Double-signal probe based on ratio fluorescence and visualization and application thereof

Abstract

The invention relates to a dual-signal probe based on ratiometric fluorescence and visualization and application thereof, and relates to the technical field of ratiometric fluorescence sensing. The double-signal probe comprises nitrogen-doped carbon dots and o-phenylenediamine. The nitrogen-doped carbon dots are synthesized through a one-step hydrothermal method, the nitrogen-doped carbon dots and o-phenylenediamine form the ratiometric fluorescent and visual dual-signal sensor, the redox reaction between copper ions and o-phenylenediamine and the fluorescence resonance energy transfer of the reaction product oxidized o-phenylenediamine to the nitrogen-doped carbon dots are ingeniously utilized, and the fluorescence resonance energy transfer of copper ions and o-phenylenediamine to the nitrogen-doped carbon dots is greatly improved. Therefore, extremely sensitive visual detection on the copper ions is realized, and the method is applied to sensitive visual detection on the content of the copper ions in industrial ethylene glycol and has a relatively high application prospect.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of ratiometric fluorescent sensing technology, and in particular to a dual-signal probe based on ratiometric fluorescence and visualization and application thereof. BACKGROUND

[0002] Ethylene glycol is an important basic chemical raw material, which is widely used in polyester fiber, antifreeze, lubricant and other fields. The traditional method for producing ethylene glycol is ethylene method, which mainly uses petroleum as raw material, and has high energy consumption and large cost fluctuation in the production process. Coal-based ethylene glycol technology mainly uses coal as raw material and is obtained by hydrogenation refining. Compared with the traditional method, coal-based ethylene glycol technology has significant advantages. However, new impurities are produced in the process of producing ethylene glycol by coal-based method. The production of these impurities not only affects the UV value of the product, but also makes it difficult for the downstream product to meet the standard requirements of the polyester grade product. Therefore, it is an urgent task to construct an effective method for detecting impurities produced in the coal-based ethylene glycol technology and to improve the intrinsic quality of the product.

