Preparation of ratio fluorescent probe based on L / J-CDs and application of ratio fluorescent probe to cetirizine hydrochloride detection

Abstract

The invention discloses preparation of an L / J-CDs-based ratiometric fluorescent probe and application of the ratiometric fluorescent probe to cetirizine hydrochloride detection, and belongs to the technical field of chemical synthesis and analysis. The ratiometric fluorescent probe is composed of a composite system of a carbon dot L-CDs solution emitting blue fluorescence and a carbon dot J-CDs solution emitting orange fluorescence, the blue fluorescence of the L-CDs in the composite system is quenched due to the addition of cetirizine hydrochloride molecules, the J-CDs are used as a reference signal, the fluorescence color is not changed, and the ratio of the fluorescence to the carbon dot L-CDs is determined. Cross reference correction is carried out through the intensity ratio of two emission peaks under the same excitation wavelength, the effects of reducing errors and improving accuracy are achieved, and visual qualitative detection assisted by naked eyes and smart phones based on an ultraviolet lamp can be achieved through the ratio fluorescent probe.

Description

Technical Field

[0001] The invention belongs to the technical field of chemical synthesis and analysis, and specifically relates to a ratio fluorescent probe based on L / J-CDs and a preparation method thereof, and uses the fluorescent probe as a detection reagent for visual detection and quantitative detection of cetirizine hydrochloride. Background Art

[0002] Cetirizine hydrochloride (CTZ) is a nonsteroidal anti-inflammatory drug that relieves allergic reactions by interacting with H1 receptors and inhibiting the release of histamine molecules. It can also be used to treat chronic urticaria. However, excessive cetirizine hydrochloride can cause adverse symptoms such as drowsiness, headache, and cognitive retardation in the human body. In addition, cetirizine hydrochloride, which repeatedly enters the environment through different water cycle pathways, is also a persistent organic pollutant that induces aquatic biological reactions and leads to aquatic ecological imbalance, including damage at the cellular level, changes in enzyme activity during oxidative stress, and enhanced immunotoxicity.

[0003] At present, instrumental technologies including liquid chromatography-tandem mass spectrometry, high performance liquid chromatography, capillary electrophoresis and electrochemical sensors have been used for the detection of cetirizine hydrochloride. However, these methods have shortcomings such as long pretreatment time, high technical skills, equipment operation and maintenance by dedicated personnel, and high testing costs during the analysis process, which limits their application in rapid trace detection. Therefore, fluorescent probes with advantages such as cost-effectiveness, rapid response, simplicity and portability, and the ability to perform in situ monitoring in various environments have become candidates for achieving real-time sample analysis.

[0004] Carbon dots (CDs) are unique zero-dimensional luminescent nanomaterials with an average diameter of less than 10 nm. They have outstanding biocompatibility, optoelectronic properties, irradiation stability, wide sources and low toxicity, and have been widely used in many fields. Heteroatom doping is an effective method to regulate the intrinsic properties of CDs. Nitrogen doping can improve the photoluminescence properties of CDs, such as increasing the quantum yield and passivating the surface to improve the self-quenching effect.

[0005] Although there have been a few published papers reporting the application of single-emission CDs fluorescent nanomaterials as probes for the analytical detection of cetirizine hydrochloride, these few papers are based on measuring the changes in a single fluorescence signal, which is easily affected by environmental and instrumental factors. Therefore, it is necessary to develop a rapid and sensitive analytical method for the detection of trace amounts of cetirizine hydrochloride in environmental water. Summary of the invention

[0006] In view of the above-mentioned deficiencies in the prior art, the present invention provides a preparation method of a ratio fluorescent probe based on L / J-CDs and its application in the detection of cetirizine hydrochloride.

[0007] The fast visual ratio fluorescence probe as the core of the probe system of the present invention realizes the selective quantitative detection of cetirizine hydrochloride by integrating the designed L-CDs and J-CDs, has good reproducibility and wide linear range, and can realize visual qualitative detection based on the naked eye under ultraviolet light and assisted by smart phones. The addition of cetirizine hydrochloride causes the blue emission fluorescence quenching of L-CDs at 445nm, while the orange fluorescence emission of J-CDs at 578nm, as a reference signal, has almost no change, so that the probe detection system produces a significant ratio color change from lavender to orange, and the intensity ratio of the two emission peaks under the same excitation wavelength is cross-referenced and corrected to achieve the effect of reducing errors and improving accuracy. The ratio fluorescence probe detection system provides a new method for constructing a visualization and quantitative system for the detection of trace drug residues.

