A method for detecting nitrogen-doped sulfonated carbon dots of ceftriaxone sodium, its preparation, and its application.

Abstract

This invention provides nitrogen-doped sulfonated carbon dots for detecting ceftriaxone sodium, their preparation method, and their applications, belonging to the fields of nanomaterial synthesis and antibiotic detection technology. The method involves: dissolving sodium diphenylamine sulfonate and sodium hydroxide in deionized water under stirring and heating; applying a direct current to obtain a mixed solution; centrifuging, filtration, and dialysis of the mixed solution; and finally, freeze-drying to obtain carbon dots. The nitrogen-doped sulfonated carbon dots prepared by this method exhibit strong blue fluorescence emission, and show fluorescence quenching and a significant change in fluorescence color upon the addition of ceftriaxone sodium, making them suitable as fluorescent nanosensors for detecting ceftriaxone sodium. Furthermore, this invention, based on the ability of nitrogen-doped sulfonated carbon dots to rapidly and sensitively detect ceftriaxone sodium in real samples, provides a new method and material for applications in the fields of biology, food safety, and environmental monitoring.

Description

Technical Field

[0001] This invention belongs to the field of nanomaterial synthesis and antibiotic detection technology, and relates to a nitrogen-doped sulfonated carbon dot for detecting ceftriaxone sodium, its preparation method and application, especially a method for preparing nitrogen-doped sulfonated fluorescent carbon dots, and the application of this carbon dot as a fluorescent detection probe to detect and identify ceftriaxone sodium in aqueous solution and milk. Background Technology

[0002] Ceftriaxone sodium (CTR) is a broad-spectrum third-generation cephalosporin antibiotic widely used in medicine and animal husbandry. However, due to frequent instances of some livestock farms failing to strictly adhere to prescribed medication guidelines, abusing antibiotics, and illegally using prohibited drugs, the levels of veterinary drug residues in animal-derived foods have significantly exceeded standards, posing a threat to human health and potentially even causing death in severe cases. Milk, as an important source of animal protein, requires serious attention due to its high levels of veterinary drug residues. Therefore, a simple, accurate, and reliable method for determining the content of ceftriaxone sodium in environmental and food samples is of great significance.

[0003] Currently, analytical methods for antibiotic residue detection include high-performance liquid chromatography (HPLC), capillary electrophoresis, microbial assays with absorbance, chemiluminescence, and electrochemical methods. However, most of these methods are time-consuming, expensive, and require complex equipment, posing a significant challenge to achieving rapid and convenient determination. Fluorescence methods, with their advantages of high sensitivity, low detection limits, and cost-effectiveness, have become an effective analytical approach. Over the past decade, fluorescent probes have made significant progress in environmental analysis and have been designed to respond to specific targets, including various novel pollutants and small molecules such as picric acid, sulfites, hypochlorites, and doxycycline. However, research on the determination of ceftriaxone sodium using fluorescence spectroscopy is still limited. The challenge in ceftriaxone sodium determination lies in finding materials or fluorescent systems with specificity and high sensitivity for identification. Developing and designing fluorescent materials for analysis is crucial. As a novel fluorescent nanomaterial that has attracted much attention in recent years, carbon dots have shown significant advantages in the field of fluorescence sensing due to their unique optical properties. This material can not only be used as a high-efficiency optical sensor for material analysis, but also effectively solves the technical bottleneck of traditional detection methods in terms of ease of operation due to its good solubility, high stability and strong resistance to photobleaching during the detection process. Summary of the Invention

[0004] This invention addresses the aforementioned problems by providing a carbon dot assay for detecting ceftriaxone sodium, its preparation method, and its applications. The nitrogen-doped sulfonated carbon dots prepared by the method of this invention exhibit strong blue fluorescence emission, and upon the addition of ceftriaxone sodium, fluorescence quenching and a significant change in fluorescence color occur, enabling its use as a fluorescent nanosensor for the determination of ceftriaxone sodium. Furthermore, this invention constructs a portable sensing device based on nitrogen-doped sulfonated carbon dots that can be used with a smartphone for rapid and sensitive detection of ceftriaxone sodium in real samples, providing a new method and material for applications in the fields of biology, food safety, and environmental monitoring.

[0005] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0006] A method for preparing nitrogen-doped sulfonated carbon dots for detecting ceftriaxone sodium involves first using sodium diphenylamine sulfonate as the carbon source. In an alkaline aqueous solution environment, a direct current is applied to the solution, with both the anode and cathode being platinum sheets. Sodium diphenylamine sulfonate molecules oxidize and polymerize on the anode surface to obtain carbon dots, which are then dispersed into the solution. Finally, the carbon dots in the mixture are separated. The preparation method includes the following steps:

[0007] Step 1: At room temperature, using sodium diphenylamine sulfonate as a carbon source, add it and an alkali to a deionized aqueous solution and stir magnetically until completely dissolved. Then, preheat the solution to obtain a clear solution. The molar concentration of sodium diphenylamine sulfonate in the clear solution is 0.05-0.2 M, and the concentration of the alkali solution is 0.5-3 M.

