CN111521589A - High-fluorescence copper nanocluster-cerium (III) fluorescent probe and preparation method and application thereof - Google Patents
High-fluorescence copper nanocluster-cerium (III) fluorescent probe and preparation method and application thereof Download PDFInfo
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- CN111521589A CN111521589A CN202010363237.1A CN202010363237A CN111521589A CN 111521589 A CN111521589 A CN 111521589A CN 202010363237 A CN202010363237 A CN 202010363237A CN 111521589 A CN111521589 A CN 111521589A
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- 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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- 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
- G01N2021/6417—Spectrofluorimetric devices
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- G—PHYSICS
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- 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
Abstract
The invention discloses a high-fluorescence copper nanocluster-cerium (III) fluorescent probe as well as a preparation method and application thereof, and belongs to the technical field of fluorescence sensing. The method is used for constructing the fluorescent sensor of hydrogen peroxide and glucose based on the phenomenon that hydrogen peroxide can linearly quench the fluorescence of the copper nanocluster-cerium (III) fluorescent probe. The method for enhancing the fluorescence characteristic of the copper nanocluster stabilized by the glutathione by using the metal cation cerium (III) is simple to operate, has obvious fluorescence enhancement degree, has the characteristics of high response speed and wide linear range when being used for detecting the hydrogen peroxide and the glucose, and can realize the visual detection of the hydrogen peroxide and the glucose.
Description
Technical Field
The invention belongs to the technical field of fluorescence sensing, and particularly relates to a high-fluorescence copper nanocluster-cerium (III) fluorescent probe, a preparation method thereof and application thereof in detection of hydrogen peroxide and glucose.
Background
Diabetes is characterized clinically by persistent hyperglycemia, and can impair or fail the functions of various tissues and organs (eyes, kidneys, heart, etc.). Due to the lack of effective treatment, complications can only be reduced by monitoring and tightly controlling blood glucose concentration in real time. Hydrogen peroxide is a representative of oxygen active species, and excessive hydrogen peroxide in the body causes various biological damages such as aging, alzheimer's disease, neurodegeneration, and even cancer. Therefore, the development of highly sensitive, highly selective, rapid glucose and hydrogen peroxide detection methods is crucial to the prevention, diagnosis and monitoring of diseases. The fluorescence sensor method has the advantages of high sensitivity, good selectivity, high response speed, simple operation and the like, and is widely researched and developed.
The metal nano-cluster is composed of several to hundreds of metal atoms, has excellent light stability and large Stokes shift, is favorable for improving the sensitivity of the fluorescence sensor and avoids the interference of background fluorescence and exciting light. Compared with gold nanoclusters and silver nanoclusters, researchers are dedicated to researching and preparing copper nanoclusters with different emission wavelengths and good fluorescence characteristics and application thereof due to the fact that precursor raw materials for preparing the copper nanoclusters are rich and low in price. Although the fluorescence sensor based on the copper nanocluster shows good sensitivity and selectivity, the prepared copper nanocluster has poor luminous performance and low fluorescence quantum yield (generally less than 10%), which is not beneficial to the practical application and development of the copper nanocluster.
Aggregation-induced emission refers to a phenomenon that light emission is remarkably enhanced after aggregation of molecules which do not emit light or emit light weakly in a solution, and is discovered by the team of Thanksgiving academicians in Tang and Dynasty in 2001. The aggregation-induced emission is that the fluorescence intensity is enhanced because the movement in the molecule is limited in an aggregation state, so that the problem of low application efficiency of the fluorescent probe is solved, the background can be reduced, and the signal-to-noise ratio and the sensitivity can be improved. Zinc ions, aluminum ions and the like are commonly used for inducing the aggregation induction effect of the copper nanocluster, so that the fluorescence of the copper nanocluster is obviously improved. However, the cerium (III) ion is used as an inducer to promote the aggregation induction effect of the copper nanocluster to prepare the high-fluorescence copper nanocluster-cerium (III) fluorescent probe, and the high-fluorescence copper nanocluster-cerium (III) fluorescent probe is used for constructing a hydrogen peroxide and glucose fluorescent sensor, so that the visible detection of hydrogen peroxide and glucose is realized, but no report is found.
Disclosure of Invention
The invention aims to improve the fluorescence of a copper nanocluster and realize the rapid and sensitive detection of hydrogen peroxide and glucose, provides a preparation method of a high-fluorescence copper nanocluster-cerium (III) fluorescent probe, and is used for constructing a hydrogen peroxide and glucose fluorescent sensor.
