Preparation method and application of fluorescent sensor array for detecting heavy metal ions

Abstract

The present invention provides a method for preparing a fluorescent sensor array for detecting heavy metal ions, the method comprising: mixing a cetylpyridinium chloride solution with a sodium hydroxide solution, then performing water bath ultrasound, purifying with dichloromethane, and then dialyzing with a dialysis bag to obtain a CPC-CDs solution; adding heavy metal ion solutions of different concentrations to CPC-CDs / XO sensing solutions under two groups of pH environments to obtain a fluorescent sensor array. Also provided is an application for detecting heavy metal ions Cr 6+ , Fe 2+ , Fe 3+ , and Hg 2+ The present invention simplifies the array construction steps by adjusting the pH environment of the system, and effectively improves the lack of affinity between carbon dots and heavy metal ions by adding xylenol, and can simultaneously detect multiple metal ions (Cr 6+ , Fe 2+ , Fe 3+ , and Hg 2+ ), with high sensitivity and good selectivity, is used for the detection of heavy metals in the environment.

Description

Technical Field

[0001] The invention belongs to the technical field of heavy metal detection, and in particular relates to a preparation method and application of a fluorescent sensor array for detecting heavy metal ions. Background Art

[0002] Heavy metal pollution, as a key and difficult point in the current environmental governance research, has attracted the extensive attention of many scholars. In the social context of the rapid development of modern industry, a large number of carcinogenic, teratogenic and mutagenic heavy metal ions are released into the environment, causing a significant decline in environmental quality or endangering human health. Choosing a fast and accurate detection method is a necessary condition for the prevention and control of heavy metal pollutants. Traditional detection methods such as enzyme analysis, chemical colorimetry and biochemical methods generally have the disadvantages of relatively high detection limits, expensive instruments, complex operations and low precision. As a method in the optical sensor array, the fluorescent sensor array has the advantages of high sensitivity, no need for a reference system, rich output signals and the ability to image. However, so far, there is a relative lack of research reports on the detection of heavy metals based on carbon dot arrays. Therefore, in existing research, it is very necessary to develop a detection method with strong specificity based on the advantages of fluorescent sensor arrays.

[0003] Carbon quantum dots (CDs) are a type of quasi-spherical nanomaterial with a size of about 10 nm and fluorescence. They have attracted extensive attention in the research field because of their wide source of raw materials and simple preparation methods. Carbon quantum dots are used as sensing units to construct fluorescent sensor arrays, which transmit information in the form of fluorescent signals through the molecular recognition process using the interaction between molecules and ions. The array sensing platform has the advantages of accuracy, diversity, simplicity, time saving and high selectivity. At present, the research on the specific detection of heavy metal pollutants using carbon quantum dots has been relatively extensive, while there are fewer studies on the construction of fluorescent sensor arrays for detection, and the research on array-based detection of heavy metal pollutants is relatively blank. At present, the research on fluorescent sensor arrays based on carbon dots mainly focuses on the enhancement or reduction of the fluorescence intensity of a single fluorophore as the response signal, and on this basis, the functional groups on the surface of carbon dots are functionally modified, or fluorescent dyes are used as raw materials to artificially synthesize fluorescent probes. Summary of the invention

[0004] The technical problem to be solved by the present invention is to provide a preparation method and application of a fluorescent sensor array for detecting heavy metal ions in view of the above-mentioned deficiencies in the prior art. The method constructs a fluorescent sensor array by adjusting the pH environment of the system, and adds xylenol orange as a receptor to bind carbon quantum dots to heavy metal ions (Cr 6+ , Fe 2+ , Fe 3+ and Hg 2+) has high detection sensitivity, high accuracy and good stability, and can be used for the detection of heavy metals in the environment.

[0005] In order to solve the above technical problems, the technical solution adopted by the present invention is: a method for preparing a fluorescent sensor array for detecting heavy metal ions, the method comprising:

[0006] S1. Preparation of CPC-CDs solution: at room temperature, transfer the sodium hydroxide solution to the cetylpyridinium chloride solution, and perform water bath sonication to obtain a reaction solution; add dichloromethane to the reaction solution, extract and purify, extract 3 times in total, let stand until the solution is colorless and transparent, dialyze in deionized water with a dialysis bag to obtain CPC-CDs solution (cetylpyridinium carbon quantum dots), and store at 4°C in the dark;

[0007] S2. Establishment of pH-regulated carbon quantum dot fluorescence sensor array: Two groups of pH detection environments were set up. The first group of pH detection environments was Tris-HCl buffer solution A with a pH of 7.4 and a concentration of 10 mM; the second group of pH detection environments was Tris-HCl buffer solution B with a pH of 8.8 and a concentration of 10 mM.

