volume of Cu with known concentration is added into each sample respectively2+Standard solution (the volume of the standard solution added is very small) so that Cu in the sample after the standard solution is added2+The concentration of (A) is as follows in sequence: 5.0X 10-4M、1.0×10-3M、2.0×10-3M, labeling the prepared solution for later use;
2. taking 4 empty centrifuge tubes, numbering firstly, secondly, thirdly and fourthly, respectively transferring 2.8mL of high-purity water into the centrifuge tubes firstly, secondly, thirdly and fourthly, then transferring 1.0mL of iron nano-cluster solution into the centrifuge tubes, mixing uniformly, continuing to add 200 mu L of high-purity water into the centrifuge tube firstly as a blank control group, and adding 200 mu L of Cu into the centrifuge tube secondly2+Has a concentration of 5.0X 10-4Practical sample of MAdding 200 mu L of Cu into No. three centrifugal tubes2+Has a concentration of 1.0X 10-3M into a sample tube, 200. mu.L of Cu was added2+Has a concentration of 2.0X 10-3M, reacting for 1min, measuring the quenching degree of the fluorescence of the iron nano-cluster, and calculating the recovery rate, as shown in the following table:
wherein the spiked concentration is the concentration of cupric ions in the system to be tested (i.e., the concentration determined from sample preparation), Cu2+The detection concentration is the concentration of the divalent copper ions obtained by detection according to the scheme of the invention, and the recovery rate is the ratio of the divalent copper ions to the divalent copper ions.
The application of the high-sensitivity fluorescent iron nano-cluster probe in selective detection of environmental pollutants is funded by a project of 'big creation plan' of Tianjin university (No.201522), a project on national science fund surface 21375095, a project of Qingjin's national science fund youth (No.17JCQNJC05800) and a project of Tianjin teacher's university doctor fund (No. 52XB1510).
By adopting the technical parameters of the invention for adjustment, the preparation of the iron nanocluster can be realized, and the detection of the divalent copper ions can be realized. The invention has been described in an illustrative manner, and it is to be understood that any simple variations, modifications or other equivalent changes which can be made by one skilled in the art without departing from the spirit of the invention fall within the scope of the invention.
Claims (6)
1. The application of the near-infrared fluorescent probe iron nanocluster in detecting bivalent copper ions is characterized in that Cu2+After the iron nano-cluster is fully reacted, the system fluorescence is quenched and the fluorescence emission light is detectedSpectrum, namely the change value of the fluorescence emission spectrum intensity can realize the iron nano-cluster dispersion system to Cu2+Detecting; in the near-infrared fluorescent probe iron nanoclusters, the particle size of the iron nanoclusters is concentrated to be 1.8-2 nm, the iron nanoclusters are distributed on a bovine serum albumin matrix, the maximum excitation wavelength is 284nm, and the maximum emission wavelength is 664 nm; the preparation method comprises the following steps:
step 1, weighing 60-120 mg of bovine serum albumin, and uniformly dispersing the bovine serum albumin in a beaker filled with 8-10 mL of high-purity water;
step 2, weighing FeCl3·6H2O is uniformly dispersed in high-purity water to form 0.1M ferric chloride aqueous solution;
step 3, dripping 80-100 mu L of the ferric chloride aqueous solution synthesized in the step 2 into the bovine serum albumin aqueous solution prepared in the step 1 at the room temperature of 20-25 ℃, and changing the solution into a hydrogel state from clarification; and dropwise adding 300-1000 mu L of hydrazine hydrate into the hydrogel, gradually dissolving the hydrogel into a yellow transparent clear solution, adding pure water until the whole system is 40-50 mL, and continuously stirring at room temperature for reacting for 2-5h to obtain the iron nano-cluster dispersion system.
2. The application of the near-infrared fluorescent probe iron nanocluster in detecting divalent copper ions according to claim 1, wherein 80-100 mg of bovine serum albumin is weighed and uniformly dispersed in a beaker filled with 10mL of high purity water in step 1.
3. The application of the near-infrared fluorescent probe iron nanocluster in detecting divalent copper ions according to claim 1, wherein in step 3, hydrazine hydrate is an aqueous solution of hydrazine hydrate, and the mass percent of the hydrazine hydrate is 80%; the dropping speed of hydrazine hydrate is 0.05-0.1mL/min, and the adding speed of high-purity water is 0.5-1 mL/s.
4. The method for detecting the divalent copper ions based on the near-infrared fluorescent probe iron nanoclusters is characterized in that a divalent copper ion detection system is formed by high-purity water, an iron nanocluster dispersion system and a sample to be detected, and the divalent copper ion detection system is used for detecting the divalent copper ions before and after the sample to be detected is addedThe fluorescence intensity changes, and the standard curve is compared to obtain the content of the divalent copper ions in the sample to be detected, wherein the linear equation is F0/F is 1.00+100.15C, F0 is the fluorescence intensity detection value when the divalent copper ions are not added, F is the fluorescence intensity detection value after the divalent copper ions are added, and C is the concentration of the divalent copper ions, namely the concentration of the divalent copper ions in the sample to be detected; the linear range of detection is 1.0 × 10-4－1.0×10-2M, detection limit of 7.23 × 10-5M; in the near-infrared fluorescent probe iron nanoclusters, the particle size of the iron nanoclusters is concentrated to be 1.8-2 nm, the iron nanoclusters are distributed on a bovine serum albumin matrix, the maximum excitation wavelength is 284nm, and the maximum emission wavelength is 664 nm; the preparation method comprises the following steps:
step 1, weighing 60-120 mg of bovine serum albumin, and uniformly dispersing the bovine serum albumin in a beaker filled with 8-10 mL of high-purity water;
step 2, weighing FeCl3·6H2O is uniformly dispersed in high-purity water to form 0.1M ferric chloride aqueous solution;
step 3, dripping 80-100 mu L of the ferric chloride aqueous solution synthesized in the step 2 into the bovine serum albumin aqueous solution prepared in the step 1 at the room temperature of 20-25 ℃, and changing the solution into a hydrogel state from clarification; and dropwise adding 300-1000 mu L of hydrazine hydrate into the hydrogel, gradually dissolving the hydrogel into a yellow transparent clear solution, adding pure water until the whole system is 40-50 mL, and continuously stirring at room temperature for reacting for 2-5h to obtain the iron nano-cluster dispersion system.
5. The method for detecting the cupric ions based on the near-infrared fluorescent probe iron nanoclusters as claimed in claim 4, wherein 2.8mL of high-purity water, 1.0mL of iron nanocluster dispersion system and 0.2mL of sample to be detected constitute 4.0mL of cupric ion detection system.
6. The method for detecting divalent copper ions based on near-infrared fluorescent probe iron nanoclusters according to claim 4, wherein the fluorescence intensity is detected after reaction for 50-70 s after a sample to be detected is added to the high-purity water and iron nanocluster dispersion system.
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| CN104985193A (en) * | 2015-07-24 | 2015-10-21 | 天津大学 | Alloy nano-particle preparation method based on protein reduction method |
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