[0008] The purpose of the present invention is achieved through the following technical solutions:

[0009] In a first aspect, the present invention provides a method for preparing a ratiometric fluorescent probe based on L / J-CDs, the method comprising the following steps:

[0010] (1) Synthesis of L-CDs: 1.50 g sodium citrate and 0.78 g acrylamide nitrogen dopant were dispersed in 30 mL deionized water and ultrasonicated for 10 min until completely dissolved. The resulting mixture was transferred to a 50 mL Teflon reactor and then reacted at 200 °C for 3 h. After naturally cooling to room temperature, the product was filtered through a filter membrane to remove larger particles. Finally, the obtained solution was diluted 10 to 1000 times with deionized water and stored at 4 °C. The resulting solution was the nitrogen-doped L-CDs solution.

[0011] (2) Synthesis of J-CDs: 0.75 g of rhodamine B was ultrasonically dispersed in 30 mL of deionized water. The resulting solution was transferred to a 50 mL Teflon reactor and treated at 160°C for 2 h. After naturally cooling to room temperature, the resulting product was centrifuged at 10,000 rpm for 10 min. After purification by dialysis and freeze-drying, the resulting powder was dispersed in 20 L of deionized water and stored at 4°C. The resulting solution was the J-CDs solution.

[0012] (3) Preparation of L / J-CDs ratio fluorescent probe: The L-CDs solution obtained in step (1) and the J-CDs solution obtained in step (2) are fully mixed at a volume ratio of 2:3 to 5:3 to obtain a L / J-CDs ratio fluorescent probe.

[0013] In step (1), the filter membrane is a 0.22 μm water-based microporous filter membrane.

[0014] Preferably, the final solution is diluted 100 times with deionized water.

[0015] In step (2), the dialysis purification is performed by filtering through a 0.22 μm water-based microporous filter membrane and dialysis purification using a dialysis bag with a molecular weight cutoff of 1000 Da.

[0016] Preferably, in step (3), the volume ratio of the L-CDs solution to the J-CDs solution is 1:1.

[0017] In a second aspect, the present invention provides a ratio fluorescent probe based on L / J-CDs, wherein the ratio fluorescent probe is prepared by the above method, and the ratio fluorescent probe is composed of a composite system of L-CDs emitting blue fluorescence and J-CDs emitting orange fluorescence.

[0018] In a third aspect, the present invention provides an application of a ratiometric fluorescent probe based on L / J-CDs in the preparation of a detection reagent, wherein the detection reagent is used for visual detection or quantitative detection of cetirizine hydrochloride.

[0019] The core of the present invention for visual detection and quantitative detection of cetirizine hydrochloride residues is the designed ratio fluorescence probe detection system of L / J-CDs. The introduction of cetirizine hydrochloride into the ratio fluorescence detection system of L / J-CDs leads to quenching of the blue emission fluorescence of L-CDs at 445nm, while the orange fluorescence emission of J-CDs at 578nm, as a reference signal, has almost no change, so that the probe detection system produces an obvious ratio color change from lavender to orange, thereby realizing the visual detection of cetirizine hydrochloride by the ratio fluorescence probe detection system, and detecting the content of cetirizine hydrochloride in the sample by the degree of fluorescence change in the ratio fluorescence system.

[0020] A visual detection method for cetirizine hydrochloride residue based on L / J-CDs ratio fluorescent probe: 0.6 mL of the ratio fluorescent probe is taken as a detection reagent and dripped into a 5 mL plastic centrifuge tube (EP tube), 1 mL of PBS buffer solution is added, and then 2.4 mL of cetirizine hydrochloride test solution is added, mixed and incubated for 0 to 15 minutes under a water bath temperature of 20 to 65° C., and obvious fluorescence color changes can be observed under ultraviolet light, thereby realizing visual detection of the cetirizine hydrochloride test solution;

[0021] The concentration of the buffer solution is 10 mM;

[0022] The pH range of the buffer solution is 3 to 9, preferably 3;

[0023] Preferably, the water bath temperature parameter is 25°C;

[0024] Preferably, the incubation time parameter is 1 min;

[0025] Fluorescence images and RGB values ​​were obtained using software under ultraviolet light source, and a relationship diagram between the ratio of R to G and the concentration of cetirizine hydrochloride was established to assist visual detection.

