A fluorescence detection method for bipyridine compounds

Abstract

The present invention provides a fluorescence detection method for bipyridine compounds. This method utilizes sulfonic acid groups and biphenyl structures on the surface of DSD acid-functionalized carbon quantum dots as recognition elements, enabling the DSD acid-functionalized carbon quantum dots to generate electrostatic and π-π interactions with compounds containing bipyridine structures. This static quenching mechanism quenches the fluorescence of the DSD acid-functionalized carbon quantum dots, thereby enabling the DSD acid-functionalized carbon quantum dots to act as a highly sensitive fluorescence sensor for detecting bipyridine compounds. Given the significant difference in binding ability between paraquat and DSD acid-functionalized carbon quantum dots in their excited and ground states, while no significant difference exists for diquat, the present fluorescence detection method can also be used to detect the presence of paraquat in diquat herbicide samples.

Description

Technical Field

[0001] The present disclosure relates to the technical field of detection of bipyridine compounds, and in particular to a fluorescence detection method for bipyridine compounds. Background Art

[0002] Environmental problems are largely caused by environmental pollutants, which also have direct impacts on ecosystems and human health. Accurately measuring trace pollutants is crucial because, despite their low concentrations in the environment, they can pose a threat to human health and ecosystems.

[0003] Paraquat and diquat are both bipyridine herbicides that are highly toxic to mammals. Due to the lack of effective treatment, poisoning from these pesticides often attracts widespread attention. After accidental ingestion of bipyridine herbicides, the pesticide molecules are rapidly absorbed by the body and ultimately enter tissues to exert their toxic effects. Therefore, prompt diagnosis and measures such as gastric lavage and hemoperfusion are essential, placing high demands on rapid diagnosis of poisoning. Furthermore, while paraquat is currently banned, diquat remains permitted. Due to its high efficacy, paraquat has been illegally mixed with diquat, leading to the need for adulteration in herbicides. Currently, the detection of paraquat and diquat mostly relies on large-scale instrumentation, using liquid chromatography-mass spectrometry to detect paraquat and diquat residues. This presents challenges such as expensive instrumentation, the need for specialized personnel, and lengthy testing times. The sodium dithionite colorimetric method is used clinically as a rapid detection method for paraquat and diquat poisoning, but the detection limit of this method is high. In particular, when the drug quickly enters the tissue from the blood, this method can no longer detect the pesticide.

[0004] Carbon quantum dots (CQDs) are excellent nanomaterials characterized by stable chemical properties, good water solubility, excellent biocompatibility, and superior fluorescence. The surface of CQDs is typically rich in functional groups, providing ample sites for further modification to enhance selectivity. The amide bond is a relatively stable chemical bond, and the amidation reaction catalyzed by N-hydroxysuccinimide (NHS) / 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC) can proceed at room temperature, making it widely applicable.

[0005] 4,4'-Diaminostilbene-2,2'-disulfonic acid (DSD acid) is a commonly used fluorescent dye intermediate. It has a planar molecular structure with conjugated double bonds and benzene rings forming a large conjugated system. It also contains amino groups as electron-donating groups, providing potential functional groups for subsequent functionalization. In some processes, it is converted into sodium DSD acid and used as a precipitant for complexing with paraquat cations in paraquat wastewater production. However, this process is generally only used for low-volume reactions and is ineffective for trace amounts of paraquat, failing to produce a precipitate. Summary of the Invention

[0006] The present disclosure aims to solve at least one of the technical problems existing in the prior art.

[0007] To this end, the first aspect of the present disclosure provides a fluorescence detection method for bipyridine compounds, which uses DSD acid-functionalized carbon quantum dots as probes to detect bipyridine compounds. The method has high sensitivity and good selectivity.

[0008] The first aspect of the present disclosure provides a fluorescence detection method for bipyridine compounds, comprising:

[0009] DSD acid-functionalized carbon quantum dots, solvent A, and standard solutions containing different concentrations of bipyridine compounds were mixed to obtain multiple homogeneous solutions B. Each homogeneous solution B was subjected to fluorescence excitation to determine the functional relationship between the concentration of the bipyridine compound and the fluorescence intensity.