[0008] 950 μL of Tris-HCl buffer solution, 20 μL of CPC-CDs solution obtained in S1 and 2 μL of 100 μM xylenol orange solution were mixed evenly and allowed to stand for 7 minutes to obtain CPC-CDs / XO sensing solutions under two pH environments; the Tris-HCl buffer solutions were Tris-HCl buffer solution A and Tris-HCl buffer solution B;

[0009] Metal ions of different concentrations were added to the CPC-CDs / XO sensing solutions under the two groups of pH environments, mixed evenly and allowed to stand for 10 minutes to obtain a CPC-CDs / XO sensing system. The fluorescence emission spectrum of CPC-CDs was recorded at an excitation wavelength of 370 nm to obtain a fluorescence sensor array.

[0010] Preferably, the volume ratio of the cetylpyridinium chloride solution and the sodium hydroxide solution in S1 is 2:1, the concentration of the cetylpyridinium chloride solution is 50 mmol / L; the concentration of the sodium hydroxide solution is 330 mmol / L; the dosage ratio of the dichloromethane and CPC-CDs solution is (1-3):1.

[0011] Preferably, the temperature of the water bath ultrasound in S1 is 20° C. to 40° C., and the time is 6 h to 9 h.

[0012] Preferably, the dialysis bag in S1 has a MWCO membrane with a molecular weight cutoff of 3000Da, and the dialysis time is 24h to 72h.

[0013] Preferably, the final concentration of the xylenol orange solution in the fluorescent sensor array in S2 is 0.2 μmol / L.

[0014] The present invention also provides the use of the fluorescent sensor array prepared above for detecting heavy metal ions, wherein the fluorescent sensor array is used for detecting heavy metal ions; the heavy metal ions are Cr 6+ , Fe 2+ , Fe 3+ and Hg 2+ .

[0015] Preferably, the final concentration of each heavy metal ion in the fluorescent sensor array is in the range of 0 μmol / L to 50 μmol / L.

[0016] Compared with the prior art, the present invention has the following advantages:

[0017] 1. The present invention solves the problem of poor affinity between fluorescent probe and analyte by adding chelating agent. CPC-CDs solution is used as fluorescent probe and metal chelating agent is used as receptor. Under the action of connecting arm, they are combined to detect heavy metal ions, forming a standard indicator-connecting arm-receptor mode (ISR). Due to the different chelation degrees between different heavy metal ions and chelating agents, different metal ion complexes are formed. Under the internal filtering effect, the fluorescent sensor array successfully distinguishes Cr 6+ , Fe 2 + , Fe 3+ , Hg 2+ Four heavy metals.

[0018] 2. The present invention simplifies the steps of constructing the sensor array by designing the detection environment pH to construct the fluorescence sensor array. According to the fluorescence intensity change I / I0 (I and I0 represent the fluorescence intensity in the presence and absence of heavy metals, respectively) under the two pH Tris-hydrochloric acid buffer solution systems, the fluorescence response spectra of two groups of heavy metal ions can be obtained. Therefore, the fluorescence sensor array can be constructed by adjusting the pH value of the detection environment to detect heavy metal ions.

[0019] 3. The pH-regulated fluorescence sensor array constructed by the present invention improves the defect of lack of affinity between CPC-CDs solution and heavy metal ions by adding chelating agents, thereby improving the detection efficiency and accuracy of multiple metal ions. The sensor array detects four heavy metal ions (Cr 6+ , Fe 2+ , Fe 3+ and Hg 2+) has high detection sensitivity, no response to other heavy metal ions, strong selectivity, and simple detection operation.

[0020] The present invention is further described in detail below with reference to the accompanying drawings and embodiments. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 is a microstructure diagram of CPC-CDs prepared in Example 1 of the present invention.

[0022] Figure 2 Graph showing the optical properties of CPC-CDs prepared in Example 1 of the present invention.

[0023] Figure 3 The four metal ion complexes (XO-Cr 6+ ,XO-Fe 2+ ,XO-Fe 3+ ,XO-Hg 2 + ) and the fluorescence emission spectrum of CPC-CDs (single metal ion concentration 10 μmol / L).