[0026] A method for quantitative detection of cetirizine hydrochloride residues based on L / J-CDs ratio fluorescent probe, comprising:

[0027] 0.6 mL of the L / J-CDs ratio fluorescent probe was dropped into a 5 mL plastic centrifuge tube (EP tube), 1 mL of PBS buffer solution was added, and then 2.4 mL of the cetirizine hydrochloride test solution was added, mixed and incubated for 0 to 15 min in a water bath temperature of 20 to 65° C., and the fluorescence emission spectrum of the solution in the range of 380 to 680 nm was recorded with an excitation wavelength of 355 nm. The linear relationship between the change in the fluorescence emission peak intensity ratio and the cetirizine hydrochloride concentration was established to achieve quantitative detection of the cetirizine hydrochloride test solution;

[0028] The concentration of the buffer solution is 10 mM;

[0029] The pH range of the buffer solution is 3 to 9, preferably 3;

[0030] Preferably, the water bath temperature parameter is 25°C;

[0031] Preferably, the incubation time parameter is 1 min.

[0032] The fluorescence intensity ratio is (I 445 / I 578 )0 / (I 445 / I 578 ), specifically, (I 445 / I 578 ) is the intensity ratio of the double emission peaks of the fluorescence spectrum after adding different concentrations of cetirizine hydrochloride, (I 445 / I 578 )0 is the intensity ratio of the double emission peaks of the blank group.

[0033] Compared with the existing detection technology, the beneficial effects of the present invention are embodied in:

[0034] (1) Compared with other single emission fluorescence detection methods, the ratio fluorescence probe detection system of the present invention effectively avoids the instability of monochromatic fluorescence intensity due to matrix effects, environmental factors and instrument factors, and has better fluorescence detection reproducibility.

[0035] (2) The present invention demonstrates the effect of visual qualitative detection by directly irradiating with ultraviolet light and analyzing the RGB values ​​of fluorescent images using a smartphone.

[0036] (3) The present invention constructs a linear relationship between the fluorescence intensity ratio and the concentration of cetirizine hydrochloride through methodological research, thereby broadening the linear range of cetirizine hydrochloride content detection.

[0037] (4) The L / J-CDs ratio fluorescent probe prepared by the present invention has a simple preparation method, low material cost, is portable and responds quickly to cetirizine hydrochloride. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 Transmission electron microscopy images of L-CDs (A) and J-CDs (B), and the insets are the particle size distribution under dynamic light scattering technology.

[0039] Figure 2 FT-IR infrared spectra of L-CDs (blue line) and J-CDs (pink line).

[0040] Figure 3 X-ray photoelectron spectroscopy (XPS) diagrams of L-CDs ((A) full range XPS spectrum, (B) high-resolution XPS spectrum of C1s, (C) high-resolution XPS spectrum of N1s, (D) high-resolution XPS spectrum of O1s).

[0041] Figure 4 X-ray photoelectron spectroscopy (XPS) images of J-CDs ((A) full range XPS spectrum, (B) high-resolution XPS spectrum of C1s, (C) high-resolution XPS spectrum of N1s, (D) high-resolution XPS spectrum of O1s).

[0042] Figure 5 (A) Ex, Em fluorescence spectra and UV-visible absorption spectra of L-CDs, the insets are the images under natural light (left) and UV light (right), (B) fluorescence emission spectra under excitation wavelengths of 295-385 nm; (C) Ex, Em fluorescence spectra and UV-visible absorption spectra of J-CDs, the insets are the images under natural light (left) and UV light (right), (D) fluorescence emission spectra under excitation wavelengths of 520-570 nm.

[0043] Figure 6 Fluorescence spectrum (A) and corresponding chromaticity diagram (B) of the detection of cetirizine hydrochloride by L / J-CDs ratio fluorescence probe under optimized conditions.