In order to achieve the above object, a first aspect of the present invention provides a high fluorescence copper nanocluster-cerium (III) fluorescent probe, which is a preparation method of the high fluorescence copper nanocluster-cerium (III) fluorescent probe, the method comprises adding a copper nanocluster into an acetic acid-sodium acetate buffer solution, mixing the mixture by vortexing, adding trivalent cerium ions into the mixture, and reacting to obtain the high fluorescence copper nanocluster-cerium (III) fluorescent probe.
The method is further provided that the copper nanocluster is prepared by the following method: 10mL of glutathione with a concentration of 50mg/mL is added dropwise to a solution containing 10mL of CuSO with a molar concentration of 10mM4The reaction is carried out for 1 hour at 37 ℃ and 650rpm, then the pH of the reaction solution is adjusted by 1M NaOH solution, the pH is adjusted to 5 by contrast with pH test paper, and the reaction is continued for 2 hours to prepare the copper nanocluster.
Further setting the pH value of the acetic acid-sodium acetate buffer solution to be 3.6-5.6.
The concentration of the trivalent cerium ions is further set to be 1-70 mM.
The second purpose of the invention is to provide a high-fluorescence copper nanocluster-cerium (III) fluorescent probe prepared by the preparation method.
The third purpose of the invention is to provide an application method of the high-fluorescence copper nanocluster-cerium (III) fluorescent probe for hydrogen peroxide detection, wherein hydrogen peroxide to be detected is added into the fluorescent probe, and hydrogen peroxide detection data are obtained by testing the fluorescence intensity of a system.
The fourth object of the invention is to provide an application method of the high-fluorescence copper nanocluster-cerium (III) fluorescent probe for glucose detection, glucose to be detected is reacted with glucose oxidase, then reaction liquid is added to the fluorescent probe, and the detection data of the glucose is obtained by testing the fluorescence intensity of a system.
The fourth purpose of the invention is to provide a preparation method of the high fluorescence copper nanocluster-cerium (III) fluorescence test paper, which comprises the steps of adding the copper nanoclusters into an acetic acid-sodium acetate buffer solution, carrying out vortex mixing, adding trivalent cerium ions into a mixed solution, carrying out vortex mixing, carrying out vacuum filtration on the mixed solution by taking a 0.22 mu m filter membrane as a carrier after reaction, airing the filtered filter membrane at room temperature, and punching the filter membrane into a round test paper by using a puncher.
In the application of the invention to detecting hydrogen peroxide, the concentration of the hydrogen peroxide is 0.05, 0.1, 0.2, 0.5, 1, 2, 5, 10 and 15mM, and the fluorescence emission peak intensity of the fluorescent probe at 650nm is measured under the condition that the excitation wavelength of the test system is 350 nm.
In the application of the glucose detection, the volume ratio of the glucose to the glucose oxidase (1.25mg/mL) is 1: 4-4: 1, the total volume is 200 muL, and the concentration of the glucose is 0.1, 0.2, 0.5, 1, 2, 5, 10 and 20 mM.
The detection range of the hydrogen peroxide related by the invention is 10-3000 mu M, the lowest detection limit is 3 mu M, the detection range of the glucose is 16-3200 mu M, and the lowest detection limit is 4.8 mu M.
In addition, the fluorescence detection method based on the high-fluorescence copper nanocluster-cerium (III) has high selectivity, and can effectively avoid coexistence of small biological molecules (D-fructose, α -lactose, D- (+) -maltose, ascorbic acid, sucrose, uric acid, lysine and tryptophan) and ions (Ca)2+、Mg2+、Zn2+) The interference of (2).
The cerium (III) is used as an inducer to promote the aggregation induction effect of the copper nanocluster to prepare the high-fluorescence copper nanocluster-cerium (III) probe, and the hydrogen peroxide can quench the fluorescence of the copper nanocluster to realize the quantitative detection of the hydrogen peroxide and the glucose. The method is simple to operate, good in selectivity and wide in linear range, provides a new method for sensitive detection of hydrogen peroxide and glucose, and can realize visual detection of hydrogen peroxide and glucose.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is within the scope of the present invention for those skilled in the art to obtain other drawings based on the drawings without inventive exercise.
FIG. 1 is a high transmission electron microscopy image of a copper nanocluster;
FIG. 2 is a graph of fluorescence contrast for copper nanoclusters, copper nanoclusters-cerium (III);
FIG. 3 is a graph of the fluorescence response of copper nanoclusters-cerium (III) at different concentrations of hydrogen peroxide;
FIG. 4 is a line graph of hydrogen peroxide detection;
FIG. 5 is a graph of the fluorescence response of copper nanoclusters-cerium (III) at different concentrations of glucose;
FIG. 6 is a line graph of glucose detection;
FIG. 7 is a visual detection of glucose based on a highly fluorescent copper nanocluster-cerium (III) fluorescent test paper.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention will be described in further detail with reference to the accompanying drawings.
