Dye-derived red fluorescent carbon dots and their preparation method and application

Abstract

The present invention belongs to the technical field of carbon-based nanoluminescent materials, and provides a dye-derived red fluorescent carbon dot, a preparation method and application thereof. Azure A dye and ethylenediaminetetraacetic acid are used as reaction precursors, and a hydrothermal reaction at 180-220°C can obtain the dye-derived red fluorescent carbon dots. The emission wavelength of the carbon dots is extended to 684 nm. The prepared carbon dots do not require surface passivation or complex modification, and the process is simple, while exhibiting excellent optical properties and good stability. A highly sensitive fluorescence sensing method for amaranth has been developed to achieve efficient and specific detection of amaranth with excellent selectivity and sensitivity. Based on the reversible change of the solution from azure to red during the response of the carbon dots to amaranth, a smartphone recognition application was constructed, which greatly improved the convenience and practicality of the detection process.

Description

Technical Field

[0001] The present invention belongs to the technical field of carbon-based nanoluminescent materials, and specifically relates to a dye-derived red fluorescent carbon dot and its preparation method and application. More specifically, the fluorescent carbon dot is used for smartphones to identify and detect amaranth. Background Art

[0002] Amaranth is a synthetic food colorant and a water-soluble azo dye widely used in food, flavoring, pharmaceutical formulations, and cosmetics. Its exceptional stability under various processing conditions, including high temperatures and strong light, makes it a highly regarded and reliable colorant in the food industry. However, studies have shown that amaranth may pose a potential threat to human health, known to trigger allergic reactions and hyperactivity in children. While amaranth can effectively enhance the visual appeal of food, its health risks and associated regulatory restrictions cannot be ignored. In recent years, research on the intake of synthetic pigments has steadily increased both domestically and internationally. Reported methods for detecting food pigments include high-performance liquid chromatography, electrochemical methods, UV-visible absorption spectroscopy, microcolumn methods, and polarography. Fluorescence has become an important complementary detection method due to its ease of operation, high sensitivity, rapid detection, and convenient analysis. However, research on the development of amaranth fluorescence sensors remains limited, and high-quality fluorescence detection methods for amaranth are still scarce.

[0003] Carbon dots (Cdots) have become a research hotspot in the field of nanomaterials due to their excellent optical properties, especially in environmental monitoring and food safety testing, which have attracted widespread attention. Although significant progress has been made in using Cdots for trace detection of food pigments, the existing Cdot fluorescence emission is mainly concentrated in the short-wavelength region and is easily interfered by the autofluorescence of biological tissues and the environmental background. In the process of extending the emission wavelength of Cdots to the required long-wavelength range, the formation of photoluminescence is usually related to the formation of polyheterocyclic molecular domains during synthesis, resulting in low emission efficiency and difficulty in precisely controlling optical properties. Considering the importance of red light in biological imaging and sensing applications, it is urgent to develop suitable fluorophores as precursors to achieve the extension of the emission wavelength of Cdots to the long-wavelength region, especially around 700 nm. Summary of the Invention

[0004] The present invention provides dye-derived red fluorescent carbon dots, as well as a preparation method and application thereof. The present invention successfully prepares red fluorescent carbon dots for the first time using Azure A dye and ethylenediaminetetraacetic acid as reaction precursors, and constructs a highly sensitive fluorescence sensing method for amaranth. Through the reversible change of the solution between azure and red during the response of the fluorescent carbon dots to amaranth, simple identification based on a smartphone is achieved.

[0005] The present invention also provides a method for preparing red fluorescent carbon dots. This method is simple to operate, uses a wide range of raw materials, and is low-cost, showing great potential for widespread application. Furthermore, based on the synthesized carbon dots, the present invention further develops a highly specific fluorescence sensing method for amaranth.

[0006] The present invention is achieved by the following technical solution: a dye-derived red fluorescent carbon dot, which uses Azure A dye and ethylenediaminetetraacetic acid as reaction precursors and can be obtained by hydrothermal reaction at 18-220°C.