[0003] Metal ions (Cu 2+ ) have been confirmed to be impurities produced in the process of producing coal-based ethylene glycol. At present, the detection of Cu 2+Methods for the determination of copper include atomic absorption spectrometry [A. M. Ure, Atomic absorption spectrometry for the determination of copper, 1 (1977) 1- 49.], inductively coupled plasma atomic emission spectrometry [A. M. Ure, Atomic absorption spectrometry for the determination of copper, 1 (1977) 1- 49.], inductively coupled plasma mass spectrometry [I. Gaubeur, M. A. Aguirre, N. Kovachev, M. Hidalgo, A. Canals, Dispersive liquid-liquid microextraction combined with laser-induced breakdown spectrometry and inductively coupled plasma optical emission spectrometry to elemental analysis, Microchemical Journal, 121 (2015) 219-226.], surface enhanced Raman scattering [Y. Guo, D. Li, S. Zheng, N. Xu, W. J. B. Deng, Bioelectronics, Utilizing Ag-Au core-satellite structures for colorimetric and surface-enhanced Raman scattering dual-sensing of Cu(II), 159 (2020) 112192. and H. Feng, Q. Fu, W. Du, R. Zhu, X. Ge, C. Wang, Q. Li, L. Su, H. Yang, J. J. A. n. Song, Quantitative assessment of copper(II) in Wilson’s disease based on photoacoustic imaging and ratiometric surface-enhanced Raman scattering, 15 (2021) 3402-3414.], chemiluminescence [J. Chen, Q. Wang, X. Liu, X. Chen, L. Wang, W. J. C. C. Yang, Black phosphorus quantum dots as novel electrogenerated chemiluminescence emitters for the detection of Cu 2+,56 (2020) 4680-4683.], electrochemical methods [X. Shao, H. Gu, Z. Wang, X. Chai, Y. Tian, G. Shi, Highly Selective Electrochemical Strategy for Monitoring of Cerebral Cu 2+ Based on a Carbon Dot-TPEAHybridized Surface, Analytical Chemistry, 85 (2013) 418-425.], colorimetric methods [S. Sun, X. Wu, Y. Huang, Q. Jiang, S. Zhu, S. Sun, Visual detection of Cu 2+in high-copper feed based on a fluorescent derivative of rhodamine B, Microchemical Journal, 171 (2021) 106858. and Z. Qing, Z. Mao, T. Qing, X. He, Z. Zou, D. He, H. Shi, J. Huang, J. Liu, K. Wang, Visual and Portable Strategy for Copper(II) Detection Based on a Striplike Poly(Thymine)-Caged and Microwell-Printed Hydrogel, Analytical Chemistry, 86 (2014) 11263-11268.], absorption spectroscopy [A. P. S. Gonzáles, M. A. Firmino, C. S. Nomura, F. R. P. Rocha, P. V. Oliveira, I. Gaubeur, Peat as a natural solid-phase for copper preconcentration and determination in a multicommuted flow system coupled to flame atomic absorption spectrometry, Analytica Chimica Acta, 636 (2009) 198-204.], electrochemistry [Y. Yan, F. Zhou, Q. Wang, Y. Huang, A sensitive electrochemical biosensor for quinolones detection based on Cu 2+ -modulated signal amplification, Microchemical Journal, 190 (2023) 108636.] and atomic emission [J. Otero-Romaní, A. A. Bermejo-Barrera, P. Bermejo-Barrera, Evaluation of commercial C18cartridges for trace elements solid phase extraction from seawater followed by inductively coupled plasma-optical emission spectrometry determination, Analytica Chimica Acta, 536 (2005) 213-218.]. However, these methods rely on instrumental detection, which has the problems of expensive instruments, complex operation, long time-consuming, tedious sample pretreatment process, etc., which is difficult to meet the analysis needs of industrial ethylene glycol. Fluorescence (FL) technology is favored by industry because of its simple operation, ultra-high sensitivity and visual advantages. However, most FL sensors focus on a single FL response mode, which is easily affected by some unavoidable factors such as environmental interference, incident light, probe concentration and biological background, and the accuracy and reliability of the single FL response mode are not high. Therefore, the double-ratio fluorescent probe based on fluorescent materials can greatly reduce background interference and further improve the quality of polyester products.

[0004] As a new type of carbon-based fluorescent nanomaterial with a diameter of less than 10 nm, carbon quantum dots have many unique properties, such as tunable and stable fluorescence emission, high water solubility, easy preparation, good biocompatibility and low toxicity [B.B. Chen, M.L. Liu, L. Zhan, C.M. Li, C.Z. Huang, Terbium(III) Modified Fluorescent Carbon Dots for Highly Selective and Sensitive Ratiometry of Stringent, Analytical Chemistry, 90 (2018) 4003-4009.]. Therefore, based on the specific cation exchange reaction between quantum dots and Cu 2+ A new type of Cu 2+ Sensor based on quantum dots has emerged. However, the manufacture of these ratiometric fluorescent sensors usually involves tedious or complex chemical modifications, which inevitably affect the quantum yield and stability. Therefore, it is still a challenge to develop a new type of direct and unmodified ratiometric fluorescent sensor for visual Cu 2+ Detection of coal-based ethylene glycol. In view of this, the present application provides a double-signal probe based on ratiometric fluorescence and visualization and its application. SUMMARY

[0005] The technical problem solved by the present application is to provide a dual signal probe based on ratio fluorescence and visualization and its application.

[0006] The technical solution of the present application to solve the above technical problems is as follows:

[0007] In the first aspect, the dual signal probe based on ratio fluorescence and visualization comprises nitrogen-doped carbon dots and o-phenylenediamine.

[0008] On the basis of the above technical solution, the present application can also be improved as follows.

[0009] Further, the mass ratio of the nitrogen-doped carbon dots to the o-phenylenediamine is (0.5-2):10.8.