Example 1
Preparing a copper nanocluster: 10mL glutathione (50mg/mL) was added dropwise to a solution containing 10mL CuSO4(10mM) in a round-bottomed flask (50mL) at 37 ℃ and 650rpmThe reaction was continued for 1 hour to give a milky white suspension. Then, the pH of the reaction solution is adjusted by NaOH (1M) solution, the pH is adjusted to 5 by contrast pH test paper, the solution color becomes light yellow, the reaction is continued for 2 hours, and the prepared copper nanocluster is stored in a refrigerator at 4 ℃. As can be seen from FIG. 1, the particle size of the prepared copper nanoclusters was about 2nm and was uniformly dispersed.
Preparing a copper nanocluster-cerium (III) fluorescent probe: to a 1mL centrifuge tube was added 600. mu.L of acetic acid-sodium acetate buffer solution (pH 5.6), 100. mu.L of the copper nanoclusters prepared in example 1, vortexed and mixed, and 100. mu.L of cerium (III) ions (20mM) was added to the mixture and reacted for 10 min.
Example 2
The copper nanocluster prepared by the method for preparing the copper nanocluster of example 1 is used.
Preparing a copper nanocluster-cerium (III) fluorescent probe: to a 1mL centrifuge tube was added 600. mu.L of acetic acid-sodium acetate buffer solution (pH 3.6), 100. mu.L of the copper nanoclusters prepared in example 1, vortexed and mixed, and 100. mu.L of cerium (III) ions (10mM) was added to the mixture and reacted for 10 min.
Example 3
The copper nanocluster prepared by the method for preparing the copper nanocluster of example 1 is used.
Preparing a copper nanocluster-cerium (III) fluorescent probe: to a 1mL centrifuge tube was added 600. mu.L of acetic acid-sodium acetate buffer solution (pH 3.6), 100. mu.L of the copper nanoclusters prepared in example 1, vortexed and mixed, and 100. mu.L of cerium (III) ions (20mM) was added to the mixture and reacted for 10 min. As shown in fig. 2, when 20mM cerium (III) was added to the copper nanocluster solution to form a copper nanocluster-cerium (III) whose fluorescence was enhanced by 40 times as compared to the copper nanoclusters.
Application example 1
Detection of hydrogen peroxide: to the mixed solution of the copper nanocluster-cerium (III) fluorescent probe prepared in example 3, 200 μ L of hydrogen peroxide (0.05, 0.1, 0.2, 0.5, 1, 2, 5, 10, 15mM) at various concentrations was added and reacted for 10min, and the intensity of the fluorescence emission peak of the fluorescent probe at 650nm was measured at an excitation wavelength of 350 nm. As shown in fig. 3, the fluorescence of the copper nanocluster-cerium (III) probe gradually decreased as the hydrogen peroxide concentration increased. The fluorescence intensity was linear with the logarithm of the hydrogen peroxide concentration, ranging from 10. mu.M to 3mM, with a detection limit of 3. mu.M (FIG. 4).
Application example 2
And (3) detection of glucose: mu.L of glucose (0.1, 0.2, 0.5, 1, 2, 5, 10, 20mM) at various concentrations was reacted with 160. mu.L of glucose oxidase (1.25mg/mL) for 30 minutes at 37 ℃. And then adding the obtained reaction solution of glucose and glucose oxidase into the mixed solution of the copper nanocluster-cerium (III) fluorescent probe prepared in example 3, and reacting for 10min, wherein the intensity of a fluorescence emission peak of the fluorescent probe at 650nm is tested under the excitation wavelength of 350 nm.
Application example 3
And (3) detection of glucose: mu.L of glucose (0.1, 0.2, 0.5, 1, 2, 5, 10, 20mM) at various concentrations was reacted with 40. mu.L of glucose oxidase (1.25mg/mL) for 30 minutes at 37 ℃. And then adding the obtained reaction solution of glucose and glucose oxidase into the mixed solution of the copper nanocluster-cerium (III) fluorescent probe prepared in example 3, and reacting for 10min, wherein the intensity of a fluorescence emission peak of the fluorescent probe at 650nm is tested under the excitation wavelength of 350 nm. As shown in FIG. 5, the fluorescence of the copper nanocluster-cerium (III) complex gradually decreased as the concentration of glucose was gradually increased, and the fluorescence intensity thereof had a good linear relationship with the logarithm of the glucose concentration in the range of 16. mu.M to 3.2mM (FIG. 6), with a detection limit of 4.8. mu.M.