[0007] The method for preparing the dye-derived red fluorescent carbon dots is as follows:

[0008] (1) Grind Azure A and EDTA evenly, then dissolve in secondary water, and ultrasonicate to obtain a uniform mixed solution; wherein the mass ratio of Azure A, secondary water and EDTA is 1-4:400-1000:1-10;

[0009] (2) Hydrothermal reaction: The above solution was transferred to a hydrothermal reactor and reacted at 180-220°C for 6-10 hours. After the reaction, it was allowed to cool to room temperature. The insoluble matter was removed by filtration and centrifuged at 10,000 rpm to collect the supernatant. Subsequently, the supernatant was dialyzed using a 500-1000 Da dialysis bag for 1-3 days to obtain a pure carbon dot aqueous solution. After the dialysis, freeze drying was performed. The freeze drying conditions were set as follows: initial temperature: -80°C, heating rate: 0.5°C / min, to ensure that the water in the solution sublimated slowly to avoid damage to the carbon dot structure. The freeze drying process lasted for 24-48 hours until a completely dry carbon dot solid powder was obtained.

[0010] Furthermore, the mass ratio of the azure A, secondary water and ethylenediaminetetraacetic acid is 1:400:1; the temperature of the hydrothermal reaction is 200° C. and the reaction is carried out for 8 hours; and the dialysis condition is dialysis treatment in a 500 Da dialysis bag for 3 days.

[0011] The present invention also provides the use of the dye-derived red fluorescent carbon dots or the dye-derived red fluorescent carbon dots prepared by the method in the simple identification of amaranth based on a smartphone.

[0012] Furthermore, the specific application method is:

[0013] (1) Preparation of R-CDs dispersion: The prepared dye-derived red fluorescent carbon dot R-CDs powder was dissolved in ultrapure water to obtain an R-CDs aqueous solution with a concentration of 3.0 mg / ml. Subsequently, 100 μl of the R-CDs aqueous solution was mixed with 2.0 ml of PBS buffer solution with a pH of 7.4, and 0-150 μM amaranth was gradually added in different concentration gradients to obtain an R-CDs dispersion.

[0014] (2) Observe the color change of the solution: The mixed sample is exposed to a light source, and the color change of the solution is captured using a smartphone camera. As the concentration of amaranth increases, the color of the carbon dot solution gradually changes from azure to red, which is recorded by image capture.

[0015] (3) Image acquisition and color analysis: The captured images are converted into RGB color values ​​through a smartphone application, and the RGB data corresponding to each image is recorded. The application automatically extracts the R, G, and B values ​​of the main colors in the image and then calculates the R / (G+B) ratio.

[0016] (4) Linear fitting and detection limit calculation: The relationship between the R / (G+B) ratio and the amaranth concentration was fitted by linear regression, and the regression equation R / (G+B)=-0.0121 C + 0.4319, R²=0.9964 was obtained. The detection limit of this method was calculated to be 1.31 μM.

[0017] This invention uses Azure A dye and ethylenediaminetetraacetic acid as precursors for the first time to successfully prepare red fluorescent carbon dots and extend the emission wavelength of the carbon dots to 684 nm. The prepared carbon dots do not require surface passivation or complex modification, the process is simple, and they exhibit excellent optical properties and good stability. A highly sensitive fluorescence sensing method for amaranth was developed to achieve efficient and specific detection of amaranth with excellent selectivity and sensitivity. Based on the reversible change of the solution from azure to red during the response of carbon dots to amaranth, a smartphone recognition application was constructed, which greatly improved the convenience and practicality of the detection process. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 This is a transmission electron micrograph of carbon dots prepared in Example 1 of the present invention; Figure 2 This is the infrared spectrum of the carbon dots prepared in Example 1 of the present invention, where the abscissa represents the detection wavelength and the ordinate represents the transmittance; Figure 3 This is the XPS spectrum of the carbon dots prepared in Example 1 of the present invention; Figure 4 The ultraviolet absorption spectrum and fluorescence excitation-emission spectrum of the carbon dots prepared in Example 1 of the present invention are shown; Figure 5 This is the excitation-emission fluorescence matrix spectrum of the carbon dots prepared in Example 1 of the present invention; Figure 6 The results of the ion selectivity study of the carbon dots prepared in Example 1 of the present invention at 684 nm fluorescence intensity are shown; Figure 7 This is a graph showing the fluorescence changes and related linear relationships of the carbon dots prepared in Example 1 of the present invention under different amaranth concentrations; Figure 8 This is a diagram illustrating the application of amaranth imaging and recognition with the assistance of a smartphone based on the carbon dots prepared in Example 1 of the present invention. DETAILED DESCRIPTION

[0019] In order to make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are part of the embodiments of the present invention, not all the embodiments; based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0020] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs, and the disclosure and materials cited therein are hereby incorporated by reference.