[0010] The DSD acid-functionalized carbon quantum dots, the solvent A, and the test sample extract containing the bipyridine compound are mixed to obtain a homogeneous solution C, the solution C is subjected to fluorescence excitation, and the concentration of the bipyridine structure compound contained in the test sample is obtained using the functional relationship between the concentration of the bipyridine structure compound and the fluorescence intensity;

[0011] The DSD acid functionalized carbon quantum dots are prepared according to the following steps:

[0012] The sulfur-nitrogen co-doped carbon quantum dots are mixed with solvent D, and an amidation reaction is carried out under the action of a catalyst and DSD acid. The product obtained by the amidation reaction is dialyzed and dried to obtain the DSD acid-functionalized carbon quantum dots.

[0013] In some embodiments, the solvent A is a mixture of any one or more of water, ethanol, acetonitrile and dimethyl sulfoxide.

[0014] In some embodiments, the standard of the bipyridine compound is paraquat, diquat, 4,4′-bipyridine, N-methyl-4,4′-bipyridine or 2,2′-bipyridine.

[0015] In some embodiments, the test article extract is selected from water samples of rice, corn, sunflower seeds, cotton seeds, blood, urine or river water.

[0016] In some embodiments, the mass ratio of the DSD acid-functionalized carbon quantum dots, the solvent A, and the standard solution is 1:100-10000000:0.05-100.

[0017] In some embodiments, the mass ratio of the DSD acid-functionalized carbon quantum dots, the solvent A, and the test article extract is 1:100-100000000:0.01-1000.

[0018] In some embodiments, the excitation wavelength is 320 nm to 360 nm.

[0019] In some embodiments, the duration of the excitation is t∈(0s, 1200s], and the excitation can be performed continuously or discontinuously.

[0020] In some embodiments, the molar ratio of the sulfur-nitrogen co-doped carbon quantum dots, the solvent D, the catalyst, and the DSD acid is 1:5000-200000:1-20:1-10.

[0021] In some embodiments, the sulfur-nitrogen co-doped carbon quantum dots are mixed with solvent D and subjected to an amidation reaction in the presence of a catalyst and DSD acid, comprising:

[0022] The sulfur-nitrogen co-doped carbon quantum dots, the solvent D and the catalyst are activated for 10 to 30 minutes to obtain an activated product;

[0023] The activated product is mixed with the DSD acid and subjected to an amidation reaction at room temperature for 3 to 24 hours.

[0024] In some embodiments, the product obtained from the amidation reaction is dialyzed for 12 hours to 48 hours.

[0025] Furthermore, the product obtained by the amidation reaction is dialyzed, and the solvent D is regularly replaced during the dialysis process to obtain a DSD acid-functionalized carbon quantum dot solution.

[0026] Furthermore, the DSD acid-functionalized carbon quantum dots solution is subjected to rotary evaporation to remove most of the solvent and then freeze-dried to obtain the DSD acid-functionalized carbon quantum dots.

[0027] Furthermore, the solvent D is a mixture of any one or more of deionized water, ethanol and N,N-dimethylformamide.

[0028] Furthermore, the catalyst is selected from a mixture of any one or more of N,N'-diisopropylcarbodiimide, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide, dicyclohexylcarbodiimide, 4-dimethylaminopyridine, carbodiimide hydrochloride, 1-hydroxybenzotriazole and 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate.

[0029] In some embodiments, the sulfur-nitrogen co-doped carbon quantum dots are prepared by a hydrothermal method, comprising the following steps:

[0030] dissolving a carbon source, a nitrogen source, and a sulfur source in solvent E and performing ultrasonic treatment to form a uniform solution A;

[0031] subjecting the solution A to a hydrothermal reaction to obtain a solution B;

[0032] The solution B is dialyzed and dried to obtain the sulfur-nitrogen co-doped carbon quantum dots.

[0033] Furthermore, the carbon source is citric acid.

[0034] Furthermore, the nitrogen source and sulfur source are selected from L-cysteine.

[0035] Furthermore, the solvent E is deionized water.

[0036] Furthermore, the temperature of the hydrothermal reaction is 160° C. to 200° C., and the reaction time is 2 h to 4 h.

[0037] Furthermore, the dialyzation time of the solution B is 48 hours to 96 hours.

[0038] Furthermore, the solvent E is regularly replaced during the dialysis of the solution B, and a sulfur-nitrogen co-doped carbon quantum dot solution is obtained after dialysis.

[0039] Furthermore, the sulfur-nitrogen co-doped carbon quantum dots solution is subjected to rotary evaporation to remove most of the solvent and then freeze-dried to obtain the sulfur-nitrogen co-doped carbon quantum dots.

[0040] Furthermore, the molar ratio of the carbon source, the nitrogen source and the sulfur source, and the solvent E is 1:0.2-0.8:500-2000.