[0008] Step 2: Apply a direct current to the clear solution obtained in Step 1 for electrolysis. Maintain a constant temperature and perform magnetic stirring during the electrolysis process. Once the electrolyte turns dark brown and shows no obvious color change, remove the current and allow it to cool naturally to room temperature to obtain a mixed solution. During electrolysis, sodium diphenylamine sulfonate molecules oxidize and polymerize on the anode surface to form carbon dots, which are then dispersed into the mixed solution.

[0009] Step 3: The mixture obtained in Step 2 is centrifuged, filtered, and dialyzed to separate the carbon dots. Then, it is freeze-dried to obtain nitrogen-doped sulfonated carbon dots in the form of a solid sample.

[0010] Furthermore, in step 1, the type of alkali is a strong alkali, including one or more of sodium hydroxide, potassium hydroxide, lithium hydroxide, cesium hydroxide, calcium hydroxide, barium hydroxide, and ammonia, preferably sodium hydroxide and potassium hydroxide.

[0011] Furthermore, in step 1: the preheating temperature is 60-80℃, and the time is 10-30 minutes.

[0012] Furthermore, in step 2: the system for applying direct current consists of two platinum electrode plates and an electrolytic cell; the electrolyte volume occupies 1 / 3-2 / 3 of the electrolytic cell volume; the direct current intensity is 0.5-3A; and the electrochemical reaction time is 1-4 hours. The electrolyte is the clear solution from step 1.

[0013] Furthermore, in step 3: the mixture is centrifuged at 8000-10000 r / min for 15-30 min, filtered with PTFE filter paper with a pore size of 0.22 μm to obtain filtrate, and dialyzed to neutral using a 500-1000 Da dialysis bag.

[0014] Furthermore, in step 3: the freeze-drying temperature is -40 to -80°C, and the time is 36 to 48 hours.

[0015] Further, in step 3, the pre-fitted linear equation is obtained by the following method: A phosphate buffer solution containing carbon dots is sequentially and quantitatively injected into centrifuge tubes. Then, equal volumes of ceftriaxone sodium samples with concentrations of 0, 4, 6, 10, 20, 30, 40, 50, and 60 μmol / L are precisely added to the system. The reaction system is uniformly mixed by vortexing for 3-10 seconds. After incubation at room temperature in the dark for 30-60 minutes, the mixture is transferred to a standard quartz cuvette for image capture using a portable detection device. The colors of the image are converted into corresponding R, G, B, H, S, and V values. A linear equation is fitted between the B×H×100 / G×S×V value corresponding to the nitrogen-doped sulfonated carbon dots and c, where c is the concentration of ceftriaxone sodium in the sample, and B, H, G, S, and V values ​​represent the corresponding blue, hue, green, saturation, and brightness values ​​of the image, respectively.

[0016] The present invention also provides a nitrogen-doped sulfonated carbon dot for detecting ceftriaxone sodium, which is prepared by the above preparation method. The nitrogen-doped sulfonated carbon dot has a nitrogen-doped carbon core structure, and its surface is modified with amino and sulfonate groups. It has the characteristics of strong water dispersibility and exhibits excellent fluorescence performance.

[0017] An application for detecting nitrogen-doped sulfonated carbon dots in ceftriaxone sodium, wherein the nitrogen-doped sulfonated carbon dots are used in the detection of ceftriaxone sodium, and the detection method includes the following steps:

[0018] Step 1: Add a phosphate buffer solution containing nitrogen-doped sulfonated carbon dots to the sample to be tested to obtain a mixed solution, and incubate for a period of time. The sample to be tested may include an aqueous solution or a milk sample. The incubation time should be greater than 30 minutes.

[0019] Step 2: Transfer the mixed solution from Step 1, after incubation for a period of time, to a standard quartz cuvette for fluorescence emission spectroscopy or fluorescence imaging.

[0020] Step 3: Convert the fluorescence emission intensity or color of the captured image measured in Step 2 into the corresponding numerical value, and substitute it into the pre-fitted linear equation to obtain the content of ceftriaxone sodium in the sample to be tested.