[0044] Figure 7The effect of the volume ratio of L-CDs to J-CDs in the ratio fluorescence probe on the ability of the ratio fluorescence detection system to recognize cetirizine hydrochloride; (A) When the volume ratio of L-CDs to J-CDs is 5:6, the fluorescence spectrum of the ratio fluorescence probe detection system changes before and after the addition of cetirizine hydrochloride; (B) The effect of different volume ratios of L-CDs to J-CDs on the fluorescence intensity ratio of cetirizine hydrochloride detected by the ratio fluorescence probe.

[0045] Figure 8 The effect of the pH value of the system on the ability of the ratio fluorescence probe detection system to identify cetirizine hydrochloride; (A) When the pH is 5, the fluorescence spectrum of the ratio fluorescence probe detection system changes before and after the addition of cetirizine hydrochloride; (B) The effect of pH value on the fluorescence intensity ratio of the ratio fluorescence probe detection of cetirizine hydrochloride.

[0046] Fig. 9 The effect of incubation temperature on the ability of the ratio fluorescence probe detection system to identify cetirizine hydrochloride; (A) When the incubation temperature is 65°C, the fluorescence spectrum of the ratio fluorescence probe detection system changes before and after the addition of cetirizine hydrochloride; (B) The effect of incubation temperature on the fluorescence intensity ratio of cetirizine hydrochloride detected by the ratio fluorescence probe.

[0047] Fig.10 Figure 1 is the fluorescence spectrum of L / J-CD composite system with different concentrations of cetirizine hydrochloride (A) (the inset is the color change diagram under ultraviolet light) and the corresponding chromaticity diagram (B); (C) and (D) are the fluorescence intensity ratios of the ratiometric fluorescent probe L / J-CDs composite system (I 445 / I 578 )0 / (I 445 / I 578 ) and the concentration of cetirizine hydrochloride.

[0048] Fig.11 (A) is a schematic diagram of smartphone color recognition when different concentrations of cetirizine hydrochloride are added to the L / J-CDs ratio fluorescence probe composite system; (B) is a graph showing the relationship between the color change (R / G) value of the ratio fluorescence probe solution and the concentration of cetirizine hydrochloride.

[0049] Fig.12 Selectivity and anti-interference diagram of ratiometric fluorescence probe for detecting cetirizine hydrochloride.

[0050] Fig.13 When L-CDs is used as a fluorescent probe alone, the fluorescence spectrum changes before and after the addition of cetirizine hydrochloride (A) and the corresponding chromaticity diagram (B).

[0051] Fig.14 When J-CDs is used alone as a fluorescent probe, the fluorescence spectrum changes before and after the addition of cetirizine hydrochloride. DETAILED DESCRIPTION

[0052] For a better understanding of the present invention, the following examples are further descriptions of the present invention. The following descriptions are illustrative rather than restrictive and should not be used to limit the scope of protection of the present invention.

[0053] Example 1

[0054] The preparation steps of L / J-CDs ratio fluorescent probe are as follows:

[0055] (1) Preparation of L-CDs: 1.50 g of sodium citrate and 0.78 g of acrylamide nitrogen dopant were dispersed in 30 mL of deionized water and ultrasonicated for 10 min until completely dissolved. The resulting mixture was transferred to a 50 mL Teflon reactor and then reacted at 200 °C for 3 h. After naturally cooling to room temperature, the product was filtered through a 0.22 μm water-based microporous filter membrane to remove larger particles. Finally, the obtained solution was diluted 100 times with deionized water and stored at 4 °C. The resulting solution was the nitrogen-doped L-CDs solution.

[0056] (2) Preparation of J-CDs: 0.75 g of rhodamine B was ultrasonically dispersed in 30 mL of deionized water. The resulting solution was transferred to a 50 mL Teflon reactor and treated at 160°C for 2 h. After naturally cooling to room temperature, the resulting deep red product was centrifuged at 10,000 rpm for 10 min. After being filtered through a 0.22 μm aqueous microporous filter membrane and purified by dialysis with a 1000 Da molecular weight cutoff bag and freeze-dried, the resulting powder was dispersed in 20 L of deionized water and stored at 4°C. The resulting solution was the J-CDs solution.