[0041] The first embodiment of the present disclosure provides a fluorescence detection method for bipyridine compounds, which has the following characteristics and

[0042] Beneficial effects:

[0043] The disclosed embodiments utilize sulfonic acid groups and biphenyl structures on the surface of DSD acid-functionalized carbon quantum dots as recognition elements, enabling the DSD acid-functionalized carbon quantum dots to generate electrostatic and π-π interactions with compounds containing bipyridine structures. This static quenching mechanism quenches the fluorescence of the DSD acid-functionalized carbon quantum dots, thereby enabling the DSD acid-functionalized carbon quantum dots to act as a highly sensitive fluorescence sensor for detecting compounds containing bipyridine structures. Because the binding energy of bipyridine compounds to the excited state of DSD acid-functionalized carbon quantum dots is greater than that to the ground state (particularly, this property of paraquat is more pronounced than that of diquat, 4,4′-bipyridine, N-methyl-4,4′-bipyridine, and 2,2′-bipyridine), a period of fluorescence excitation can effectively promote the quenching of the fluorescence of the DSD acid-functionalized carbon quantum dots by bipyridine compounds, enhancing the detection effect.

[0044] A fluorescence detection method for bipyridine compounds provided in an embodiment of a second aspect of the present disclosure is specifically used to detect whether a diquat herbicide sample contains paraquat, comprising the following steps:

[0045] Diluting the diquat herbicide sample with water according to its nominal mass concentration to obtain a diluent G, mixing the DSD acid-functionalized carbon quantum dots, solvent A, and the diluent G to obtain a homogeneous solution H, subjecting the homogeneous solution H to fluorescence excitation, measuring a time-resolved spectrum of the homogeneous solution H, and determining whether the diquat herbicide sample contains paraquat using a functional relationship between fluorescence intensity and time;

[0046] The DSD acid functionalized carbon quantum dots are prepared according to the following steps:

[0047] The sulfur-nitrogen co-doped carbon quantum dots are mixed with solvent D, and an amidation reaction is carried out under the action of a catalyst and DSD acid. The product obtained by the amidation reaction is dialyzed and dried to obtain the DSD acid-functionalized carbon quantum dots.

[0048] In some embodiments, the dilution factor is: mass concentration × 10 10 ~mass concentration × 10 8 .

[0049] In some embodiments, the mass ratio of the DSD acid-functionalized carbon quantum dots to the solvent A is 1:100-10,000,000, and the mass ratio of the solvent A to the diluent G is 1:0.1-10.

[0050] In some embodiments, the relationship between fluorescence intensity and time function is used to determine whether a diquat herbicide sample contains paraquat, specifically:

[0051] Two points are selected from the time-resolved spectrum in chronological order, with a time difference of Δt between the two points. The fluorescence intensities of the two points are F1 and F2, respectively. When the ratio of F2 to F1 exceeds a set threshold, it is considered that the diquat herbicide sample does not contain paraquat; otherwise, it is considered that the diquat herbicide sample contains paraquat. The measurement time of the time-resolved spectrum is 10s to 1200s.

[0052] In some embodiments, the wavelength of fluorescence excitation is 320 nm to 360 nm, and the excitation mode is continuous excitation or intermittent excitation.

[0053] A fluorescence detection method for bipyridine compounds provided in the second embodiment of the present disclosure has the following characteristics and

[0054] Beneficial effects:

[0055] The disclosed embodiments utilize paraquat's ability to bind to DSD acid-functionalized carbon quantum dots in the excited state, which is much better than its ability to bind to the ground state. However, there is no significant difference in the binding ability of diquat to DSD acid-functionalized carbon quantum dots between the excited and ground states. Consequently, time-resolved spectroscopy reveals that the fluorescence intensity of DSD acid-functionalized carbon quantum dots does not change with irradiation time after diquat addition, while that of paraquat decreases significantly over time. This difference can be used to identify whether paraquat has been incorporated into diquat. This method can significantly and sensitively distinguish whether diquat has been adulterated with paraquat, offers rapid detection, and is cost-effective. It can be further developed into a visual colorimetric method for real-time, on-site detection of pesticides. BRIEF DESCRIPTION OF THE DRAWINGS

[0056] Figure 1 This is the working curve of Example 1 for detecting paraquat, a compound containing a bipyridine structure, using DSD acid-functionalized carbon quantum dots.

[0057] Figure 2 This is a transmission electron microscope lattice diffraction image of the DSD acid-functionalized carbon quantum dots prepared in Example 1. The morphology and lattice spacing of the carbon quantum dots can be seen.