[0021] Further, in step 1: the concentration of the phosphate buffer solution is 0.05-0.5M, and the pH value is 7-9; the concentration of nitrogen-doped sulfonated carbon dots in the phosphate buffer solution is 0.01-0.1 mg / mL. The milk sample is prepared by mixing milk and acetonitrile in a 1:1 ratio, sonicating for 10-30 min, centrifuging at 12000-15000 rpm for 10 min, and then filtering through a 0.22 μm microporous membrane to obtain a clear and transparent pretreated milk solution; the nitrogen-doped sulfonated carbon dopant phosphate buffer solution is mixed with the milk sample or aqueous solution at a 1:1 volume ratio.

[0022] The principle and innovation of this invention are as follows:

[0023] In a strongly alkaline environment, water electrolysis produces hydroxyl radicals (·OH) and superoxide anions (·O). 2- The reaction disrupts the -NH- group of sodium diphenylamine sulfonate. The sulfonate group in sodium diphenylamine sulfonate increases conductivity and promotes the electrochemical oxidation process. In the anodic region, aromatic fragments from sodium diphenylamine sulfonate after bond breakage undergo condensation and carbonization with nitrogen-containing organic small molecules to form nitrogen-doped carbon cores. The -NH- group is converted to pyrrole nitrogen and pyridine nitrogen. The surface is modified by the sulfonate group of sodium diphenylamine sulfonate and the amino groups generated during the reaction, thus exhibiting excellent fluorescence properties. This method of electrochemical oxidation of small molecules differs from the previously reported top-down electrochemical stripping method and is characterized by easy synthesis and controllability. Simultaneously, the pyrrole nitrogen in the nitrogen-doped sulfonated carbon dots can selectively recognize ceftriaxone sodium in the test sample, and detection is achieved through changes in fluorescence intensity or color. This method is simple, portable, and allows for rapid real-time detection of ceftriaxone sodium.

[0024] Compared with the prior art, the beneficial effects of the present invention are:

[0025] (1) The carbon dots prepared by this invention have a unique composition and surface functional groups, with nitrogen doping in the core and sulfonate and amino groups on the surface;

[0026] (2) The preparation method of the present invention is simple, reliable, green and environmentally friendly, low in cost, and suitable for large-scale production;

[0027] (3) The carbon dots provided by the present invention have fluorescence that can be efficiently quenched by ceftriaxone sodium, and have high sensitivity and excellent anti-interference performance.

[0028] (4) The fluorescent detection probe and portable detection device provided by the present invention have high sensitivity and good selectivity.

[0029] (5) The present invention is simple to operate and does not require large instruments. It can perform rapid on-site detection through smartphone software, which can better overcome the problems of complex food matrices such as milk and the susceptibility of detection to interference. Attached Figure Description

[0030] Figure 1 (a) is a transmission electron microscope image of nitrogen-doped sulfonated carbon dots prepared in Example 1 of the present invention. Figure 1 (b) is a high-magnification transmission electron microscope image and lattice fringe pattern of the nitrogen-doped sulfonated carbon dots prepared in Example 1 of the present invention. Figure 1 (c) is Figure 1 (a) The corresponding particle size distribution diagram.

[0031] Figure 2 The infrared spectrum of the nitrogen-doped sulfonated carbon dots prepared in Example 1 of this invention is shown.

[0032] Figure 3 The UV-Vis absorption spectrum and fluorescence excitation-emission spectrum of the nitrogen-doped sulfonated carbon dots prepared in Example 1 of this invention are shown.

[0033] Figure 4 This is a spectrum showing the fluorescence emission curve of nitrogen-doped sulfonated carbon dots prepared in Example 1 of the present invention as a function of excitation wavelength.

[0034] Figure 5 This is a selectivity diagram for the detection of ceftriaxone sodium by nitrogen-doped sulfonated carbon dots. Figure 5 (a) Changes in fluorescence intensity at an excitation wavelength of 423 nm after adding different metal ions (300 μM) to nitrogen-doped sulfonated carbon dot solution; Figure 5 (b) The change in fluorescence intensity of nitrogen-doped sulfonated carbon dot solution at an excitation wavelength of 423 nm after the addition of different small molecules (300 μM);

[0035] Figure 6 A schematic diagram illustrating the detection of ceftriaxone sodium concentration based on fluorescence intensity changes using nitrogen-doped sulfonated carbon dots; where... Figure 6 (a) is a graph showing the fluorescence emission spectrum changes of nitrogen-doped sulfonated carbon dots at concentrations of ceftriaxone sodium ranging from 0 to 300 μM; Figure 6 (b) is a graph showing the fluorescence emission spectrum changes of nitrogen-doped sulfonated carbon dots at concentrations of 0-24 μM for ceftriaxone sodium; Figure 6 (c) The fluorescence intensity ratio of nitrogen-doped sulfonated carbon dots before and after the addition of ceftriaxone sodium is a piecewise linear relationship with the concentration of ceftriaxone sodium in the range of 0-300 μM; Figure 6(d) shows the linear relationship between the fluorescence intensity ratio of nitrogen-doped sulfonated carbon dots before and after the addition of ceftriaxone sodium and the concentration of ceftriaxone sodium in the range of 0-24 μM.

