CN107345910B - Fluorescent wide-chromaticity test paper for visually detecting copper ions and preparation method and application thereof - Google Patents
Fluorescent wide-chromaticity test paper for visually detecting copper ions and preparation method and application thereof Download PDFInfo
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Abstract
A fluorescence wide-chromaticity test paper for visually detecting copper ions and a preparation method and application thereof are disclosed, wherein a three-color ratio fluorescence probe is used as ink, and filter paper is used as a solid phase carrier through an ink-jet printer to be uniformly printed to obtain the test paper for visually detecting the copper ions; the three-color ratio fluorescent probe is a blue carbon dot, a green quantum dot and a red quantum dot, and the quantum dots are modified by 3-mercaptopropionic acid. The invention firstly utilizes the principle of double quenching to construct three-color ratio fluorescent test paper and is used for visually detecting copper ions, the prepared test paper has the advantage of wider color change range than the existing double-ratio fluorescent probe test paper, and the test paper realizes that the color of the test paper can be obviously identified by naked eyes from light red color, light flesh color, dark orange color, grass green color, dark olive green color, slate blue color, palygreen color and fine blue color (figure 4) along with the addition of a detected substance.
Description
Technical Field
The invention belongs to the technical field of copper ion detection, and particularly relates to a fluorescent wide-chromaticity test paper for visually detecting copper ions, and a preparation method and application thereof.
Background
Copper is an important element of human health, and enzymes use the redox activity of this metal to perform energy production, neurotransmitter and pigment synthesis, and epigenetic modification functions. On the one hand, copper deficiency can lead to a number of diseases, such as anemia, pancytopenia and bone marrow abnormalities. On the other hand, abnormal copper metabolism is associated with many diseases including cancer, neurodegenerative alzheimer's disease, parkinson's and huntington's disease, and hereditary diseases such as curling syndrome and wilson's disease. As a typical copper-mediated disease, wilson's disease is an autosomal recessive genetic disease with copper accumulation in tissues. Patients with wilson's disease often show high urinary copper levels (>100 μ g/24 h, compared to normal). Normal human reference values for urine copper content at 24 hours have been reported differently from laboratory to laboratory, most with 40. mu.g/24 hours (0.6. mu. mol/24 hours) as the upper limit of normal. Therefore, urine copper can be detected as a clinical diagnosis index.
Conventional methods for detecting copper ions include inductively coupled plasma mass spectrometry, atomic absorption spectrometry, atomic emission spectrometry, and mass spectrometry. However, the conventional large-scale laboratory instrument has the defects of high price, huge and heavy volume, complex sample pretreatment before detection, operation by specially trained professionals and the like, and in addition, the sample is separated from the detection field and sent to an analysis laboratory, so that the real-time field detection cannot be realized. Therefore, finding a simple, fast, low cost, and portable method for field testing is a significant challenge for analysts.
In recent years, semiconductor quantum dots and carbon dots have a significant advantage over conventional organic dyes, and thus become optical labels with great potential for use in the chemical/biological sensing field. The advantages of quantum dots attract the attention of researchers, such as good optical performance, good photochemical stability, long fluorescence lifetime, good water solubility and the like. In addition, the fluorescence sensor has another incomparable advantage that only a portable ultraviolet lamp is needed to realize the visual detection of the analyte by direct observation with naked eyes. Based on the wide application of the classical pH test paper, the fluorescence detection test paper often fixes fluorescent probe molecules on a substrate in an assembly or printing manner, and the detection test paper prepared by the method has the advantages of low cost, convenience in operation, easiness in carrying and the like.
However, such a fluorescence test strip similar to a pH test strip still presents a difficult problem. At present, the establishment of fluorescent test strips is mostly realized by using fluorescent probes with single color, but the fluorescent probes with single color can only detect analytes through the change of fluorescence intensity generated by fluorescence turn-on or quenching, and the method greatly limits the quantification capability of the test strips. Later, dual emission rate probes were developed to achieve more accurate visual detection of targets using color changes instead of single color intensities. Unfortunately, dual emission ratiometric probes remain unsuccessful because when two fluorescent probes of different colors are mixed, the resulting color of the ratiometric probe must be an intermediate composite color of the two, thereby narrowing the range of color variations. When a fluorescent probe is used on the test strip, the fluorescent test strip exhibits only a two-part color change in nature, which is not significant. Generally, the existing fluorescence test paper generally has the problems of low sensitivity to the chromaticity change of the target dose, narrow spectrum evolution range, incapability of performing visual quantitative detection on the target and the like, and the fluorescence test paper capable of performing visual quantitative detection on the target is rare.