[0058] Figure 3 This is a transmission electron microscope image of the DSD acid-functionalized carbon quantum dots prepared in Example 1, from which the morphology of the carbon quantum dots can be seen.

[0059] Figure 4 This is the X-ray diffraction spectrum of the DSD acid-functionalized carbon quantum dots prepared in Example 1.

[0060] Figure 5 This is the infrared spectrum of the DSD acid functionalized carbon quantum dots prepared in Example 1. Figure 5 Medium 1699cm -1 and 1552cm -1The absorption at corresponds to the amide I peak and the amide II peak, indicating that the formation of amide bonds enables the DSD acid to covalently bind to the carbon quantum dots.

[0061] Figure 6 This is the change in fluorescence intensity after the DSD acid-functionalized carbon quantum dots and paraquat are mixed under light excitation in Example 1, which is used to illustrate that the excitation time is an important reference factor in this detection scheme.

[0062] Figure 7 This is the working curve of Example 2 for detecting diquat containing a bipyridine structure using DSD acid-functionalized carbon quantum dots.

[0063] Figure 8 This is the working curve of Example 3 using DSD acid-functionalized carbon quantum dots to detect 2,2′-bipyridine, a compound containing a bipyridine structure.

[0064] Figure 9 This is Example 4, in which DSD acid-functionalized carbon quantum dots are used to detect time-resolved spectra of a diquat sample and a diquat sample spiked with paraquat. DETAILED DESCRIPTION

[0065] In order to make the purpose, technical solutions and advantages of this application more clearly understood, this application is further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain this application and are not intended to limit this application.

[0066] On the contrary, this application covers any alternatives, modifications, equivalents, and solutions made within the spirit and scope of this application as defined by the claims. Furthermore, to facilitate a better understanding of this application, certain specific details are described in detail below in the detailed description of this application. Those skilled in the art will be able to fully understand this application without these details.

[0067] The embodiments of the present application are described in detail below.

[0068] The technical solution disclosed herein is not limited to the specific embodiments listed below, but also includes various applications of using the DSD acid structure for identifying compounds containing a bipyridine structure.

[0069] Example 1:

[0070] The fluorescence detection method for bipyridine compounds provided in Example 1 specifically uses DSD acid-functionalized carbon quantum dots to detect the concentration of paraquat in a test sample, and specifically includes the following steps:

[0071] 1. Detection of paraquat concentration in standard: Take water as solvent A, mix DSD acid functionalized carbon quantum dots and solvent A at a mass ratio of 1:100000 to obtain DSD acid functionalized carbon quantum dot solution, take 100 μL of DSD acid functionalized carbon quantum dot solution, and mix it with 900 μL of aqueous solutions of paraquat of different concentrations, then continuously irradiate the mixture for 30 seconds at an excitation wavelength of 343 nm, and record the fluorescence spectra of paraquat of different concentrations and the fluorescence intensity at 417 nm. Among them, the fluorescence intensity when no paraquat is added is recorded as F0, and the fluorescence intensity after adding paraquat is recorded as F. According to the results measured after the interaction between paraquat of different concentrations and DSD acid functionalized carbon quantum dots, a graph of paraquat concentration x versus y = F0 / F (ratio of fluorescence intensity) is made, and the working curve for detecting paraquat concentration is obtained according to the fitting results, see Figure 1 The equation of the working curve is: y = 0.8498x + 1.0139, and the linearity of the working curve is R 2 =0.9979, greater than 0.99.

[0072] 2. Paraquat Concentration in the Test Article: Take 2g of blood (urine or river water samples may also be used) and extract with 2mL of methanol. Take 900μL of the test article extract and add 100μL of DSD acid-functionalized carbon quantum dot solution. Test the test article using the paraquat concentration method described in Step 1 above. Calculate the paraquat concentration in the test article using the working curve method.

[0073] Furthermore, in this Example 1, DSD acid-functionalized carbon quantum dots were prepared according to the following steps:

[0074] 1) Preparation of sulfur-nitrogen co-doped carbon quantum dots: 2 g of citric acid, 1 g of L-cysteine, and 5 mL of deionized water were mixed and added to a hydrothermal reactor. The reaction time was set to 2 h and the temperature was set to 200°C. After the reaction was completed, the mixture was cooled to room temperature and the product was filtered through a microporous filter membrane to obtain a yellow-brown solution. The solution was dialyzed through a dialysis bag (3500 Da), concentrated by rotary evaporation, and the dialyzate was freeze-dried to obtain sulfur-nitrogen co-doped carbon quantum dots.