[0024] Figure 4 The ultraviolet absorption spectra of CPC-CDs, four metal ion complexes, CPC-CDs+metal ion complexes and CPC-CDs+metal ion complexes of Example 2 of the present invention (single metal ion concentration 10 μmol / L).

[0025] Figure 5 It is the fluorescence response degree of CPC-CDs and CPC-CDs-XO in different pH buffer solutions of Example 2 of the present invention.

[0026] Figure 6 It is the fluorescence response of the metal ions of Example 2 of the present invention and the sensor array system in different pH environments (the final concentration of all metal ions is 0-50 μmol / L).

[0027] Figure 7 This is an optimization diagram of the XO concentration in the Tris-HCl buffer solution with pH = 7.4 according to Example 2 of the present invention.

[0028] Figure 8 This is an optimization diagram of the reaction time of CPC-CDs and XO in a Tris-HCl buffer solution with a pH of 7.4 according to Example 2 of the present invention.

[0029] Fig. 9This is an optimization diagram of the incubation time of the CPC-CDs / XO sensing system at pH = 7.4 for metal ions in Example 2 of the present invention (single metal ion concentration 10 μmol / L). DETAILED DESCRIPTION

[0030] Example 1

[0031] This example is the preparation of carbon quantum dot CPC-CDs solution:

[0032] At room temperature, 10 mL of a 330 mmol / L sodium hydroxide solution was transferred to 20 mL of a 50 mmol / L cetylpyridinium chloride solution (CPC), and ultrasonicated in a water bath at 20°C for 9 h to obtain a reaction solution; dichloromethane was added to the reaction solution, extracted and purified for a total of 3 times, and allowed to stand until the solution was colorless and transparent, and dialyzed in deionized water with a dialysis bag (MWCO membrane, with a molecular weight cutoff of 3000 Da) for 72 h to obtain carbon quantum dots (CDs), referred to as CPC-CDs solution, which was stored in a dark place at 4°C; the dosage ratio of the dichloromethane to the CPC-CDs solution was 3:1.

[0033] In this embodiment, the conditions of water bath ultrasound can be 20°C to 40°C for 6h to 9h; the dialysis time can be 24h to 72h; the usage ratio of dichloromethane and CPC-CDs solution can be (1 to 3):1.

[0034] Fluorescence spectra were recorded using a fluorescence spectrophotometer (F-4500 Hitachi, Japan). UV-visible spectra were obtained using a UV-3600 UV absorption spectrometer (China General Analysis Company). Fourier transform infrared (FT-IR) spectra were recorded by a VERTEX 70 spectrometer (Bruker, Germany). X-ray photoelectron spectroscopy (XPS; ESCALAB 250Xi, USA) was processed using an advanced Avantage data system. The morphology, particle size, and lattice fringes of CPC-CDs were observed using a high-resolution transmission electron microscope (HRTEM; JEM-2100 series, 200 kV).

[0035] like Figure 1 As shown, Figure 1 (a) TEM (inset: size distribution of CPC-CDs); (b) HRTEM image of CPC-CDs; (c) infrared spectrum of CPC-CDs; (d) XPS scan of CPC-CDs. High-resolution transmission electron microscopy (HRTEM) shows that CPC-CDs have fine lattice fringes, with a typical lattice spacing of 0.20 nm ( Figure 1 a, Figure 1 b). The surface structure was studied by FT-IR and XPS spectroscopy ( Figure 1 c, Figure 1 d), the OH / NH structure was determined at 3468 eV, indicating good water solubility.

[0036] In terms of optical properties, such as Figure 2 As shown in the figure, the fluorescence excitation (a line) and emission (b line) spectra of CPC-CDs, (Inset: photos of CPC-CDs solution under natural light (right) and 365nm ultraviolet light (left)). CPC-CDs is a light yellow solution in ambient light and emits strong yellow fluorescence under 365nm ultraviolet excitation ( Figure 2 ). The maximum emission wavelength is 535nm, and the excitation wavelength is 370nm. The quantum yield measured in aqueous solution is 26.2%.

[0037] Example 2

[0038] This example is the establishment of a pH-regulated carbon quantum dot fluorescence sensor array:

[0039] Four heavy metals (Cr 6+ , Fe 2+ , Fe 3+ , Hg 2+ ), Tris-hydrochloric acid buffer solution (pH 7.4 and 8.8), XO (metal chelator xylenol orange) and the CPC-CDs solution prepared in Example 1 to establish an array sensing platform.