Disclosure of Invention
The technical problem to be solved is as follows: the invention aims to provide a three-color ratio fluorescent test paper and a method for visually detecting copper ions, and the method has the characteristics of high selectivity, high sensitivity, quantitative detection, visual detection result and obvious color change.
The technical scheme is as follows: a preparation method of fluorescent wide-chromaticity test paper for visually detecting copper ions comprises the steps of taking a three-color ratio fluorescent probe as ink, taking filter paper as a solid-phase carrier through an ink-jet printer, and uniformly printing to obtain the test paper for visually detecting the copper ions; the three-color ratio fluorescent probe is a blue carbon dot, a green quantum dot and a red quantum dot, and the quantum dots are modified by 3-mercaptopropionic acid.
The preparation of the three-color ratiometric fluorescent probe comprises the following steps: (1) preparing cadmium telluride quantum dots: in proportion, 0.2284g of cadmium chloride (CdCl)2·2.5H2O) was added to 100mL of deoxygenated ultrapure water, followed by addition of 0.21mL of 3-mercaptopropionic acid (MPA), and the pH thereof was adjusted to 10 with 1M sodium hydroxide solution to give a mixed solution; 0.0319g of tellurium powder and 0.05g of sodium borohydride are taken and added into 2mL of ultrapure water, ice bath is carried out for 8 hours under the protection of nitrogen, and sodium hydrogen telluride solution is generated through reaction; 10mL of a 0.5M sulfuric acid solution was injected into the produced sodium hydrogen telluride solution, and the produced H was removed by flowing nitrogen gas2Completely introducing Te into the mixed solution, stirring for 20 minutes at room temperature, changing the color of the mixed solution from colorless to orange yellow, and continuously heating and refluxing for 50 minutes at 100 ℃ to obtain mercaptopropionic acid modified green quantum dots; or heating and refluxing for 24-48 hours to obtain red quantum dots; (2) preparation of blue carbon dots: dissolving 1-2g of citric acid and 100-500 mu L of ethylenediamine in 20mL of ultrapure water according to the proportion, then transferring the solution into a 30mL polytetrafluoroethylene reaction kettle, reacting for 4-8 hours at 200 ℃, and dialyzing the obtained carbon spots for 24-48 hours by using a dialysis bag with the molecular weight cut-off of 500 for later use; (3) trichromatic ratio fluorescencePreparation of an optical probe: and proportionally adding 90 mu L of green quantum dot solution, 40 mu L of red quantum dot solution and 30 mu L of blue carbon dot solution into a beaker, adding 10 mMN-2-hydroxyethyl piperazine-N' -2 ethanesulfonic acid buffer solution with the pH value of 7.420 mL, and uniformly mixing to obtain the three-color mixed ratio fluorescent probe solution.
Preferably, the volume ratio of the blue carbon dot solution, the green quantum dot solution and the red quantum dot solution is 3:9: 4.
Preferably, the quantum dot stock solution obtained in step (1) is irradiated under an ultraviolet lamp of 15W for 1 hour to improve the quantum yield.
Preferably, the quantum dots obtained in step (1) are washed three times with acetone, and the precipitate is dispersed in ultrapure water for later use.
The preparation method of the test paper comprises the following steps: cleaning purchased commercial printer cartridge with deionized water, oven drying to obtain blank cartridge, injecting three-color ratio probe into the blank cartridge with syringe, and printing a 7 × 3cm print on common filter paper2Repeatedly printing for about 30 times, naturally standing for 5 min, and cutting into 3 × 1cm2The strip-shaped test paper.
Preferably, the thickness of the fluorescent probe layer is 0.05 to 0.1. mu.m.
The fluorescence wide-chromaticity test paper for visually detecting copper ions is prepared by the preparation method.