[0075] 2) Preparation of DSD acid-functionalized carbon quantum dots: 3 mg of sulfur-nitrogen co-doped carbon quantum dots obtained in step 1) were dissolved in 30 mL of deionized water, 50 mg of catalyst N,N'-diisopropylcarbodiimide and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide were added, and the reaction was continued for 15 min. Then 100 mg of DSD acid was added and the reaction was continued for 24 h. The product was filtered through a 0.22 μm microporous filter membrane to obtain a yellow-brown solution; dialyzed through a dialysis bag (3500 Da), concentrated by rotary evaporation, and the dialyzate was freeze-dried to obtain DSD acid-functionalized carbon quantum dots.

[0076] Figures 2 to 5 This is the characterization result of DSD acid functionalized carbon quantum dots. Figure 2 This is a high-resolution transmission electron microscope image showing that DSD acid-functionalized carbon quantum dots have obvious graphite lattice characteristics, and their core is composed of regularly arranged carbon atoms. Figure 3 This is a transmission electron microscope image, which shows that the DSD acid-functionalized carbon quantum dots have a nearly circular morphology and an average particle size of 5.27±1.31nm. Figure 4 This is the X-ray diffraction pattern of DSD acid-functionalized carbon quantum dots, which shows a broad diffraction peak centered at 24.2°, which is the diffraction peak of graphite and also indicates that it has the structural characteristics of graphite. Figure 5 This is the infrared spectrum of DSD acid functionalized carbon quantum dots. The main information is 1699cm -1 and 1552cm -1 The absorption at corresponds to the amide I peak and the amide II peak, indicating that the formation of amide bonds enables the DSD acid to covalently bind to the carbon quantum dots.

[0077] The preparation method of the DSD acid functionalized carbon quantum dots provided in the present embodiment 1 utilizes a hydrothermal method to react citric acid with L-cysteine ​​to form a structure of highly conjugated carbon quantum dots. At the same time, the incorporation of heteroatoms such as sulfur and nitrogen further enhances the fluorescence properties of the carbon quantum dots by changing the molecular energy level, resulting in bright blue fluorescence. At the same time, the raw materials used are easy to obtain and have extremely low cost. The product has good water solubility and is non-toxic at working concentrations. The DSD acid functionalized carbon quantum dots prepared by this method are covalently linked to the carbon quantum dots by an amidation reaction. This modification of the carbon quantum dot surface can produce conjugation with the carbon quantum dots without destroying the fluorescence properties. It also causes a large number of DSD acid groups to exist on the carbon quantum dot surface, which can provide a structural basis for subsequent electrostatic interactions and π-π interactions.

[0078] Furthermore, in order to explore the effect of the excitation duration of fluorescence excitation on the detection effect of paraquat, without changing other parameters of this example, the changes in the fluorescence intensity of the DSD acid functionalized carbon quantum dots and paraquat in Example 1 under different fluorescence excitation durations were measured. The measurement results are shown in Figure 6 .Depend on Figure 6It can be seen that the fluorescence intensity gradually decreases with the extension of the excitation time. To explain this phenomenon, quantum chemical DFT and TD-DFT methods were used to calculate the binding energies of DSD acid-functionalized carbon quantum dots and paraquat in the ground state and excited state, respectively, and ghost atom processing was used to reduce the basis set superposition error. The results show that this phenomenon is caused by the fact that the binding energy of paraquat with the excited state of DSD acid-functionalized carbon quantum dots is greater than that with the ground state (diquat, 4,4′-bipyridine, N-methyl-4,4′-bipyridine and 2,2′-bipyridine also have this characteristic. The calculation method of binding energy is the same as that of paraquat and will not be repeated here). Under the premise of a certain paraquat concentration, the fluorescence can be better quenched with the extension of the excitation time, thus making the detection method more sensitive for paraquat. Given that paraquat and DSD acid-functionalized carbon quantum dots are based on a static quenching mechanism, this excitation is not limited to continuous excitation of light, and intermittent excitation can also achieve a similar effect of improving detection ability.