[0040] Set up heavy metal ion solutions with different heavy metal ions (Cr 6+ , Fe 2+ , Fe 3+ , Hg 2+ ), and the final concentration of each heavy metal ion solution in the fluorescence sensor array is set to a different concentration gradient (0, 1 μM, 10 μM, 20 μM, 30 μM, 40 μM, 50 μM);

[0041] CPC-CDs and XO were reacted in Tris-HCl buffer solution (pH 7.4 and 8.8), and the optimal reaction time of CPC-CDs and XO was 7 min. On this basis, metal ion solution was added, and the incubation time of CPC-CDs / XO sensing system for metal ions was 10 min ( Fig. 9 ).

[0042] 5 mL of a Tris-HCl solution with a pH of 7.4 and an initial concentration of 1 M and 3.3 mL of a Tris-HCl solution with a pH of 8.8 and an initial concentration of 1.5 M were diluted with deionized water to obtain two sets of pH detection environments;

[0043] The first group of pH test environments were: pH 7.4, 10 mM Tris-HCl buffer solution;

[0044] The second group of pH test environment is: pH 8.8, concentration 10mM Tris-HCl buffer solution;

[0045] Next, 950 μL of Tris-HCl buffer solution (two groups: 10 mM, pH 7.4, 950 μL / 10 mM, pH 8.8, 950 μL), 20 μL of CPC-CDs solution and 2 μL of 100 μM XO were mixed in a 1 mL centrifuge tube, mixed evenly and allowed to stand for 7 min to obtain CPC-CDs / XO sensing solutions under two pH environments;

[0046] Metal ions of different concentrations were added to the CPC-CDs / XO sensing solutions under two groups of pH environments, mixed evenly and allowed to stand for 10 min to obtain the CPC-CDs / XO sensing system. The fluorescence emission spectrum of CPC-CDs was recorded at an excitation wavelength of 370 nm to obtain a fluorescence sensor array.

[0047] In the detection system, Figure 3 The results show that the absorption spectra of the four metal ion complexes overlap well with the excitation spectra of CPC-CDs. Therefore, when heavy metal ion complexes are added to the two pH detection systems, the fluorescence intensity of CPC-CDs can be effectively reduced. Figure 4 The ultraviolet absorption spectra of CPC-CDs, four metal ion complexes, CPC-CDs+metal ion complex, and CPC-CDs and metal ion complex and their sum were measured, showing that the experimental curve (absorption spectrum of CPC-CDs / metal ion complex) was close to the theoretical curve (physical superposition of the curves of CPC-CDs solution and metal ion complex solution), indicating that no complex was formed between CPC-CDs and the metal ion complex.

[0048] The results showed that four heavy metals (Cr 6+ , Fe 2+ , Fe 3+ , Hg 2+ ) can effectively quench the IFE fluorescence intensity of CPC-CDs under the action of the small molecule linker. Therefore, the present invention uses a metal chelator as a small molecule linker, and monitors the fluorescence response changes of CPC-CDs while undergoing a chelation reaction with the metal under an environment in which the pH value of the system is adjusted. A sensor array with strong specificity and high sensitivity was produced. In order to verify the detection performance of the sensor array. First, we tested the fluorescence response of CPC-CDs in the range of pH 5-10 to obtain the optimal pH value for array detection. Figure 5The fluorescence response of CPC-CDs and CPC-CDs-XO in different pH buffer solutions shows that in the pH range of 5-10, the difference in the change of fluorescence emission intensity of CPC-CDs and CPC-CDs-XO at 535nm is small. It can be seen that XO has little effect on the fluorescence emission intensity of CPC-CDs, can be used to connect analytes and carbon dots, and has the advantage of enhancing the specificity of the detection system. We selected two pH Tris-buffer solutions (7.4 and 8.8), used XO as a small molecule linker, and made a sensor array for detecting heavy metals by monitoring the fluorescence changes of CPC-CDs. First, the fluorescence response of the sensor array to a single analyte was tested to verify the performance of the sensor system. Then, four heavy metals at a concentration of 10 μM were added to the two pH sensing units. As a result, the fluorescence intensity of CPC-CDs was quenched to varying degrees under the two pH sensing units. As Figure 6 ,The fluorescence response of metal ions and sensor array system under different pH environments (left pH 7.4, right pH 8.8) shows that the relative fluorescence response changes I / I0 of the four heavy metals are different, indicating that the sensor array can be used for the identification of heavy metals.