[0036] Figure 7 A schematic diagram illustrating the detection of ceftriaxone sodium concentration based on color changes of nitrogen-doped sulfonated carbon dots; where... Figure 7 (a) The fluorescence color change of nitrogen-doped sulfonated carbon dots at concentrations of 0-30 μM in ceftriaxone sodium; Figure 7 (b) is a graph showing the relationship between the B×H×100 / G×S×V value and the concentration of added ceftriaxone sodium. Detailed Implementation

[0037] The present invention will be further described below with reference to specific embodiments, but the present invention is not limited to the following embodiments.

[0038] Example 1:

[0039] Step 1: Dissolve 2.768g sodium diphenylamine sulfonate and 4.000g sodium hydroxide in 100mL deionized water with magnetic stirring, and preheat from room temperature to 70℃ for 10min.

[0040] Step 2: Apply a 1A DC current to the clear solution obtained in Step 1 for electrolysis. Maintain a constant temperature of 70℃ and magnetic stirring conditions during the process. The electrolysis time is 2 hours. After electrolysis is completed, allow it to cool naturally to room temperature.

[0041] Step 3: The mixture obtained in Step 2 was centrifuged at 9000 r / min for 20 min, filtered through a 0.22 μm microporous membrane, dialyzed to neutral using a 1000 Da dialysis bag, and then freeze-dried to obtain carbon dot solid powder. The freeze-drying temperature was -40℃ and the time was 48 h. The absolute quantum efficiency of the 0.01 mg / mL carbon dot solution was measured to be 23.34% using a UV-NIR absolute quantum yield analyzer.

[0042] Example 2:

[0043] Step 1: Dissolve 1.384g sodium diphenylamine sulfonate and 5.611g potassium hydroxide in 100mL deionized water with magnetic stirring, and preheat from room temperature to 60℃ for 20min.

[0044] Step 2: Apply a 0.5A DC current to the clear solution obtained in Step 1 for electrolysis. Maintain a constant temperature of 60℃ and magnetic stirring conditions during the process. The electrolysis time is 1 hour. After the electrolysis is completed, allow it to cool naturally to room temperature.

[0045] Step 3: The mixture obtained in Step 2 was centrifuged at 8000 r / min for 15 min, filtered through a 0.22 μm microporous membrane, dialyzed to neutral using a 500 Da dialysis bag, and then freeze-dried to obtain carbon dot solid powder. The freeze-drying temperature was -80℃ and the time was 36 h. The absolute quantum efficiency of the 0.01 mg / mL carbon dot solution was measured to be 11.53% using a UV-NIR absolute quantum yield analyzer.

[0046] Example 3:

[0047] Step 1: Dissolve 5.536g sodium diphenylamine sulfonate and 8.000g sodium hydroxide in 100mL deionized water with magnetic stirring, and preheat from room temperature to 80℃ for 30min.

[0048] Step 2: Apply a 2A DC current to the clear solution obtained in Step 1 for electrolysis. Maintain a constant temperature of 80℃ and magnetic stirring conditions during the process. The electrolysis time is 3 hours. After the electrolysis is completed, allow it to cool naturally to room temperature.

[0049] Step 3: The mixture obtained in Step 2 was centrifuged at 8000 r / min for 25 min, filtered through a 0.22 μm microporous membrane, dialyzed to neutral using a 1000 Da dialysis bag, and then freeze-dried to obtain carbon dot solid powder. The freeze-drying temperature was -60℃ and the time was 40 h. The absolute quantum efficiency of the 0.01 mg / mL carbon dot solution was measured to be 13.33% using a UV-NIR absolute quantum yield analyzer.

[0050] Example 4:

[0051] Step 1: Dissolve 8.304g sodium diphenylamine sulfonate and 12.001g sodium hydroxide in 100mL deionized water with magnetic stirring, and preheat from room temperature to 75℃ for 15min.

[0052] Step 2: Apply a 3A DC current to the clear solution obtained in Step 1 for electrolysis. Maintain a constant temperature of 75℃ and magnetic stirring conditions during the process. The electrolysis time is 4 hours. After the electrolysis is completed, allow it to cool naturally to room temperature.

[0053] Step 3: The mixture obtained in Step 2 was centrifuged at 10000 r / min for 30 min, filtered through a 0.22 μm microporous membrane, dialyzed to neutral using a 500 Da dialysis bag, and then freeze-dried to obtain carbon dot solid powder. The freeze-drying temperature was -70℃ and the time was 38 h. The absolute quantum efficiency of the 0.01 mg / mL carbon dot solution was measured to be 14.66% using an ultraviolet-near-infrared absolute quantum yield meter.