[0079] Example 2:

[0080] The fluorescence detection method for bipyridine compounds provided in Example 2 specifically uses DSD acid-functionalized carbon quantum dots to detect the concentration of diquat in the test sample, and specifically includes the following steps:

[0081] 1. Detection of diquat concentration in standard: Take dimethyl sulfoxide as solvent A, mix DSD acid functionalized carbon quantum dots and solvent A in a mass ratio of 1:1000 to obtain DSD acid functionalized carbon quantum dot solution, take 50 μL of DSD acid functionalized carbon quantum dot solution, and mix it with 950 μL of aqueous solutions of paraquat of different concentrations, then continuously irradiate the mixture for 1200 s at an excitation wavelength of 360 nm, and record the fluorescence spectra of diquat of different concentrations and the fluorescence intensity at 417 nm. Among them, the fluorescence intensity when diquat is not added is recorded as F0, and the fluorescence intensity after adding diquat is recorded as F. According to the results measured after the interaction between diquat of different concentrations and DSD acid functionalized carbon quantum dots, a graph of diquat concentration x versus y = F0 / F (ratio of fluorescence intensity) is made, and the working standard curve for detecting diquat concentration is obtained according to the fitting results, see Figure 7 The equation of the working curve is: y = 0.1262x + 0.9972, and the linearity of the working curve is R 2 =0.9972, greater than 0.99.

[0082] 2. Determination of Diquat Concentration in the Test Article: Extract 2g of sunflower seeds (rice or cotton seeds may also be used) with 2mL of ethanol. Take 950μL of the test article extract and add 50μL of DSD acid-functionalized carbon quantum dot solution. Test the test article using the diquat concentration method described in Step 1 above. Calculate the diquat concentration in the test article using the working curve method.

[0083] Furthermore, in this Example 2, DSD acid-functionalized carbon quantum dots were prepared according to the following steps:

[0084] 1) Preparation of sulfur-nitrogen co-doped carbon quantum dots: 2 g of citric acid, 1 g of L-cysteine ​​and 5 mL of deionized water were mixed and added to a hydrothermal reactor. The reaction time was set to 4 h and the temperature was set to 160°C. After the reaction was completed, the solution was cooled to room temperature and filtered through a microporous filter membrane to obtain a yellow-brown solution. The solution was dialyzed through a dialysis bag (3500 Da), concentrated by rotary evaporation, and the dialyzate was freeze-dried to obtain sulfur-nitrogen co-doped carbon quantum dots.

[0085] 2) Preparation of DSD acid-functionalized carbon quantum dots: 10 mg of sulfur-nitrogen co-doped carbon quantum dots obtained in step 1) was dissolved in 30 mL of ethanol, 10 mg of catalyst dicyclohexylcarbodiimide and 100 mg of catalyst 4-dimethylaminopyridine were added, the reaction was continued for 15 min, and then 50 mg of DSD acid was added and the reaction was continued for 12 h. The product was filtered through a 0.22 μm microporous filter membrane to obtain a yellow-brown solution; dialyzed through a dialysis bag (3500 Da), concentrated by rotary evaporation, and the dialyzate was freeze-dried to obtain DSD acid-functionalized carbon quantum dots.

[0086] Example 3:

[0087] The fluorescence detection method for bipyridine compounds provided in Example 3 specifically uses DSD acid-functionalized carbon quantum dots to detect the concentration of 2,2′-bipyridine contained in the test sample, and specifically includes the following steps:

[0088] 1. Detection of 2,2′-bipyridine concentration in the standard: Take ethanol as solvent A, mix DSD acid-functionalized carbon quantum dots and solvent A in a mass ratio of 1:10000 to obtain a DSD acid-functionalized carbon quantum dot solution, take 500 μL of the DSD acid-functionalized carbon quantum dot solution, and mix it with 500 μL of an aqueous solution of paraquat of different concentrations (the preparation method of DSD acid-functionalized carbon quantum dots in this embodiment is the same as that in Example 1), then intermittently irradiate the mixed solution for 10 seconds at an excitation wavelength of 320 nm, and record the fluorescence spectrum of 2,2′-bipyridine of different concentrations and the fluorescence intensity at 417 nm. Among them, the fluorescence intensity when no 2,2′-bipyridine is added is recorded as F0, and the fluorescence intensity after adding 2,2′-bipyridine is recorded as F. Based on the results of the interaction between different concentrations of 2,2′-bipyridine and DSD acid functionalized carbon quantum dots, a graph of 2,2′-bipyridine concentration x versus y = F0 / F (ratio of fluorescence intensity) was prepared, and a working curve for detecting 2,2′-bipyridine concentration was obtained based on the fitting results. Figure 8 The equation of the working curve is: y = 0.0136x + 0.99, and the linearity of the working curve is R 2 =0.9954, greater than 0.99.