[0010] Further, the nitrogen-doped carbon dots are prepared by the following steps:

[0011] Citric acid or citrate and ethylenediamine are added to ultrapure water for hydrothermal reaction, and then separated, and the supernatant is collected; the supernatant is dialyzed and dried to obtain nitrogen-doped carbon dots.

[0012] Further, the ratio of the amount of citric acid to the amount of ethylenediamine is (1-4)g:(0.5-2)mL; and the volume ratio of the ethylenediamine to the ultrapure water is (0.5-1):(30-50).

[0013] Further, the hydrothermal reaction conditions are as follows: the temperature is 180-200℃, and the time is 4-6h.

[0015] The dialysis conditions are as follows: the clear liquid is purified in a dialysis bag with a molecular weight cutoff of 500Da for (12-24)h, and the water is changed every (3-5)h.

[0016] In the second aspect, the application of the dual signal probe based on ratio fluorescence and visualization is used for detecting copper ions in industrial ethylene glycol.

[0017] Further, the industrial ethylene glycol comprises at least one of rich ethylene glycol, industrial-grade ethylene glycol and polyester-grade ethylene glycol.

[0018] In the third aspect, the sensing system for detecting copper ions in industrial ethylene glycol comprises Tris-HCl buffer, o-phenylenediamine and nitrogen-doped carbon dots.

[0019] Furthermore, the concentration of o-phenylenediamine in the sensing system is 0.5-2 mM, preferably 1 mM; the concentration of nitrogen-doped carbon dots in the sensing system is 5-10 μg / mL, preferably 10 μg / mL.

[0020] This invention constructs a dual-signal probe based on ratiometric fluorescence and visualization, namely nitrogen-doped carbon dots and o-phenylenediamine. This dual-signal probe can sensitively detect Cu in ethylene glycol. 2+ In this detection method, Cu 2+ It undergoes a redox reaction with o-phenylenediamine to form o-phenylenediamine oxide. O-phenylenediamine oxide not only exhibits a fluorescence signal at a wavelength of 570 nm but also shows an ultraviolet absorption peak at an absorption wavelength of 450 nm. Simultaneously, fluorescence resonance energy transfer occurs between the nitrogen-doped carbon dots and o-phenylenediamine oxide, quenching the fluorescence intensity of the nitrogen-doped carbon dots. Due to the reaction between o-phenylenediamine and Cu... 2+ The selective oxidation and colorimetric reactions of Cu were utilized to achieve a method targeting Cu 2+ A ratio fluorescence and visualization dual-signal sensing system for Cu 2+ It exhibits good selectivity and sensitivity, with a detection limit (LOD) of 76 nM. It was applied to the detection of Cu in industrial ethylene glycol (rich ethylene glycol, industrial-grade ethylene glycol, and polyester-grade ethylene glycol). 2+ Its recovery rate ranged from 95.95% to 107.8%. This ratio, along with the fluorescent and visual dual-signal probe, is suitable for the routine detection of Cu in industrial ethylene glycol. 2+ It offers a promising outlook.

[0021] The beneficial effects of this invention are:

[0022] (1) This invention synthesizes nitrogen-doped carbon dots through a one-step hydrothermal method and constructs a ratio fluorescence and visualization dual-signal sensor with o-phenylenediamine. It cleverly utilizes the redox reaction between copper ions and o-phenylenediamine, as well as the fluorescence resonance energy transfer of the reaction product oxidized o-phenylenediamine to nitrogen-doped carbon dots, thereby achieving highly sensitive visualization detection of copper ions. It is applied to the sensitive visualization detection of copper ion content in industrial ethylene glycol and has high application prospects.

[0023] (2) Compared with single-signal fluorescence detection, this method allows for clear visual identification of fluorescence color changes. It exhibits specific selectivity and anti-interference capabilities, a wide detection range, and a low detection limit. It has been successfully applied to Cu in ethylene glycol, industrial-grade ethylene glycol, and polyester-grade ethylene glycol. 2+ The detection. Attached Figure Description

[0024] Figure 1 Images of N-CDs are shown in electron microscope images; (a) is a TEM image, and (b) is a high-resolution transmission electron microscope image.