[0057] (3) Preparation of L / J-CDs ratio fluorescent probe: The L-CDs solution obtained in step (1) and the J-CDs solution obtained in step (2) are fully mixed in a volume ratio of 1:1 to obtain the L / J-CDs ratio fluorescent probe.

[0058] Considering that the detection sensitivity of the probe to the analyte is related to its own properties, FT-IR, TEM, DLS, and XPS were used to determine the morphology and elemental composition of the components of the ratio fluorescence probe system. Figure 1 It can be seen that the prepared L-CDs and J-CDs are evenly dispersed, and the average particle sizes are 1.07nm and 2.45nm, respectively. Figure 2 It can be seen that there are active functional groups such as amino and hydroxyl groups on the surfaces of L-CDs and J-CDs. Figure 3 It can be seen that L-CDs are composed of C, N, and O, and there are amino and pyrrolic nitrogen on the surface, which proves that nitrogen element is successfully doped in the preparation of L-CDs. Figure 4 It also confirms the structural characteristics of J-CDs in the infrared spectrum.

[0059] In addition, the spectral characteristics of the ratiometric fluorescent probe system components were studied by UV-vis and fluorescence spectroscopy. From the fluorescence excitation and emission spectra of L-CDs, it can be seen that at the optimal excitation wavelength of 355nm, the strongest emission peak is at 445nm ( Figure 5 A), and also exhibited strong blue fluorescence under 365nm ultraviolet light source ( Figure 5 A illustration), Figure 5 B shows that the position of the fluorescence emission peak of L-CDs does not change with the change of excitation wavelength, indicating that it has no excitation dependence, which indicates that the size distribution and surface state of L-CDs are uniformly distributed. From the fluorescence excitation and emission spectra of J-CDs, it can be seen that under the optimal excitation wavelength of 555nm, the strongest emission peak is at 578nm ( Figure 5 C), and also exhibited strong orange fluorescence under 365nm ultraviolet light source ( Figure 5 C illustration), Figure 5 D shows that the J-CDs have no excitation dependence, which indicates that the size distribution and surface state distribution of J-CDs are also uniform.

[0060] The above optical property research and morphology characterization results can prove the successful preparation of the ratio fluorescence probe system components L-CDs and J-CDs, and have excellent optical properties.

[0061] Example 2

[0062] 0.6 mL of the L / J-CDs ratio fluorescent probe prepared in step (3) of Example 1 was dripped into a 5 mL EP tube, 1 mL of PBS buffer solution (10 mM, pH = 3) was added, and then 2.4 mL of cetirizine hydrochloride solution (concentration in the EP tube was 200 μM) was added, mixed and incubated for 1 min in a water bath at 25°C, and the fluorescence emission spectrum of the solution in the range of 380 to 680 nm was recorded with an excitation wavelength of 355 nm. Figure 6 It can be seen that the fluorescence emission peak at 445 nm is significantly reduced after the addition of cetirizine hydrochloride.

[0063] Example 3

[0064] The L-CDs synthesized in step (1) of Example 1 and the J-CDs synthesized in step (2) were fully mixed at a volume ratio of 5:6, and 0.6 mL was dripped into a 5 mL EP tube. 1 mL PBS buffer solution (10 mM, pH = 3) was added, and then 2.4 mL of cetirizine hydrochloride solution (concentration of 200 μM) was added. The mixture was mixed and incubated for 1 min in a water bath at 25°C. The fluorescence emission spectrum of the solution in the range of 380 to 680 nm was recorded at an excitation wavelength of 355 nm. The effect of the volume ratio of L-CDs to J-CDs on the ability of the ratiometric fluorescent probe detection system to recognize cetirizine hydrochloride was also investigated. Figure 7 It can be seen that the recognition effect of cetirizine hydrochloride is best when L-CDs and J-CDs are mixed in a volume ratio of 1:1 in the ratio fluorescence probe composite system.