According to the fluorescence wide-chromaticity test paper for visually detecting the copper ions, the three-color ratio fluorescence probe can respectively emit blue, green and red fluorescence under the excitation of the single-wavelength light source; wherein the wavelength excited by the single-wavelength light source is 360 nm; the blue fluorescence emission wavelength is 440 nm; the green fluorescence emission wavelength is 510 nm; the red fluorescence emission wavelength was 600 nm.
The fluorescent wide-chromaticity test paper is applied to visual detection of copper ions.
The technical scheme of the invention comprises that the preparation method comprises the steps of mixing blue carbon dots, green quantum dots and red quantum dots to obtain a fluorescent probe, and printing the fluorescent probe on test paper. The effectiveness of the in-situ visual detection of copper ions is proved by a double quenching method. The blue carbon dots are fluorescence stable to copper ions, whereas the fluorescence of green and red quantum dots can be rapidly quenched by copper ions. Compared with the traditional visual test paper, when the concentration of copper ions is relatively low, the test paper prepared based on the three-color probe has the characteristic of high macroscopic resolution. The fluorescent test paper can visually detect the content of copper ions in a water sample and a urine sample.
Has the advantages that: the invention firstly utilizes the principle of double quenching to construct three-color ratio fluorescent test paper and is used for visually detecting copper ions. The test paper prepared by the invention has the advantage of wider color change range than the existing double-ratio fluorescent probe test paper, realizes that the test paper has the color from light red color to light flesh color to dark orange color to grass green color to dark olive green color to slate blue color to paleness blue color (figure 4) along with the addition of a detected substance, and can be obviously identified by naked eyes. The invention widens the application range of the test paper detection technology and provides a diagnosis index for clinically diagnosing diseases caused by copper ion deficiency or copper ion metabolic disorder. The preparation method is simple, the raw materials are simple and easy to obtain, special technical personnel are not needed, large-scale instruments can be avoided to a certain extent, visual detection can be carried out only by one handheld ultraviolet lamp, and the method is simple to operate, rapid and convenient, high in sensitivity and remarkable in effect. The method can effectively avoid the interference of other impurities in the sample and has good selectivity. The prepared test paper can visually detect copper ions on site in real time on line.
Drawings
Fig. 1 is a fluorescence spectrum of blue carbon dots (a), green quantum dots (b), red quantum dots (c), and a quantum dot/carbon dot mixed system (d).
FIG. 2 is a graph showing the fluorescence spectrum and color change of a mixed system of copper ions with different concentrations and quantum dots/carbon dots. The color of the solution gradually changes with the increase of the concentration of copper ions (0, 3,6,10,20,40,60,80,110,140,180,230,280,330,430nM from left to right).
FIG. 3 shows the selectivity of the test strip for various other ions.
Fig. 4 is a visual photograph of a test paper for detecting copper ions in an aqueous solution, wherein the first action is the color change of the test paper under an ultraviolet lamp, and the second action is the color change of the test paper under sunlight. The concentration of copper ions is 0,6,20,60,110,180,280 and 430nM from left to right.
Fig. 5 is a visual photograph of the test paper for detecting copper ions in urine, wherein the first action is the color change of the test paper under sunlight, and the second action is the color change of the test paper under an ultraviolet lamp. The copper ion concentration was 58.7,26.3,178nM from left to right.
Detailed Description
The following examples further illustrate the present invention but are not to be construed as limiting the invention. Modifications and substitutions to methods, procedures, or conditions of the invention may be made without departing from the spirit and substance of the invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art.
Example 1
1. Preparation of quantum dots
0.2284g of cadmium chloride (CdCl)2·2.5H2O) was added to 100mL of deoxygenated ultrapure water, followed by addition of 0.21mL of 3-mercaptopropionic acid (MPA), and then the pH thereof was adjusted to 10 with 1M sodium hydroxide solution to give a mixed solution; 0.0319g of tellurium powder and 0.05g of sodium borohydride are taken and added into 2mL of ultrapure water, ice bath is carried out for 8 hours under the protection of nitrogen, and sodium hydrogen telluride is generated through reaction; injecting 10mL0.5M sulfuric acid solution into the generated sodium hydrogen telluride solution, and generating H by flowing nitrogen2And (3) completely introducing Te into the mixed solution, stirring for 20 minutes at room temperature, changing the color of the mixed solution from colorless to orange yellow, continuously heating and refluxing for 50 minutes at 100 ℃ to obtain green cadmium telluride quantum dots modified by mercaptopropionic acid, heating and refluxing for 24-48 hours if red quantum dots are obtained, irradiating the obtained quantum dot stock solution for 1 hour under a 15W ultraviolet lamp, washing with acetone for three times, and dispersing the precipitate into ultrapure water for later use.