[0021] Technical equivalents to the specific embodiments described that are apparent to those skilled in the art using no more than routine experimentation are intended to be encompassed by this application.

[0022] The experimental methods in the following examples, unless otherwise specified, are all conventional methods. The instruments and equipment used in the following examples, unless otherwise specified, are all conventional laboratory instruments and equipment; the experimental materials used in the following examples, unless otherwise specified, are all purchased from conventional biochemical reagent stores.

[0023] Example 1: Preparation of dye-derived red fluorescent carbon dots, the specific method is as follows:

[0024] Step 1: Weigh 0.05 g of Azure A and 0.05 g of ethylenediaminetetraacetic acid in a mortar and grind them thoroughly. Then, dissolve them in 20 mL of secondary water and sonicate to obtain a uniform mixed solution. Transfer the solution to a 50 mL hydrothermal reactor.

[0025] Step 2: Place the hydrothermal reactor in an oven and react at 200° C. for 8 hours to obtain a red solution.

[0026] Step 3: After filtering the insoluble matter, dialyze the solution in a glass container using a 500 Da dialysis bag for 3 days to obtain a pure carbon dot aqueous solution.

[0027] Step 4: freeze-dry the fluorescent carbon dot aqueous solution to obtain red fluorescent carbon dots with a relative quantum yield of 14.6%.

[0028] The structural characterization of the carbon dots prepared in Example 1 is shown in Figure 1 、 Figure 2 and Figure 3 . Figure 1Transmission electron microscopy (TEM) image of carbon dots prepared in Example 1 shows that the average particle size of the carbon dots is 4.90±0.07 nm, and high-resolution TEM shows a lattice spacing of 0.22 nm, corresponding to the (100) crystal plane of graphitic carbon.

[0029] Figure 2 This is the infrared spectrum (FT-IR) of carbon dots prepared in Example 1. The carbon dots have an FT-IR wavelength of 3447 cm - There is a characteristic peak at ¹, corresponding to the stretching vibration of OH / NH; 2833 cm - ¹ and 2715 cm - The peaks at ¹ are attributed to the asymmetric and symmetric stretching vibrations of CH, respectively; 1605 cm - The peak at ¹ indicates the stretching vibration of C=O; while the peak at 1365 cm - ¹ and 1124 cm - The peaks of ¹ are associated with the vibrations of CN and CO, respectively.

[0030] Figure 3 This is the full X-ray photoelectron spectroscopy (XPS) spectrum of carbon dots prepared in Example 1 of the present invention, where C 1s 、N 1s and O 1s The main peaks appear at atomic percentages of 59.04%, 13.00%, and 27.96%, respectively, with corresponding binding energies at 284.8 eV, 400.2 eV, and 531.1 eV. The combined results indicate that the carbon dots' structural characteristics are characterized by their internal sp² aromatic conjugated domains interspersed with sp³ defect sites, and their surface modification with hydrophilic functional groups such as -NH² and -COOH. These complex structural elements collectively impart unique electronic and chemical properties to the material.

[0031] The optical properties of the carbon dots prepared in Example 1 are as follows: Figure 4 and Figure 5 shown. Figure 4 The UV absorption spectrum and fluorescence excitation-emission spectrum of the carbon dots prepared in Example 1 are shown. The UV absorption spectrum shows two distinct absorption peaks at approximately 200 nm and 292 nm, corresponding to π-π transitions in the aromatic structure and n-π transitions of the C=O and C=N groups, respectively. In addition, a broad absorption peak was observed at 655 nm, associated with low-energy surface states. Fluorescence excitation-emission spectroscopy indicates that the optimal excitation wavelength (λex) and emission wavelength (λem) of the carbon dots are 666 nm and 684 nm, respectively. Figure 5 The excitation-emission fluorescence matrix spectrum of the carbon dots prepared in Example 1 shows that when the excitation wavelength changes from 300 nm to 680 nm, the emission peak position of the carbon dots at 684 nm remains constant, which is attributed to the uniform surface state of the carbon dots.