[0089] Example 4:

[0090] The fluorescence detection method for bipyridine compounds provided in Example 3 specifically utilizes DSD acid-functionalized carbon quantum dots to identify whether a diquat herbicide sample contains paraquat, and specifically includes the following steps:

[0091] A 1 g sample labeled as diquat herbicide (nominal mass concentration of 20%) was diluted with water to a gradient dilution concentration of 0.05 mg / L of the active ingredient. 900 μL of the diluent was added to 100 μL of a 10 mg / L DSD acid-functionalized carbon quantum dot solution (prepared in the same manner as in Example 1) to obtain a homogeneous solution H. The homogeneous solution H was subjected to fluorescence excitation, and a time-resolved spectrum of the homogeneous solution H was measured, with the fluorescence intensity recorded once per second. Specifically, the homogeneous solution H was irradiated continuously for 30 seconds at an excitation wavelength of 343 nm. The fluorescence intensity 1 second after the start of irradiation was recorded as F1, and the fluorescence intensity at 30 seconds (i.e., Δt = 29 seconds) was recorded as F2. If the F2 / F1 value is greater than the set threshold, the set threshold is obtained by fitting to be 0.99-0.00014×Δt. Considering that in actual applications, trace amounts of paraquat will not be adulterated with diquat, the set threshold is set to 0.95. That is, when F2 / F1>0.95, it can be considered that the sample does not contain paraquat, but only diquat. Otherwise, it indicates that paraquat has been adulterated in the herbicide.

[0092] In Example 4, the excited state binding ability of paraquat with DSD acid functionalized carbon quantum dots is much better than that with the ground state, while there is no significant difference in the excited state binding ability of diquat with DSD acid functionalized carbon quantum dots and the ground state. Therefore, in the time-resolved spectrum, the fluorescence intensity of DSD acid functionalized carbon quantum dots does not change with the irradiation time after the addition of diquat, while the fluorescence intensity of paraquat decreases significantly with time. The test results are shown in Figure 9 , this difference can be used to identify whether paraquat is mixed with diquat.

[0093] In summary, the detection method of bipyridine compounds provided by the present disclosure is a method for detecting bipyridine compounds using DSD acid-functionalized carbon quantum dots. Specifically, the sulfonic acid groups, biphenyl structures and other groups contained on the surface of the DSD acid-functionalized carbon quantum dots are used as recognition elements, so that the DSD acid-functionalized carbon quantum dots can generate electrostatic interactions and π-π interactions with compounds containing bipyridine structures, and the fluorescence of the DSD acid-functionalized carbon quantum dots is quenched through a static quenching mechanism, thereby realizing the high sensitivity detection of compounds containing bipyridine structures by using the DSD acid-functionalized carbon quantum dots as a fluorescence sensor. At the same time, since the binding energy of bipyridine compounds to the excited state of DSD acid-functionalized carbon quantum dots is greater than that to the ground state, fluorescence excitation for a certain period of time can effectively promote the fluorescence quenching of DSD acid-functionalized carbon quantum dots by bipyridine compounds, enhance the detection effect, and realize quantitative detection of the concentration of bipyridine compounds; further, taking advantage of the fact that the binding ability of paraquat to DSD acid-functionalized carbon quantum dots in the excited state is much better than that in the ground state, and the difference in the binding ability of diquat to DSD acid-functionalized carbon quantum dots in the excited state and the ground state is not as significant as that of paraquat, qualitative detection of whether diquat contains paraquat can be realized.

[0094] Throughout this specification, reference to terms such as "one embodiment," "some embodiments," "illustrative embodiments," "examples," "specific examples," or "some examples" means that a specific feature, structure, material, or characteristic described in conjunction with that embodiment or example is included in at least one embodiment or example of the present invention. In this specification, illustrative uses of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.

[0095] Although the embodiments of the present application have been shown and described, those skilled in the art will appreciate that various changes, modifications, substitutions, and variations may be made to the embodiments without departing from the principles and intent of the present application, and that the scope of the present application is defined by the claims and their equivalents.

Claims

1. A fluorescence detection method for bipyridine compounds, characterized in that: include: Mixing DSD acid-functionalized carbon quantum dots, solvent A, and standard solutions containing different concentrations of bipyridine compounds to obtain multiple homogeneous solutions B, performing fluorescence excitation on each of the homogeneous solutions B, and determining the functional relationship between the concentration of the bipyridine compound and the fluorescence intensity; The DSD acid-functionalized carbon quantum dots, the solvent A, and the test sample extract containing the bipyridine compound are mixed to obtain a homogeneous solution C, the solution C is subjected to fluorescence excitation, and the concentration of the bipyridine compound contained in the test sample extract is obtained using the functional relationship between the concentration of the bipyridine compound and the fluorescence intensity; The DSD acid functionalized carbon quantum dots are prepared according to the following steps: The sulfur-nitrogen co-doped carbon quantum dots are mixed with solvent D, and an amidation reaction is carried out under the action of a catalyst and DSD acid. The product obtained by the amidation reaction is dialyzed and dried to obtain the DSD acid-functionalized carbon quantum dots.