2. Preparation of carbon dots
1-2g of citric acid and 500 mu L of ethylenediamine are dissolved in 20mL of ultrapure water, then the solution is transferred to a 30mL polytetrafluoroethylene reaction kettle and reacted at 200 ℃ for 4-8 hours, and the obtained carbon spots are dialyzed for 24-48 hours by a dialysis bag with the molecular weight cut-off of 500 for later use.
3. Preparation of trichromatic ratiometric fluorescent probes
Adding 90 mu L of green quantum dots, 40 mu L of red quantum dots and 30 mu L of blue carbon dots into a beaker, adding 20mL of 10mM N-2-hydroxyethylpiperazine-N' -2 ethanesulfonic acid buffer solution (pH 7.4), and uniformly mixing to obtain the three-color ratiometric fluorescent probe solution. The fluorescence spectrum is shown in FIG. 1.
4. Visual detection of copper ions by three-color ratio fluorescent probe mixed system
And adding the copper ion solution to be detected into a three-color ratio fluorescent probe mixed system for fluorescence visual detection. With the increasing concentration of copper ions, the fluorescence color is from light red color to light flesh color to dark orange color to grass green color to dark olive green color to slate blue color to gem blue color to sunny blue color. At this moment, under the ultraviolet lamp, the step change of the color can be seen, and the visual detection is realized. The visualization is shown in fig. 2.
Example 2
1. Preparation of quantum dots
The preparation method of the quantum dots in this example is the same as example 1.
2. Preparation of carbon dots
The method for preparing the carbon dots in this example is the same as in example 1.
3. Preparation of trichromatic ratiometric fluorescent probes
The three-color ratiometric fluorescent probes in this example were prepared as in example 1.
4. Standard stock solutions of 0.3mM copper ions and 3mM other ions were prepared, respectively.
5. Adding a certain volume of the different ion standard stock solution in the step 4 into the solution of the three-color ratiometric fluorescent probe.
6. The selectivity of the test strip for various other ions is shown in FIG. 3.
Example 3
1. Preparation of quantum dots
The preparation method of the quantum dots in this example is the same as example 1.
2. Preparation of carbon dots
The method for preparing the carbon dots in this example is the same as in example 1.
3. Preparation of trichromatic ratiometric fluorescent probes
The three-color ratiometric fluorescent probes in this example were prepared as in example 1.
4. Preparation of test paper
And cleaning the purchased commercial printer ink box with deionized water, and drying to obtain a blank ink box. The three color ratio probe was injected into a blank cartridge using a syringe and printed on plain filter paper in a 7X 3cm format2Repeatedly printing for about 30 times, naturally standing for 5 min, and cutting into 3 × 1cm2The test paper which responds to the copper ions of the target object can be obtained, and the thickness of the fluorescent probe layer is 0.05-0.1 mu m.
5. Visual detection of copper ions in water
After the test paper is dried, copper ions with different concentrations are slowly dripped on the prepared test paper, and after the test paper is dried for 5 minutes at room temperature, the test paper is irradiated by ultraviolet light excited by 360nm and the change of the fluorescence color of the test paper is observed. The visual picture is shown in fig. 4.
Example 4
1. Preparation of quantum dots
The preparation method of the quantum dots in this example is the same as example 1.
2. Preparation of carbon dots
The method for preparing the carbon dots in this example is the same as in example 1.
3. Preparation of trichromatic ratiometric fluorescent probes
The three-color ratiometric fluorescent probes in this example were prepared as in example 1.