[0032] The results of the study on the responsiveness of the carbon dots prepared in Example 1 to amaranth are shown in FIG. Figure 6 and Figure 7 In the standard operation, the prepared R-CDs powder was first dissolved in ultrapure water to obtain a concentration of 3.0 mg·mL - Subsequently, 100 μL of R-CDs solution (concentration 3.0 mg mL - ¹) was mixed with 2.0 mL of PBS buffer (pH = 7.4), and various concentrations of amaranth (0-150 µM) were gradually added to prepare R-CDs dispersions. Fluorescence emission spectra of each sample were measured at an excitation wavelength of 666 nm, with each measurement repeated three times. The interfering effects of other chemicals in the presence of R-CDs were also evaluated using the same experimental procedures as for amaranth. Figure 6 It was shown that in the presence of different metal ions, representative biological molecules and food additives, the fluorescence intensity of R-CDs did not change significantly. Only amaranth could significantly inhibit the fluorescence emission of carbon dots, showing obvious selectivity. Figure 7 Further confirmation was obtained that the fluorescence intensity of the carbon dots at 684 nm decreased significantly with increasing amaranth concentration, and exhibited a good linear response within the concentration range of 0.1-2.5 µM. The linear regression equation was ΔF = 209553.5885C + 11749.3502, R² = 0.9917, and the limit of detection (LOD) was 16.7 nM. These results demonstrate that the prepared R-CDs have excellent selectivity and sensitivity and can be used for the efficient detection of amaranth.

[0033] The application of smartphone-assisted amaranth imaging recognition based on the carbon dots prepared in Example 1 is as follows Figure 8 shown. Figure 8The assay process is demonstrated: 1) Sample Preparation: First, amaranth solutions of varying concentrations are mixed with a carbon dot solution to ensure uniform mixing and a stable dispersion. Stirring or ultrasonic treatment can be used to ensure complete dissolution and uniform distribution of the amaranth. 2) Observation of Solution Color Change: The mixed sample is exposed to an appropriate light source, and the color change of the solution is captured using a smartphone camera. As the amaranth concentration increases, the color of the carbon dot solution gradually changes from azure to red, and this change can be captured using an image. 3) Image Acquisition and Color Analysis: Images of the solution are captured using a smartphone camera, ensuring that factors such as the angle and lighting source have minimal impact on color recognition. A smartphone application converts the captured images into RGB color values, and the corresponding RGB data for each image is recorded. The application automatically extracts the R, G, and B values ​​of the primary colors in the image. 4) Data Processing and Analysis: The extracted RGB values ​​are input into data processing software to calculate the R / (G+B) ratio. By comparing the R / (G+B) ratio at different amaranth concentrations, a linear curve between the concentration and the ratio was drawn, and their correlation was further analyzed. 5) Linear fitting and detection limit calculation: The relationship between the R / (G+B) ratio and the amaranth concentration was fitted by linear regression, and the regression equation R / (G+B)=-0.0121 C + 0.4319, R²=0.9964 was obtained, and the detection limit of this method was calculated to be 1.31μM. The results showed that the smartphone-assisted imaging colorimetry method still has high detection sensitivity at low amaranth concentrations. 6) Field application and validation: Using a smartphone and its application, rapid quantitative detection of amaranth was carried out in a field environment. This convenient detection method does not rely on traditional large-scale laboratory equipment and can provide accurate detection results under real-time conditions, significantly improving the efficiency and flexibility of amaranth detection.

[0034] Example 2: Preparation of dye-derived red fluorescent carbon dots. Except that the amount of EDTA was 0.02 g, the other conditions were the same as those in Example 1. The measured relative quantum yield was 11.2%.

[0035] Example 3: Preparation of dye-derived red fluorescent carbon dots. Except that the amount of EDTA was 0.08 g, the other conditions were the same as those in Example 1. The measured relative quantum yield was 8.2%.