[0049] Next, the detection conditions were optimized to improve the detection performance of the sensor. First, a fluorescence titration experiment was performed to adjust the optimal concentration of XO in the array system ( Figure 7 ), add different concentrations of XO to the Tris-HCl buffer solution at pH = 7.4. As the concentration of XO increases, the fluorescence of CPC-CDs begins to quench. However, when the concentration of XO is too low, the chelation reaction between the XO and the metal ions is poor, and it is difficult to improve the specificity of the detection system. Therefore, the final concentration of XO in the fluorescence sensor array is 0.2 μM as the optimal reaction concentration. Secondly, the reaction time is optimized ( Figure 8 ), the fluorescence intensity of CPC-CDs-XO at 535nm increased with the increase of reaction time, and finally the optimal reaction time was found to be 7min.

[0050] The above is only a preferred embodiment of the present invention and does not limit the present invention in any way. Any simple modification, change and equivalent change made to the above embodiment according to the technical essence of the invention still falls within the protection scope of the technical solution of the present invention.

Claims

1. A method for preparing a fluorescent sensor array for detecting heavy metal ions, characterized in that: The method is: S1. Preparation of carbon quantum dots: at room temperature, transfer a sodium hydroxide solution to a cetylpyridinium chloride solution, perform water bath sonication, and obtain a reaction solution; add dichloromethane to the reaction solution, extract and purify, extract for 3 times in total, let stand until the solution is colorless and transparent, dialyze in deionized water with a dialysis bag to obtain carbon quantum dots, and store at 4°C in the dark; S2. Establishment of pH-regulated carbon quantum dot fluorescence sensor array: Two groups of pH detection environments were set up. The first group of pH detection environments was Tris-HCl buffer solution A with a pH of 7.4 and a concentration of 10 mM. The second group of pH detection environments was Tris-HCl buffer solution B with a pH of 8.8 and a concentration of 10 mM. 950 μL of Tris-HCl buffer solution, 20 μL of carbon quantum dots obtained in S1 and 2 μL of 100 μM xylenol orange solution were mixed evenly and allowed to stand for 7 min to obtain two groups of carbon quantum dots / xylenol orange sensing solutions under two pH environments; the Tris-HCl buffer solutions were Tris-HCl buffer solution A and Tris-HCl buffer solution B respectively; Different concentrations of metal ions were added to the carbon quantum dots / xylenol orange sensing solutions under the two groups of pH environments, mixed evenly and allowed to stand for 10 minutes to obtain a carbon quantum dots / xylenol orange sensing system. The fluorescence emission spectrum of the carbon quantum dots was recorded at an excitation wavelength of 370 nm to obtain a fluorescence sensor array.

2. The method for preparing a fluorescent sensor array for detecting heavy metal ions according to claim 1, characterized in that: The volume ratio of the cetylpyridinium chloride solution and the sodium hydroxide solution in S1 is 2:1, the concentration of the cetylpyridinium chloride solution is 50 mmol / L; the concentration of the sodium hydroxide solution is 330 mmol / L; the dosage ratio of the dichloromethane and the carbon quantum dots is (1~3):

1.

3. The method for preparing a fluorescent sensor array for detecting heavy metal ions according to claim 1, characterized in that: The temperature of the water bath ultrasound in S1 is 20°C to 40°C, and the time is 6h to 9h.

4. The method for preparing a fluorescent sensor array for detecting heavy metal ions according to claim 1, characterized in that: The dialysis bag in S1 has a MWCO membrane with a molecular weight cutoff of 3000Da, and the dialysis time is 24h~72h.

5. The method for preparing a fluorescent sensor array for detecting heavy metal ions according to claim 1, characterized in that: The final concentration of the xylenol orange solution in the fluorescent sensor array described in S2 is 0.2 μmol / L.

6. An application of a fluorescent sensor array prepared according to any one of claims 1 to 5 for detecting heavy metal ions, characterized in that: The fluorescent sensor array is used for the detection of heavy metal ions, and the heavy metal ions are Cr 6+ , Fe 2+ , Fe 3+ and Hg 2+ .

7. The use according to claim 6, characterized in that: The final concentration range of each heavy metal ion in the fluorescent sensor array is 0 μmol / L to 50 μmol / L.

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