[0065] Example 4

[0066] 0.6 mL of the L / J-CDs ratio fluorescent probe prepared in step (3) of Example 1 was dripped into a 5 mL EP tube, 1 mL of PBS buffer solution (10 mM, pH = 5) was added, and then 2.4 mL of cetirizine hydrochloride solution (concentration of 200 μM) was added. The mixture was mixed and incubated for 1 min in a water bath at 25°C. The fluorescence emission spectrum of the solution in the range of 380 to 680 nm was recorded with an excitation wavelength of 355 nm. The effect of pH value on the ability of the ratio fluorescent probe detection system to recognize cetirizine hydrochloride was also investigated. Figure 8 It can be seen that the ratiometric fluorescent probe L / J-CDs composite system has the best recognition effect on cetirizine hydrochloride at pH = 3.

[0067] Example 5

[0068] 0.6 mL of the L / J-CDs ratiometric fluorescent probe prepared in step (3) of Example 1 was dripped into a 5 mL EP tube, 1 mL of PBS buffer solution (10 mM, pH = 3) was added, and then 2.4 mL of cetirizine hydrochloride solution (concentration of 200 μM) was added. The mixture was mixed and incubated in a water bath at 65°C for 1 min, and the fluorescence emission spectrum of the solution in the range of 380 to 680 nm was recorded at an excitation wavelength of 355 nm. The effect of different incubation temperatures on the ability of the ratiometric fluorescent probe detection system to recognize cetirizine hydrochloride was investigated. Fig. 9 It can be seen that the ratiometric fluorescent probe L / J-CDs composite system has the best recognition effect on cetirizine hydrochloride at an incubation temperature of 25°C.

[0069] Example 6

[0070] The L / J-CDs ratio fluorescent probe solution is used for the detection of cetirizine hydrochloride, and the steps are as follows:

[0071] (1) 0.6 mL of the L / J-CDs ratio fluorescent probe prepared in step (3) of Example 1 was dripped into a 5 mL EP tube, 1 mL of PBS buffer solution (10 mM, pH = 3) was added, and then 2.4 mL of cetirizine hydrochloride solution of different concentrations (concentration range is 0.05-300 μM) was added. The mixture was mixed and incubated for 1 min in a water bath at 25° C. The fluorescence emission spectrum of the solution in the range of 380-680 nm was recorded with an excitation wavelength of 355 nm. By establishing the fluorescence emission peak intensity ratio (I 445 / I 578 )0 / (I 445 / I 578 ) changes and the concentration of cetirizine hydrochloride to achieve quantitative detection of cetirizine hydrochloride.

[0072] (2) Drawing of standard curve: After adding different concentrations of cetirizine hydrochloride solution to the composite system of L / J-CDs ratio fluorescence probe, the fluorescence intensity was tested. The results showed that the blue fluorescence at 445 nm was gradually quenched, and the orange fluorescence at 578 nm remained basically unchanged. By establishing the fluorescence emission peak intensity ratio (I 445 / I 578 )0 / (I 445 / I 578 ) and the concentration of cetirizine hydrochloride, the quantitative detection of cetirizine hydrochloride can be achieved. Fig.10 It can be seen from the fluorescence spectrum that with the increase of the concentration of cetirizine hydrochloride added, the fluorescence emission peak intensity ratio (I 445 / I 578 )0 / (I 445 / I 578 ) has two linear relationships with the concentration of cetirizine hydrochloride, and the calculated detection limit is 10.2nM. Compared with the reported single-emission carbon dot N-CDs detection system for the determination of cetirizine, the linear range is expanded from 0.08-48μΜ to 0.05-60μΜ and 60-290μΜ.

[0073] Example 7

[0074] The L / J-CDs ratio fluorescent probe solution is used for visual detection of cetirizine hydrochloride, and the steps are as follows:

[0075] (1) Construction of a smartphone-assisted visualization detection platform: including enzyme label strips, UV dark box, UV lamp and smartphone.

[0076] (2) Visual detection of cetirizine hydrochloride: The preferred detection conditions of step (1) of Example 6 were used to mix the ratiometric fluorescent probe solution and cetirizine hydrochloride of different concentrations, and then titrated onto the enzyme label strip, respectively. The fluorescence color change can be visually observed in a dark environment. The fluorescence image can be captured by a smartphone, and the RGB value of the image can be further identified.