[0025] Figure 2 XPS for N-CDs; where (a) is the full spectrum, and (b) to (c) are high-resolution C1s, N1s, and O1s plots;

[0026] Figure 3 The images show the UV absorption and fluorescence emission spectra of N-CDs; where (a) is the UV absorption and fluorescence emission spectrum of N-CDs, and (b) is the fluorescence spectrum of N-CDs under different excitations.

[0027] Figure 4 For different concentrations of Cu 2+ Fluorescence intensity and ratio in the presence of Cu 2+ The relationship between concentrations is shown in the graph, where (a) Cu at different concentrations 2+ The following fluorescence intensity diagram exists, (b) ratio with Cu 2+ Concentration relationship graph;

[0028] Figure 5 Cu at different concentrations 2+ There are changes in ultraviolet absorption and ultraviolet absorption and Cu 2+ The relationship between concentrations; where (a) Cu at different concentrations 2+ There is a graph showing the change in ultraviolet absorption, (b) ultraviolet absorption and Cu 2+ A graph showing the relationship between concentrations. Detailed Implementation

[0029] The principles and features of this invention are described below. The examples given are for illustrative purposes only and are not intended to limit the scope of the invention. Where specific techniques or conditions are not specified in the embodiments, they should be performed according to the techniques or conditions described in the literature in this field, or according to the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased through legitimate channels.

[0030] Example

[0031] 1. Experimental methods.

[0032] (1) Preparation of nitrogen-doped carbon dots:

[0033] 2.0 g of citric acid was dispersed in 40 mL of ultrapure water and stirred until completely dissolved. Then, 1 mL of ethylenediamine was added to the solution. After thorough mixing, the mixture was placed in a stainless steel autoclave and subjected to a hydrothermal reaction at 200 °C for 5 h. After the reaction cooled to room temperature, the reaction product was centrifuged at 10000 r / min, and the supernatant was collected. This supernatant was dialyzed and then freeze-dried to obtain nitrogen-doped carbon dot powder (N-CDs). Finally, the freeze-dried powder was dissolved in distilled water to obtain a 1 mg / mL nitrogen-doped carbon dot solution for later use.

[0034] (2) Standard curve of copper ion concentration, fluorescence intensity ratio F570 / F450 and absorption intensity A450 was constructed:

[0035] Tris-HCl buffer solution, o-phenylenediamine solution and different concentrations of copper ion solution were uniformly mixed and incubated under certain conditions in the dark. Then, the mixture was fully mixed with nitrogen-doped carbon quantum dots and rested for 5 min to obtain a ratio fluorescence and visualization dual signal probe solution, which was the sensing system. The pH value of the Tris-HCl buffer solution was 7.4, and the amount used was 3 mL. The concentration of the o-phenylenediamine solution was 300 mM, and the amount was 10 μL. The incubation condition was incubation at 37 °C for 180 min. In the ratio fluorescence and visualization dual signal probe solution, the initial fluorescence intensity ratio (F570 / F450) of nitrogen-doped carbon dots and o-phenylenediamine was 1:9. The concentrations of the copper ion solution were 0 μM, 10 μM, 20 μM, 30 μM, 40 μM, 50 μM, 60 μM, 70 μM, 80 μM, 90 μM and 100 μM, and the volumes were 0, 3, 6, 9, 12, 15, 18, 21, 24, 27 and 30 μL, respectively. The fluorescence spectrum and absorption spectrum data containing different concentrations of copper ions were obtained by means of a fluorescence spectrophotometer and an ultraviolet absorption spectrophotometer, respectively, and then the standard linear relationship curves of copper ion concentration, fluorescence intensity ratio F570 / F450 and absorption intensity A450 were constructed.