2. The fluorescence detection method according to claim 1, wherein The standard substance of the bipyridine compound is paraquat, diquat, 4,4'-bipyridine, N-methyl-4,4'-bipyridine or 2,2'-bipyridine.

3. The fluorescence detection method according to claim 1, wherein The test article extract is selected from water samples of rice, corn, sunflower seeds, cotton seeds, blood, urine or river water.

4. The fluorescence detection method according to claim 1, wherein The mass ratio of the DSD acid-functionalized carbon quantum dots, the solvent A, and the standard solution is 1:100-10,000,000:0.05-100; and / or The mass ratio of the DSD acid-functionalized carbon quantum dots, the solvent A, and the test sample extract is 1:100-100000000:0.01-1000.

5. The fluorescence detection method according to claim 1, wherein The duration of fluorescence excitation is t∈(0s,1200s].

6. A fluorescence detection method for bipyridine compounds, characterized in that: Used to detect the presence of paraquat in diquat herbicide samples, including: Diluting the diquat herbicide sample with water according to its nominal mass concentration to obtain a diluent G, mixing the DSD acid-functionalized carbon quantum dots, solvent A, and the diluent G to obtain a homogeneous solution H, subjecting the homogeneous solution H to fluorescence excitation, measuring a time-resolved spectrum of the homogeneous solution H, and determining whether the diquat herbicide sample contains paraquat using a functional relationship between fluorescence intensity and time; The DSD acid functionalized carbon quantum dots are prepared according to the following steps: The sulfur-nitrogen co-doped carbon quantum dots are mixed with solvent D, and an amidation reaction is carried out under the action of a catalyst and DSD acid. The product obtained by the amidation reaction is dialyzed and dried to obtain the DSD acid-functionalized carbon quantum dots.

7. The fluorescence detection method according to claim 6, characterized in that: The dilution multiple is: mass concentration × 10 10 ~mass concentration × 10 8 ; The mass ratio of the DSD acid-functionalized carbon quantum dots to the solvent A is 1:100-10,000,000, and the mass ratio of the solvent A to the diluent G is 1:0.1-10; and / or The relationship between fluorescence intensity and time function was used to determine whether paraquat was present in the diquat herbicide sample. Specifically: Two points are selected from the time-resolved spectrum in chronological order, with a time difference of Δt between the two points. The fluorescence intensities of the two points are F1 and F2, respectively. When the ratio of F2 to F1 exceeds a set threshold, it is considered that the diquat herbicide sample does not contain paraquat; otherwise, it is considered that the diquat herbicide sample contains paraquat. The measurement time of the time-resolved spectrum is 10s to 1200s.

8. The fluorescence detection method according to claim 1 or 6, characterized in that: The wavelength of fluorescence excitation is 320nm~360nm, and the excitation mode is continuous excitation or intermittent excitation.

9. The fluorescence detection method according to claim 1 or 6, characterized in that: The solvent A is a mixture of any one or more of water, ethanol, acetonitrile and dimethyl sulfoxide; The solvent D is a mixture of any one or more of deionized water, ethanol and N,N-dimethylformamide; The catalyst is selected from a mixture of any one or more of N,N'-diisopropylcarbodiimide, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide, dicyclohexylcarbodiimide, 4-dimethylaminopyridine, carbodiimide hydrochloride, 1-hydroxybenzotriazole and 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate; and / or The molar ratio of the sulfur-nitrogen co-doped carbon quantum dots, the solvent D, the catalyst and the DSD acid is 1:5000-200000:1-20:1-10.

10. The fluorescence detection method according to claim 1 or 6, characterized in that: The sulfur-nitrogen co-doped carbon quantum dots are mixed with solvent D, and an amidation reaction is carried out under the action of a catalyst and DSD acid, comprising: The sulfur-nitrogen co-doped carbon quantum dots, the solvent D and the catalyst are activated for 10 to 30 minutes to obtain an activated product; The activated product is mixed with the DSD acid and subjected to an amidation reaction at room temperature for 3 to 24 hours.

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