[0054] Comparative Example 1 (compared to Example 1):

[0055] Step 1: Dissolve 1.692g of diphenylamine and 4.000g of sodium hydroxide in 100mL of deionized water with magnetic stirring, and preheat from room temperature to 70℃ for 15min.

[0056] Step 2: Apply a 1A DC current to the clear solution obtained in Step 1 for electrolysis. Maintain a constant temperature of 70℃ and magnetic stirring conditions during the process. The electrolysis time is 2 hours. After electrolysis is completed, allow it to cool naturally to room temperature.

[0057] Step 3: The mixture obtained in Step 2 was centrifuged at 9000 r / min for 20 min, filtered through a 0.22 μm microporous membrane, dialyzed to neutral using a 500 Da dialysis bag, and then freeze-dried to obtain carbon dot solid powder. The freeze-drying temperature was -40℃ and the time was 48 h. The absolute quantum yield of the 0.01 mg / mL carbon dot solution was measured to be 2.31% using a UV-NIR absolute quantum yield analyzer.

[0058] Comparative Example 1 is compared with Example 1, the difference being that the carbon source was changed, with sodium diphenylamine sulfonate replaced by diphenylamine. The results show the importance of the sulfonate group in sodium diphenylamine sulfonate for promoting the electrolysis reaction process.

[0059] Comparative Example 2 (compared to Example 1):

[0060] Step 1: Dissolve 2.768g of sodium diphenylamine sulfonate magnetometer in 100mL of 1M hydrochloric acid solution by stirring, and preheat from room temperature to 70℃ for 15min.

[0061] Step 2: Apply a 1A DC current to the clear solution obtained in Step 1 for electrolysis. Maintain a constant temperature of 70℃ and magnetic stirring conditions during the process. The electrolysis time is 2 hours. After electrolysis is completed, allow it to cool naturally to room temperature.

[0062] Step 3: The mixture obtained in Step 2 was centrifuged at 9000 r / min for 20 min, filtered through a 0.22 μm microporous membrane, dialyzed to neutral using a 1000 Da dialysis bag, and then freeze-dried to obtain carbon dot solid powder. The freeze-drying temperature was -40℃ and the time was 48 h. The absolute quantum efficiency of the 0.01 mg / mL carbon dot solution was measured to be 1.86% using a UV-NIR absolute quantum yield analyzer.

[0063] This Comparative Example 1 is compared with Example 1. The difference is that the electrolyte environment was changed, and sodium hydroxide was replaced with hydrochloric acid. The results show that an alkaline environment is one of the important conditions for the generation of carbon dots by electrolysis.

[0064] The morphology characterization of the nitrogen-doped sulfonated carbon dots prepared in Example 1 of this invention is as follows: Figure 1 As shown. Figure 1 A transmission electron microscope image shows that the carbon dots are in a monodisperse state. Figure 1 b is a high-resolution transmission electron microscope image; a lattice spacing of 0.343 nm corresponds to graphite (sp... 2 The diffraction plane of carbon. Figure 1 Statistical analysis of the carbon dots clearly shows that their particle size distribution is between 6 and 10 nm, with an average particle size of approximately 8.2 nm.

[0065] The structural characterization of the nitrogen-doped sulfonated carbon dots prepared in Example 1 of this invention is as follows: Figure 2 As shown in the figure. Infrared spectroscopy reveals a significant difference in peak shape between the nitrogen-doped sulfonated carbon dots and the precursor sodium diphenylamine sulfonate, confirming the successful occurrence of the reaction. At 1034 cm⁻¹ -1 and 1205cm -1 The asymmetric stretching vibration of the O=S=O group at the position, and 631 cm⁻¹ -1 The stretching vibration of the CS bond at the location indicates the presence of sulfonic acid groups retained by sodium diphenylamine sulfonate in the nitrogen-doped sulfonated carbon dots. Characteristic vibrations involving nitrogen atoms occur in the 1650-1300 cm⁻¹ range. -1 The area holds special significance. 1633cm -1 1590cm -1 and 1490cm -1 The strong absorption peaks appearing at these locations correspond to the stretching vibration coupling modes of the C=C bonds in the pyrrole ring skeleton and the C+C and CN bonds in the pyridine ring skeleton, respectively, confirming that the nitrogen-doped sulfonated carbon dots have a different nitrogen presence form than that in sodium diphenylamine sulfonate and possess an aromatic conjugated structure. Furthermore, the 3500-3300 cm⁻¹... -1 and 3600-3200cm -1 The broad and strong absorption bands within the range are characteristic of electronegative -OH, -COOH, and -NH2 groups, supporting the conclusion that nitrogen and oxygen atoms have been doped onto the surface of nitrogen-doped sulfonated carbon dots. Comparison with the original material shows that the nitrogen-doped sulfonated carbon dots retain the sulfonic acid functional groups of sodium diphenylamine sulfonate, while also exhibiting the characteristics of novel nitrogen-containing groups.