[0036] (3) Detection of copper ion concentration in the sample:

[0037] The sample to be tested was filtered to remove insoluble impurities, mixed with the ratio fluorescence and visualization dual signal probe solution (without adding Cu 2+ ) obtained in the above (2) and rested for 5 min, and was subjected to standard addition treatment (10 μM, 20 μM and 30 μM). Then, the fluorescence intensity ratio F570 / F450 and the absorption intensity A450 of the sample to be tested were obtained by means of a fluorescence spectrophotometer and an ultraviolet absorption spectrophotometer, respectively, and the content of copper ions contained in the sample to be tested was quantitatively determined according to the standard linear relationship curves obtained in the above (2).

[0038] 2. Experimental results.

[0039] (1) Characterization of the structure and properties of nitrogen-doped carbon dots.

[0040] The structure and properties of nitrogen-doped carbon dots were characterized, as shown in a, which showed that N-CDs were spherical in shape and uniformly dispersed, with an average diameter of 2.6 nm. Figure 1 Figure 1 ​Figure b shows that the N-CDs have high crystallinity with a lattice spacing of 0.23 nm, corresponding to the (100) crystal plane of graphitic carbon. XPS further confirmed the presence of C, N, and O elements in the prepared N-CDs, with contents of 72.66%, 9.31%, and 18.03%, respectively. Figure 2 The "ad" in the text indicates the presence of carboxyl, hydroxyl, and amino functional groups. For example... Figure 3 As shown in Figures a and b, the characteristic absorption center of N-CDs is located near 340 nm, and the emission wavelength of N-CDs is located at 450 nm.

[0041] (2) Standard curves of copper ion concentration versus fluorescence intensity F570 / F450 ratio and absorption intensity A450:

[0042] Cu of different concentrations 2+ A mixed solution of Cu and o-phenylenediamine was added to N-CDs. The fluorescence intensity of the N-CDs themselves was not affected by Cu alone. 2+ And o-phenylenediamine has an effect. Furthermore, o-phenylenediamine alone does not exhibit fluorescence emission. When o-phenylenediamine and Cu... 2+ When coexisting, Cu 2+ It undergoes a redox reaction with o-phenylenediamine to form o-phenylenediamine oxide. O-phenylenediamine oxide exhibits fluorescence at 570 nm when excited at 340 nm, and the fluorescence intensity increases with Cu. 2+ The fluorescence intensity increases significantly with increasing Cu concentration. Simultaneously, fluorescence resonance energy transfer occurs between N-CDs and o-phenylenediamine oxide; at excitation of 340 nm, the fluorescence intensity of N-CDs is quenched, and increases further with increasing Cu concentration. 2+ With increasing concentration, the fluorescence intensity of N-CDs gradually decreases. Meanwhile, o-phenylenediamine shows no absorption peak at 450 nm in the ultraviolet region, while o-phenylenediamine oxide exhibits an absorption peak at this wavelength, and this peak decreases with increasing Cu concentration. 2+ The effect is significantly enhanced with increasing concentration. For example... Figure 4 As shown in Figure a, with Cu 2+ With increasing concentration, the fluorescence of N-CDs gradually decreases at 450 nm, while it gradually increases at 570 nm. For example... Figure 4 As shown in b, Cu 2+ The concentration change from 0 μM to 80 μM showed a good linear relationship with the fluorescence ratio F570 / F450, and the relationship was: F570 / F450 = 0.04549 + 0.01752 [Cu 2+ Correlation coefficient R 2 =0.992, Cu 2+ The detection limit (3σ / s) is 0.076 μM.

[0043] like Figure 5 As shown in Figure a, with Cu 2+With the increase of concentration, the UV absorption peak at 450 nm gradually increased. Figure 5 As shown in b, the Cu 2+ concentration in the range of 0-40 μM, the absorbance at 450 nm showed a good linear relationship, and the relationship was: A450=-0.01353+0.01633[Cu 2+ ](R 2 =0.992). In the range of Cu 2+ concentration of 40 μM-90 μM, the absorbance at 450 nm showed a good linear relationship, and the relationship was: A450=0.31424+0.0101[Cu 2+ ](R 2 =0.991).