Application example 4
Detecting glucose by using high-fluorescence copper nanocluster-cerium (III) fluorescence test paper: (1) adding 60mL of acetic acid-sodium acetate buffer solution (pH 3.6) and 10mL of the copper nanocluster prepared in example 1 into a 100mL centrifuge tube, uniformly mixing by vortex, adding 10mL of 20mM cerium (III) ions into the mixed solution, uniformly mixing by vortex, reacting for 10min, vacuum-filtering the mixed solution by taking a 0.22-micrometer filter membrane as a carrier, drying the filtered filter membrane at room temperature, and punching the filter membrane into a circular test paper with the diameter of 3mM by using a puncher. (2) mu.L of glucose (0.1, 0.2, 0.5, 1, 2, 5, 10, 20mM) at various concentrations was reacted with 40. mu.L of glucose oxidase (1.25mg/mL) for 30 minutes at 37 ℃. And then dropwise adding the obtained reaction liquid of glucose and glucose oxidase onto the prepared round copper nanocluster-cerium (III) fluorescent test paper, and reacting for 10 min. Then, the photo was taken under 365nm UV light. As shown in FIG. 7, when the concentration of glucose gradually increased, the fluorescent bright spot gradually became dark, indicating that the fluorescent probe based on the copper nanocluster-cerium (III) can realize the visual detection of glucose.
The above disclosure is only for the purpose of illustrating the preferred embodiments of the present invention, and it is therefore to be understood that the invention is not limited by the scope of the appended claims.
Claims (8)
1. A preparation method of a high-fluorescence copper nanocluster-cerium (III) fluorescent probe is characterized by comprising the following steps: adding the copper nanocluster into an acetic acid-sodium acetate buffer solution, uniformly mixing by vortex, adding trivalent cerium ions (cerium (III)) into the mixed solution, and reacting to obtain the high-fluorescence copper nanocluster-cerium (III) fluorescent probe.
2. The method of claim 1, wherein: the copper nanocluster is prepared by the following method: 10mL of glutathione with a concentration of 50mg/mL is added dropwise to a solution containing 10mL of CuSO with a molar concentration of 10mM4The reaction is carried out for 1 hour at 37 ℃ and 650rpm, then the pH of the reaction solution is adjusted by 1M NaOH solution, the pH is adjusted to 5 by contrast with pH test paper, and the reaction is continued for 2 hours to prepare the copper nanocluster.
3. The method of claim 1, wherein: the pH value of the acetic acid-sodium acetate buffer solution is 3.6-5.6.
4. The method of claim 1, wherein: the concentration of the trivalent cerium ions is 1-70 mM.
5. A high fluorescence copper nanocluster-cerium (III) fluorescent probe prepared by the preparation method of any one of claims 1 to 4.
6. The application method of the high fluorescence copper nanocluster-cerium (III) fluorescent probe as claimed in claim 5 in hydrogen peroxide detection, which is characterized in that: and adding hydrogen peroxide to be detected into the fluorescent probe, and obtaining hydrogen peroxide detection data through the fluorescence intensity of a test system.
7. The method for applying the high-fluorescence copper nanocluster-cerium (III) fluorescent probe as claimed in claim 5 to glucose detection, wherein the method comprises the following steps: and reacting glucose to be detected with glucose oxidase, adding the reaction solution into the fluorescent probe, and obtaining detection data of the glucose through the fluorescence intensity of a test system.
8. A preparation method of high-fluorescence copper nanocluster-cerium (III) fluorescence test paper is characterized by comprising the following steps: adding copper nanoclusters into an acetic acid-sodium acetate buffer solution, uniformly mixing in a vortex mode, adding trivalent cerium ions into a mixed solution, uniformly mixing in a vortex mode, after reaction, taking a filter membrane of 0.22 mu m as a carrier, carrying out vacuum filtration on the mixed solution, airing the filtered filter membrane at room temperature, and punching the filter membrane into round test paper by using a puncher.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN115322771A (en) * | 2022-07-11 | 2022-11-11 | 安徽师范大学 | Cerium-doped copper nano material and preparation method thereof |
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| CN109724957A (en) * | 2018-12-28 | 2019-05-07 | 南京工业大学 | One kind is based on the aluminium ion induction phosphorescence copper nano-cluster aggregation enhancing aluminum ions method and its application of fluorescence detection |
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| US7504356B1 (en) * | 2006-02-17 | 2009-03-17 | University Of Central Florida Research Foundation, Inc. | Nanoparticles of cerium oxide having superoxide dismutase activity |
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| CN109724957A (en) * | 2018-12-28 | 2019-05-07 | 南京工业大学 | One kind is based on the aluminium ion induction phosphorescence copper nano-cluster aggregation enhancing aluminum ions method and its application of fluorescence detection |
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| CN115322771A (en) * | 2022-07-11 | 2022-11-11 | 安徽师范大学 | Cerium-doped copper nano material and preparation method thereof |
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