[0066] The optical property spectrum of the nitrogen-doped sulfonated carbon dots prepared in Example 1 of this invention is shown below. Figure 3 , 4 As shown. Figure 3 The optical properties of nitrogen-doped sulfonated carbon dots were analyzed using ultraviolet-visible absorption spectroscopy. The results showed two closely spaced absorption bands. The first absorption peak appeared at 270 nm, attributed to the nucleus sp state. 2The second absorption band, observed at 326 nm, is associated with the n→π* transition of the C=O bond and the surface functional groups of the nitrogen-doped sulfonated carbon dots. This can be attributed to the surface functional groups and the doping of nitrogen and sulfur atoms. Furthermore, the optimal excitation wavelength for the nitrogen-doped sulfonated carbon dots is 310 nm, corresponding to the emission peak at 423 nm. Figure 4 The images show the emission spectra of the carbon dot at different excitation wavelengths. When using excitation wavelengths of 270-340 nm, we observed a central emission wavelength of 423 nm. Notably, despite the change in excitation wavelength, the maximum fluorescence emission peak of the nitrogen-doped sulfonated carbon dot remained consistent. This consistency indicates that the carbon dot exhibits excitation wavelength independence and that the nitrogen-doped sulfonated carbon dot has a uniform size, resulting in a relatively stable fluorescence emission peak.

[0067] Application verification:

[0068] To investigate the selectivity of the prepared nitrogen-doped sulfonated carbon dots for ceftriaxone sodium, using the nitrogen-doped sulfonated carbon dots prepared in Example 1 as an example, solutions of BaCl2, CaCl2, CdCl2, AgNO3, CoCl2, CuCl2, FeCl2, FeCl3, MgCl2, MnCl2, NiCl2, and ZnCl2 with a concentration of 0.1 mol / L were prepared and diluted to 1 mmol / L. 1 mL of each salt solution was placed in a centrifuge tube, and then 1 mL of a phosphate buffer solution containing 0.02 mg of nitrogen-doped sulfonated carbon dots was added. The mixture was vortexed for 3 seconds. The reaction system was then homogeneously mixed. Solutions of valine, threonine, proline, glutamic acid, alanine, riboflavin, oxytetracycline, tetracycline, penicillin, ampicillin, and amoxicillin with a concentration of 0.01 mol / L were prepared and diluted to 500 μmol / L. 1 mL of each of these small molecule solutions was placed in a centrifuge tube, and 1 mL of a phosphate buffer solution containing 0.02 mg of nitrogen-doped sulfonated carbon dots was added. The mixture was vortexed for 3 seconds to achieve homogeneous mixing. The fluorescence of the nitrogen-doped sulfonated carbon dots was then tested using a fluorescence spectrometer under 310 nm incident light to determine whether the fluorescence was quenched upon the addition of these substances. Figure 5 This indicates that nitrogen-doped sulfonated carbon dots have good selectivity for the detection of ceftriaxone sodium. Figure 5 (a) indicates that most metal ions do not have a fluorescence quenching effect on nitrogen-doped sulfonated carbon dots; Figure 5 (b) indicates that some common small molecules have a negligible effect on the fluorescence quenching of nitrogen-doped sulfonated carbon dots.