[0044] (3) The detection results of copper ion concentration in the sample.

[0045] The method of constructing N-CDs fluorescent probe for detecting Cu 2+ was applied to the actual sample for detecting Cu 2+ . Randomly selected three kinds of samples of ethylene glycol, industrial grade ethylene glycol and polyester grade ethylene glycol were diluted 100 times, and the interference substances in the sample were eliminated by dilution. As shown in Table 1, the recovery rate was 95.79%-107.8%. The results showed that the method can be used for detecting Cu 2+ in the actual sample.

[0046] Table 1

[0047]

[0048] As can be seen from the above, the nitrogen-doped carbon dots and the ratio fluorescence and visual double signal probe of o-phenylenediamine are successfully prepared, which ingeniously utilizes the redox reaction between copper ions and o-phenylenediamine, and the fluorescence resonance energy transfer of the reaction product oxidized o-phenylenediamine to nitrogen-doped carbon dots, so as to realize the visual detection of copper ions. The sensing system has good selectivity and sensitivity for Cu 2+ , and the detection limit is 76 nM; it is applied to the detection of Cu 2+ in industrial ethylene glycol (ethylene glycol, industrial grade ethylene glycol and polyester grade ethylene glycol), and the recovery rate is between 95.79% and 107.8%. The ratio fluorescence and visual double signal probe provides a good prospect for the conventional detection of Cu 2+ in industrial ethylene glycol.

[0049] Although the embodiments of the present application have been shown and described above, it is understood that the above-described embodiments are exemplary and are not to be construed as limiting the present application, and that changes, modifications, substitutions and variations can be made by those skilled in the art without departing from the scope of the present application.

Claims

1. A dual signal probe based on ratiometric fluorescence and visualization, characterized in that, The dual signal probe comprises nitrogen-doped carbon dots and o-phenylenediamine.

2. The dual signal probe based on ratiometric fluorescence and visualization according to claim 1, wherein, The mass ratio of the nitrogen-doped carbon dots to the o-phenylenediamine is (0.5-2):10.

8.

3. The dual signal probe based on ratiometric fluorescence and visualization according to claim 1, wherein, The nitrogen-doped carbon dots are prepared by the following steps: Citric acid or citrate and ethylenediamine are added into ultrapure water for hydrothermal reaction, and then separated, and the supernatant is collected; the supernatant is dialyzed, and then dried to obtain the nitrogen-doped carbon dots.

4. The dual signal probe based on ratiometric fluorescence and visualization according to claim 3, wherein, The dosage ratio of the citric acid to the ethylenediamine is (1-4)g:(0.5-2)mL; and the volume ratio of the ethylenediamine to the ultrapure water is (0.5-1):(30-50).

5. The dual signal probe based on ratiometric fluorescence and visualization according to claim 3, wherein, The hydrothermal reaction is carried out at a temperature of 180-200℃ for 4-6h.

6. Use of a dual signal probe based on ratiometric fluorescence and visualization, characterized in that, The dual signal probe based on ratio fluorescence and visualization according to any one of claims 1-5 is used for detecting copper ions in industrial ethylene glycol.

7. The use of a dual signal probe based on ratiometric fluorescence and visualization according to claim 6, characterized in that, The industrial ethylene glycol comprises at least one of rich ethylene glycol, industrial-grade ethylene glycol and polyester-grade ethylene glycol.

8. A sensing system for the detection of copper ions in industrial ethylene glycol, characterized by, The sensing system comprises Tris-HCl buffer and the dual signal probe based on ratio fluorescence and visualization according to any one of claims 1-5.

9. The sensing system for the detection of copper ions in industrial ethylene glycol according to claim 8, characterized by the fact that, The concentration of the o-phenylenediamine in the sensing system is 0.5-2mM; and the concentration of the nitrogen-doped carbon dots in the sensing system is 5-10μg / mL.

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