[0036] Example 4: Preparation of dye-derived red fluorescent carbon dots. Except that the amount of EDTA was 0.15 g, the other conditions were the same as those in Example 1. The measured relative quantum yield was 7.8%.

[0037] Example 5: Preparation of dye-derived red fluorescent carbon dots: The mass ratio of Azure A, secondary water, and EDTA was controlled to be 4:800:10; the hydrothermal reaction temperature was 180°C for 10 h; and the dialysis treatment was performed using a 500 Da dialysis bag for 2 days. The remaining methods were the same as those described in Example 1.

[0038] Example 6: Preparation of dye-derived red fluorescent carbon dots: The mass ratio of Azure A, secondary water, and EDTA was controlled to be 3:600:5; the hydrothermal reaction temperature was 220°C for 6 hours; and the dialysis treatment was performed using a 1000Da dialysis bag for 1 day. The remaining methods were the same as those described in Example 1.

[0039] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the above embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A dye-derived red fluorescent carbon dot, characterized by: Using Azure A dye and ethylenediaminetetraacetic acid as reaction precursors, dye-derived red fluorescent carbon dots can be obtained by hydrothermal reaction at 180-220℃.

2. The method for preparing the dye-derived red fluorescent carbon dots according to claim 1, characterized in that: The specific method is: (1) Grind Azure A and EDTA evenly, then dissolve in secondary water, and ultrasonicate to obtain a uniform mixed solution; wherein the mass ratio of Azure A, secondary water and EDTA is 1-4:400-1000:1-10; (2) Hydrothermal reaction: The above solution was transferred to a hydrothermal reactor and reacted at 180-220°C for 6-10 hours. After the reaction, the solution was allowed to cool to room temperature, insoluble matter was removed by filtration, and the solution was centrifuged at 10,000 rpm to collect the supernatant. Subsequently, the supernatant was dialyzed using a 500-1000 Da dialysis bag for 1-3 days to obtain a pure carbon dot aqueous solution. After the dialysis, the solution was freeze-dried. The freeze-drying conditions were set as follows: initial temperature: -80°C, heating rate controlled at 0.5°C / min, and freeze-drying process lasted for 24-48 hours until completely dry carbon dot solid powder was obtained.

3. The preparation method according to claim 2, characterized in that: The mass ratio of the azure A, secondary water and ethylenediaminetetraacetic acid is 1:400:1; the temperature of the hydrothermal reaction is 200° C. and the reaction is carried out for 8 hours; and the dialysis condition is dialysis treatment in a 500 Da dialysis bag for 3 days.

4. Use of the dye-derived red fluorescent carbon dots according to claim 1 or the dye-derived red fluorescent carbon dots prepared by the method according to claim 2 in the simple identification of amaranth based on a smartphone.

5. Use of the dye-derived red fluorescent carbon dots prepared by the method of claim 3 in the simple identification of amaranth based on a smartphone, characterized in that: The specific application method is: (1) Preparation of R-CDs dispersion: The prepared dye-derived red fluorescent carbon dot R-CDs powder was dissolved in ultrapure water to obtain an R-CDs aqueous solution with a concentration of 3.0 mg / ml. Subsequently, 100 μl of the R-CDs aqueous solution was mixed with 2.0 ml of PBS buffer solution with a pH of 7.4, and 0-150 μM amaranth was gradually added in different concentration gradients to obtain an R-CDs dispersion. (2) Observe the color change of the solution: The mixed sample is exposed to a light source, and the color change of the solution is captured using a smartphone camera. As the concentration of amaranth increases, the color of the carbon dot solution gradually changes from azure to red, which is recorded by image capture. (3) Image acquisition and color analysis: The captured images are converted into RGB color values ​​through a smartphone application, and the RGB data corresponding to each image is recorded. The application automatically extracts the R, G, and B values ​​of the main colors in the image and then calculates the R / (G+B) ratio. (4) Linear fitting and detection limit calculation: The relationship between the R / (G+B) ratio and the amaranth concentration was fitted by linear regression, and the regression equation R / (G+B)=-0.0121 C + 0.4319, R²=0.9964 was obtained. The detection limit of this method was calculated to be 1.31 μM.

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