[0069] The nitrogen-doped sulfonated carbon dots obtained in Examples 1, 2, 3, 4, Comparative Example 1, and Comparative Example 2 were used as fluorescent probes for the detection of ceftriaxone sodium. The specific method was as follows: 1 mL of phosphate buffer solution containing 0.02 mg of nitrogen-doped sulfonated carbon dots was quantitatively injected into a 4 mL centrifuge tube. Then, 1 mL of ceftriaxone sodium samples with concentrations of 4, 8, 16, 20, 24, 28, 32, 36, 40, 44, and 48 μmol / L were accurately transferred into the system. The reaction system was uniformly mixed by vortexing for 3 seconds. After incubation at room temperature in the dark for 30 minutes, the mixture was transferred to a standard quartz cuvette for fluorescence emission spectroscopy measurement. The changes in fluorescence intensity were recorded, and the F0 / F and c(CTR) equations for each example were fitted, where c(CTR) is the concentration of ceftriaxone sodium in the test solution, F0 is the fluorescence intensity value at 423 nm of the nitrogen-doped sulfonated carbon dot phosphate buffer solution, and F is the fluorescence intensity value at 423 nm of the mixed solution after a 1:1 mixture of the nitrogen-doped sulfonated carbon dot phosphate buffer solution and the ceftriaxone sodium sample. Comparative Examples 1 and 2 showed no detection effect on ceftriaxone sodium, and the detection limits for Examples 1, 2, 3, and 4 were 0.328 μM, 0.767 μM, 0.723 μM, and 0.785 μM, respectively. For Example 1, in Figure 6 In (a), we observed that the fluorescence of nitrogen-doped sulfonated carbon dots quenched with increasing ceftriaxone sodium concentration when the concentration ranged from 0 to 300 μM. Figure 6 In (b), we can perform piecewise linear fitting on this fluorescence intensity change. Figure 6 In (c), it can be seen that as the concentration of ceftriaxone sodium increases from 2 μM to 24 μM, the blue fluorescence emission intensity at 423 nm gradually decreases, indicating that fluorescence quenching can be sensitively triggered even in the presence of low concentrations of ceftriaxone sodium. This results in a stepwise change in the fluorescence intensity ratio F0 / F, ranging from 1 to 1.3. Figure 6 In (d), it can be seen that the fluorescence intensity ratio F0 / F exhibits a strong linear correlation with the CTR concentration in the range of 0-30 μM (R0). 2 =0.9957), the fitted equation was F0 / F = 0.0107 × c(CTR) + 0.9994, and the limit of detection (LOD) was calculated to be 0.298 μM according to the formula LOD = 3σ / k. Using this method to detect ceftriaxone sodium spiked aqueous solution, the spiked recovery rate was found to be 98.7% (n = 3).

[0070] The nitrogen-doped sulfonated carbon dots prepared in Example 1 were used to detect ceftriaxone sodium by observing color changes. Quantitative amounts of phosphate buffer solution were sequentially injected into centrifuge tubes containing the carbon dots. Then, ceftriaxone sodium samples with concentrations of 0, 10, 20, 30, 40, 50, and 60 μmol / L were precisely added to the system. The reaction system was homogeneously mixed by vortexing for 10 seconds. After incubation at room temperature in the dark for 30 minutes, the mixture was transferred to a standard quartz cuvette and imaged using a portable detection device. The colors of the images were converted into corresponding R, G, B, H, S, and V values. A linear equation was fitted between B×H×100 / G×S×V and c(CTR) for the nitrogen-doped sulfonated carbon dots, where c(CTR) is the concentration of ceftriaxone sodium in the sample, and the B, H, G, S, and V values ​​represent the corresponding blue, hue, green, saturation, and brightness values ​​of the image, respectively. The content of ceftriaxone sodium in the sample can be obtained by substituting the B×H×100 / G×S×V value corresponding to the photograph of the unknown sample into the equation. Figure 7 (a) is a series of fluorescence images of milk samples containing 0-30 μM ceftriaxone sodium. Figure 7 (b) The concentration of ceftriaxone sodium was assessed by calculating the B×H×100 / G×S×V value. The B×H×100 / G×S×V value varied with the concentration of ceftriaxone sodium and showed a strong linear relationship in the range of 0-30 μM (R0). 2 =0.9805), the fitted linear equation was B×H×100 / G×S×V=0.1748c(CTR)+9.4379, and the calculated limit of detection (LOD) was 0.38 μM. This method was used to detect ceftriaxone sodium in milk samples, and the recovery rate was found to be 103.6% (n=3).

[0071] Finally, it should be noted that the above embodiments are merely illustrative of the implementation methods of the present invention, but should not be construed as limiting the scope of the present invention. It should be pointed out that those skilled in the art can make several modifications and improvements without departing from the concept of the present invention, and these all fall within the protection scope of the present invention.

Claims

1. A method for the preparation of nitrogen-doped sulfonated carbon dots for the detection of ceftriaxone sodium, characterized by, The preparation method comprises the following steps: Step 1: Sodium diphenylamine sulfonate is used as a carbon source, and the sodium diphenylamine sulfonate and alkali are added into a deionized water solution and magnetically stirred until completely dissolved, and preheating treatment is performed to obtain a clear solution; the concentration of sodium diphenylamine sulfonate in the clear solution is 0.05-0.2 M, and the concentration of the alkali solution is 0.5-3 M; Step 2: Direct current is applied to the clear solution obtained in step 1 to perform electrolysis, and the electrolysis process is kept at constant temperature and magnetically stirred; during the electrolysis process, the sodium diphenylamine sulfonate molecules are oxidized and polymerized on the surface of the anode to obtain carbon dots, which are then dispersed into the mixed solution; the system for applying the direct current is composed of two platinum electrodes and an electrolytic cell, the volume of the electrolyte accounts for 1 / 3-2 / 3 of the volume of the electrolytic cell, the direct current intensity is 0.5-3 A, and the electrochemical reaction time is 1-4 h; Step 3: The mixed solution obtained in step 2 is subjected to centrifugation, suction filtration and dialysis to separate the carbon dots, and then freeze-drying is performed to obtain nitrogen-doped sulfonated carbon dots.