4. Preparation of test paper
And cleaning the purchased commercial printer ink box with deionized water, and drying to obtain a blank ink box. The three color ratio probe was injected into a blank cartridge using a syringe and printed on plain filter paper in a 7X 3cm format2Repeatedly printing for about 30 times, naturally standing for 5 min, and cutting into 3 × 1cm2To obtain the target copper ionsThe thickness of the fluorescent probe layer of the responsive test paper is 0.05-0.1 μm.
5. Visual detection of copper ions in urine
After the test paper is dried, slowly dripping the sample-added copper ion urine sample containing different concentrations on the prepared test paper, drying for 5 minutes at room temperature, irradiating by ultraviolet light excited by 360nm, and observing the change of the fluorescence color of the sample-added copper ion urine sample. The visual picture is shown in fig. 5.
Claims (4)
1. A preparation method of fluorescent wide-chromaticity test paper for visually detecting copper ions is characterized by comprising the following steps of (1) preparation of quantum dots: 0.2284g of CdCl2•2.5 H2Adding O into 100mL of deoxygenated ultrapure water, then adding 0.21mL of 3-mercaptopropionic acid, and adjusting the pH value of the mixture to 10 by using 1M sodium hydroxide solution to obtain a mixed solution; 0.0319g of tellurium powder and 0.05g of sodium borohydride are taken and added into 2mL of ultrapure water, ice bath is carried out for 8 hours under the protection of nitrogen, and sodium hydrogen telluride is generated through reaction; 10mL of a 0.5M sulfuric acid solution was injected into the produced sodium hydrogen telluride solution, and the produced H was removed by flowing nitrogen gas2Completely introducing Te into the mixed solution, stirring for 20 minutes at room temperature, changing the color of the mixed solution from colorless to orange yellow, and continuously heating and refluxing for 50 minutes at 100 ℃ to obtain green cadmium telluride quantum dots modified by mercaptopropionic acid; or heating and refluxing for 24-48 hours, irradiating the obtained quantum dot stock solution for 1 hour under a 15W ultraviolet lamp, washing with acetone for three times, and dispersing the precipitate in ultrapure water to obtain red cadmium telluride quantum dots; (2) preparation of blue carbon dots: dissolving 1-2g of citric acid and 500 mu L of 100-one ethylene diamine in 20mL of ultrapure water, then transferring the solution into a 30mL polytetrafluoroethylene reaction kettle, reacting for 4-8 hours at 200 ℃, and dialyzing the obtained carbon spots for 24-48 hours by using a dialysis bag with the molecular weight cutoff of 500 for later use; (3) preparation of trichromatic ratiometric fluorescent probes: adding 90 mu L of green quantum dots, 40 mu L of red quantum dots and 30 mu L of blue carbon dots into a beaker, adding 20mL of 10mM buffer solution with pH = 7.4N-2-hydroxyethylpiperazine-N' -2 ethanesulfonic acid, and uniformly mixing to obtain a three-color ratio fluorescent probe solution; (4) preparation of detection test paper: will purchase backWashing commercial printer cartridge with deionized water, oven drying to obtain blank cartridge, injecting three-color ratio probe solution into the blank cartridge with syringe, and printing on common filter paper to obtain 7 × 3cm print2Repeatedly printing 30 times, naturally standing for 5 min, and cutting into 3 × 1cm2The thickness of the fluorescent probe layer of the strip-shaped test paper is 0.05-0.1 μm.
2. The wide-chromaticity fluorescence test paper for visually detecting copper ions, which is prepared according to the preparation method of claim 1.
3. The wide-chromaticity fluorescence test paper for visually detecting copper ions according to claim 2, wherein the three-color ratio fluorescence probe can emit blue, green and red fluorescence respectively under the excitation of a single-wavelength light source; wherein the wavelength excited by the single-wavelength light source is 360 nm; the blue fluorescence emission wavelength is 440 nm; the green fluorescence emission wavelength is 510 nm; the red fluorescence emission wavelength was 600 nm.
4. The use of the wide-colorimetric fluorescent strip of claim 2 for the visual detection of copper ions.
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| CN109900693A (en) * | 2019-04-12 | 2019-06-18 | 浙江科技学院 | A kind of copper ion test paper and its detection method |
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| CN111040757A (en) * | 2019-11-01 | 2020-04-21 | 浙江工业大学 | Preparation method and application of ratiometric fluorescent probe for detecting copper ions |
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