[0077] (3) Drawing of standard curve: In a dark environment, the RGB value of the fluorescent color was further identified by irradiation with 365nm ultraviolet light and fluorescence images captured by a smartphone. It was observed that the R / G value showed a linear relationship with the concentration of cetirizine hydrochloride ( Fig.11 ).

[0078] Example 8

[0079] L / J-CDs ratiometric fluorescent probe solution for specific detection of cetirizine hydrochloride

[0080] Using the preferred detection conditions in step (1) of Example 6, 200 μM of CTZ, promethazine hydrochloride, tobramycin sulfate, chlorpheniramine, fluconazole, fluoxetine hydrochloride and some amino acids (such as phenylalanine Phe, glycine Gly, threonine Thr, histidine His, arginine Arg, leucine Leu, methionine Met) were added to the ratiometric fluorescent probe detection system. Only CTZ greatly increased (I 445 / I 578 )0 / (I 445 / I 578 ) values. When different substrates (200 μM) coexisted with CTZ (200 μM), other substrates had almost no effect on the detection of CTZ. 500 μM of metal cations (Cd 2+ 、Al 3+ , K + Mg 2+ 、Zn 2+ 、Na +) and common anions (SO4 2- 、NO3 - , Cl - ), fluorescence intensity ratio (I 445 / I 578 )0 / (I 445 / I 578 ) remains basically unchanged, such as Fig.12 The results show that the system has good selectivity and anti-interference for cetirizine hydrochloride.

[0081] Comparative Example 1

[0082] L-CDs as a fluorescent probe for the detection of cetirizine hydrochloride

[0083] (1) Preparation of L-CDs: The preparation process of this step is the same as that of Example 1.

[0084] (2) 0.3 mL of L-CDs was dropped into a 5 mL EP tube, 1 mL of PBS buffer solution (10 mM, pH = 3), 0.3 mL of deionized water, and 2.4 mL of cetirizine hydrochloride solution (concentration of 200 μM) were added, mixed and incubated for 1 min in a water bath at 25°C, and the fluorescence emission spectrum of the solution in the range of 380 to 680 nm was recorded with an excitation wavelength of 355 nm. Fig.13 It can be seen that the fluorescence emission intensity is significantly quenched after the addition of cetirizine hydrochloride.

[0085] Comparative Example 2

[0086] J-CDs alone as a fluorescent probe for the detection of cetirizine hydrochloride

[0087] (1) Preparation of J-CDs: The preparation process of this step is the same as that of Example 1.

[0088] (2) 0.3 mL of J-CDs was dropped into a 5 mL EP tube, 1 mL of PBS buffer solution (10 mM, pH = 3), 0.3 mL of deionized water, and 2.4 mL of cetirizine hydrochloride solution (concentration of 200 μM) were added, mixed and incubated in a water bath at 25°C for 1 min, and the fluorescence emission spectrum of the solution in the range of 380 to 680 nm was recorded with an excitation wavelength of 355 nm. Fig.14 It can be seen that the fluorescence emission intensity does not change substantially after the addition of cetirizine hydrochloride.

[0089] The present invention investigates the results of using the fluorescent probes prepared in Example 2, Comparative Example 1 and Comparative Example 2 for the detection of cetirizine hydrochloride. Figure 6 and Fig.13 It can be seen that after adding cetirizine hydrochloride, when L-CDs is used as a single fluorescent probe or a ratio fluorescent probe, the fluorescence emission intensity at 445nm will be significantly reduced with the addition of cetirizine hydrochloride. Fig.13 The fluorescence color change in the ratio fluorescence probe detection system shown in the CIE chromaticity diagram has a larger span than that of the single-emission fluorescence probe, which is conducive to visual resolution. Fig.14 The fluorescence emission intensity did not change significantly, and J-CDs could not be used alone for the detection of cetirizine hydrochloride.

[0090] Obviously, the use of L / J-CDs ratiometric fluorescent probe for the detection of cetirizine hydrochloride is conducive to the development of a simple method for visually assisted detection.

[0091] The above is only a preferred embodiment of the present invention, and certainly cannot be used to limit the scope of the present invention. For ordinary technicians in this technical field, without departing from the principle of the present invention, some improvements can be made using the technical content suggested above, and these improvements and changes are also considered to be within the scope of protection of the present invention.