2. A method for the preparation of nitrogen-doped sulfurbated carbon dots for the detection of ceftriaxone sodium as claimed in claim 1, wherein, In step 1, the type of the alkali is a strong alkali selected from one or more of sodium hydroxide, potassium hydroxide, lithium hydroxide, cesium hydroxide, calcium hydroxide, barium hydroxide and ammonia water; the preheating treatment temperature is 60-80 ℃, and the time is 10-30 min.

3. A method for the preparation of nitrogen-doped sulfurbated carbon dots for the detection of ceftriaxone sodium as claimed in claim 1, wherein, In step 1, the type of the alkali is sodium hydroxide or potassium hydroxide.

4. A method for the preparation of nitrogen-doped sulfurbated carbon dots for the detection of ceftriaxone sodium as claimed in claim 1, wherein, In step 2, the direct current is removed after the electrolyte turns black brown and there is no obvious color change, and the mixed solution is naturally cooled to room temperature.

5. The method for preparing nitrogen-doped sulfurbated carbon dots for detecting ceftriaxone sodium according to claim 1, characterized in that, In step 3, the mixed solution is centrifuged at a speed of 8000-10000 r / min for 15-30 min, the filtrate is obtained by suction filtration using a PTFE filter paper with a pore size of 0.22 μm, and the filtrate is dialyzed to neutral using a dialysis bag with a molecular weight cut-off of 500-1000 Da; the freeze-drying temperature is -40~-80 ℃, and the time is 36~48 h.

6. A nitrogen-doped sulfonated carbon dot for detecting ceftriaxone sodium, characterized in that, The nitrogen-doped sulfonated carbon dots are prepared by the preparation method of any one of claims 1-5; the nitrogen-doped sulfonated carbon dots have a nitrogen-doped carbon core structure, are modified with amino and sulfonate groups on the surface, have the characteristic of strong water dispersibility, and exhibit fluorescence performance.

7. Use of the nitrogen-doped sulfurb carbon dots for detecting ceftriaxone sodium as claimed in claim 6, wherein, The nitrogen-doped sulfonated carbon dots are applied to the detection of ceftriaxone sodium.

8. Use of nitrogen-doped sulfurb carbon dots for detecting ceftriaxone sodium according to claim 7, characterized in that, The detection method comprises the following steps: Step 1: A phosphate buffer solution of the nitrogen-doped sulfonated carbon dots is added to a sample to be detected to obtain a mixed solution, and incubation is performed, wherein the incubation time is greater than 30 min; Step 2: The mixed solution of step 1 is transferred to a standard quartz cuvette for fluorescence emission spectrum determination or fluorescence photograph shooting; Step 3: The fluorescence emission intensity or the color of the photographed image determined in step 2 is converted into a corresponding numerical value, which is substituted into a linear equation fitted in advance to obtain the content of ceftriaxone sodium in the sample to be detected.

9. Use of nitrogen-doped sulfurb carbon dots for detecting ceftriaxone sodium according to claim 8, characterized in that, In the detection method: In step 1, the sample to be detected includes an aqueous solution or a milk sample; wherein the milk sample is prepared by mixing milk and acetonitrile at a ratio of 1:1, ultrasonic treatment, centrifugation and filtration; In step 1, the concentration of the phosphate buffer solution is 0.05-0.5 M, and the pH value is 7-9; The concentration of the nitrogen-doped sulfonated carbon dots in the phosphate buffer solution in the step 1 is 0.01-0.1 mg / mL; In the step 1, the nitrogen-doped sulfonated carbon dots in the phosphate buffer solution are mixed with the sample to be detected at a volume ratio of 1:1; In the step 3, the linear equation fitted in advance is obtained by the following method: the centrifugal tube is sequentially quantitatively injected with the phosphate buffer solution of the carbon dots, and then the same volume of ceftriaxone sodium samples with concentrations of 0, 4, 6, 10, 20, 30, 40, 50 and 60 μmol / L is accurately removed and added to the system; the reaction system is uniformly mixed by vortex oscillation, and after incubation at room temperature in the dark, it is transferred to a standard quartz colorimetric cell for image shooting; the color of the image is converted into corresponding R, G, B, H, S and V values, and the linear equation of the BxHx100 / GxSxV value of the nitrogen-doped sulfonated carbon dots corresponding to c is fitted, wherein c is the concentration of ceftriaxone sodium in the sample to be detected, and B, H, G, S and V values represent the color of the image converted into corresponding blue value, hue value, green value, saturation and brightness, respectively.

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