Claims

1. A method for preparing a ratiometric fluorescent probe based on L / J-CDs, characterized in that: The method comprises the following steps: (1) Synthesis of L-CDs: 1.50 g sodium citrate and 0.78 g acrylamide nitrogen dopant were dispersed in 30 mL deionized water and ultrasonicated for 10 min until completely dissolved. The resulting mixture was transferred to a 50 mL Teflon reactor and then reacted at 200 °C for 3 h. After naturally cooling to room temperature, the product was filtered through a filter membrane to remove larger particles. Finally, the obtained solution was diluted 10 to 1000 times with deionized water and stored at 4 °C. The resulting solution was the nitrogen-doped L-CDs solution. (2) Synthesis of J-CDs: 0.75 g of rhodamine B was ultrasonically dispersed in 30 mL of deionized water. The resulting solution was transferred to a 50 mL Teflon reactor and treated at 160°C for 2 h. After naturally cooling to room temperature, the resulting product was centrifuged at 10,000 rpm for 10 min. After purification by dialysis and freeze-drying, the resulting powder was dispersed in 20 L of deionized water and stored at 4°C. The resulting solution was the J-CDs solution. (3) Preparation of L / J-CDs ratio fluorescent probe: The L-CDs solution obtained in step (1) and the J-CDs solution obtained in step (2) are fully mixed at a volume ratio of 2:3 to 5:3 to obtain a L / J-CDs ratio fluorescent probe.

2. The method for preparing a ratiometric fluorescent probe based on L / J-CDs according to claim 1, characterized in that: The filter membrane in step (1) is a 0.22 μm water-based microporous filter membrane.

3. The method for preparing a ratiometric fluorescent probe based on L / J-CDs according to claim 1, characterized in that: The dilution factor in step (1) is 100 times.

4. The method for preparing a ratiometric fluorescent probe based on L / J-CDs according to claim 1, characterized in that: The dialysis purification in step (2) is performed by filtering through a 0.22 μm microporous filter membrane and dialysis purification using a dialysis bag with a molecular weight cutoff of 1000 Da.

5. The method for preparing a ratiometric fluorescent probe based on L / J-CDs according to claim 1, characterized in that: In step (3), the volume ratio of the L-CDs solution to the J-CDs solution is 1:

1.

6. A ratiometric fluorescent probe based on L / J-CDs, characterized in that: The ratio fluorescent probe is prepared by the method according to any one of claims 1 to 5, and the ratio fluorescent probe is composed of a composite system of L-CDs emitting blue fluorescence and J-CDs emitting orange fluorescence.

7. Use of the L / J-CDs-based ratiometric fluorescent probe according to claim 6 in the preparation of a detection reagent, characterized in that: The detection reagent is used for visual detection or quantitative detection of cetirizine hydrochloride.

8. The use according to claim 7, characterized in that: The application comprises the following steps: taking 0.6 mL of the ratio fluorescent probe as a detection reagent and dropping it into a 5 mL plastic centrifuge tube, adding 1 mL of PBS buffer solution, and then adding 2.4 mL of a cetirizine hydrochloride test solution, mixing and incubating for 0 to 15 minutes under a water bath temperature of 20 to 65° C., and observing obvious fluorescent color changes under ultraviolet light to achieve visual detection of the cetirizine hydrochloride test solution.

9. The use according to claim 8, characterized in that: Fluorescence images and RGB values ​​were obtained using software under ultraviolet light, and a relationship diagram between the ratio of R to G and the concentration of cetirizine hydrochloride was established to assist in visual detection.

10. The use according to claim 7, characterized in that: 0.6 mL of the ratio fluorescent probe is taken as a detection reagent and dropped into a 5 mL plastic centrifuge tube, 1 mL of PBS buffer solution is added, and then 2.4 mL of the cetirizine hydrochloride test solution is added, and the mixture is mixed and incubated for 0 to 15 minutes under a water bath temperature of 20 to 65° C., and the fluorescence emission spectrum of the solution in the range of 380 to 680 nm is recorded with an excitation wavelength of 355 nm. The relationship between the change in the fluorescence emission peak intensity ratio and the cetirizine hydrochloride concentration is established to achieve quantitative detection of the cetirizine hydrochloride test solution.

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