Non-excitation dependent carbon dot and preparation method and fluorescence detection application thereof

Abstract

The invention relates to the technical field of carbon dot preparation and application, in particular to a non-excitation dependent carbon dot and a preparation method and fluorescence detection application thereof. The pyrogallic acid is introduced into a thiourea-o-phenylenediamine-DMF solvothermal system, the pyrogallic acid serves as a surface state engineering agent and a structure-directing agent to participate in carbon dot formation, part of high-energy non-radiation composite defects are repaired, and the types of surface functional groups are promoted to tend to be single; meanwhile, the pyrogallic acid can reconstruct a surface electronic state, and a plurality of originally dispersed and excitation-dependent surface emission centers are integrated into a single dominant luminescence center, so that an effective luminescence transition path in the carbon dot tends to be single, and the carbon dot has good non-excitation dependence. When the carbon dots are applied to detection of tetracycline hydrochloride, the stability of fluorescence signals, the repeatability of detection results and the accuracy of quantitative analysis are improved, so that rapid, sensitive and convenient detection of tetracycline hydrochloride is realized.

Description

Technical Field

[0001] This application relates to the field of carbon dot preparation and application technology, specifically to an excitation-independent carbon dot, its preparation method, and its fluorescence detection application. Background Technology

[0002] Modern analytical detection technologies are developing towards high efficiency, greenness, and portability. Traditional fluorescent probes generally suffer from high cost, significant biotoxicity, and insufficient stability, making it difficult to simultaneously meet the diverse needs of rapid and accurate detection in fields such as environmental monitoring, biomedicine, and food safety. Carbon dots, as a novel zero-dimensional carbon-based fluorescent nanomaterial, possess advantages such as wide availability of raw materials, mild preparation processes, good water solubility, easy surface functionalization, good biocompatibility, no heavy metal toxicity, and strong resistance to photobleaching. They can maintain relatively stable fluorescence signals in complex systems, thus becoming an important research direction in the field of fluorescent probes and providing a new material basis for the development of accurate and convenient detection technologies.

[0003] However, excitation dependence is an inherent optical property of most common carbon dots. Excitation dependence refers to the significant shift in the maximum emission wavelength of a carbon dot as the excitation wavelength changes. This is typically manifested as a synchronized redshift in emission wavelength and fluctuations in fluorescence intensity when the excitation wavelength redshifts. This is usually caused by the uneven particle size distribution and diverse types of surface defect states and luminescent sites during carbon dot synthesis, resulting in multiple luminescent centers with different energies within the carbon dot. When excited by light of different wavelengths, photons of different energies selectively excite surface states under different chemical conditions, thus causing the emission spectrum to change with the excitation wavelength. Surface states typically originate from functional groups, defects, and heteroatom doping on the carbon dot surface, introducing a series of discrete trap energy levels into the carbon nucleus band gap.

[0004] Due to this ubiquitous excitation-dependent characteristic, conventional carbon dots have significant limitations in probe applications. On one hand, the emission wavelength is not fixed, making it difficult to match with standardized detection equipment and increasing the complexity of device and integrated design. On the other hand, the fluorescence signal is easily affected by fluctuations in excitation conditions, leading to decreased repeatability and reliability of detection data, thus increasing the difficulty of signal acquisition and data analysis, and hindering the accurate and stable quantitative detection of trace targets. In contrast, excitation-independent carbon dots can maintain a essentially constant maximum emission wavelength under different excitation wavelengths, with only a small, controllable change in fluorescence intensity. Therefore, they are more suitable for adaptation to single-wavelength detection devices, more conducive to the development of portable, integrated rapid detection instruments, and can effectively avoid excitation condition interference, significantly improving the stability, reliability, and repeatability of detection results.

[0005] Furthermore, tetracycline hydrochloride, as a common antibiotic, plays a significant role in residue detection in environmental analysis, food safety, and biological sample testing. Existing research indicates that carbon dot materials can be used to construct fluorescent probes for tetracycline hydrochloride, enabling a certain degree of quantitative detection. However, existing carbon dot probe systems generally suffer from problems such as fluorescence emission being significantly affected by excitation conditions, insufficient probe signal stability, and unsatisfactory detection repeatability, thus limiting their further application in highly sensitive, rapid, and low-cost quantitative detection. Summary of the Invention

[0006] To address the above problems, this invention provides a non-excitation-dependent carbon dot, which is prepared from pyrogallol, thiourea, and o-phenylenediamine using a solvothermal method. This pyrogallol-thiourea-o-phenylenediamine carbon dot is abbreviated as PTO-CDs.

[0007] On the other hand, the present invention provides a method for preparing non-excitation-dependent carbon dots, comprising the following steps: Step 1: Place pyrogallol, thiourea and o-phenylenediamine in a reaction vessel, add DMF solvent, and mix to obtain a precursor mixture; Step 2: Sonicate the precursor mixture; Step 3: Place the sealed reactor in an oven to carry out the solvothermal reaction; Step 4: After the reaction is complete, cool to room temperature; Step 5: Centrifuge to remove solid impurities; Step 6: Filter the supernatant to obtain the carbon dot solution; Step 7: Dry the carbon dot solution to obtain carbon dot powder.

[0008] In this invention, after introducing pyrogallic acid into the thiourea-o-phenylenediamine-DMF solvothermal system, pyrogallic acid does not merely exist as a common reactant, but participates in carbon dot formation as a surface engineering agent and structure directing agent. Its abundant phenolic hydroxyl groups can interact with the active sites on the carbon dot surface under high-temperature conditions to form a more uniform and stable surface coating layer, repairing some high-energy nonradiative recombination defects and promoting the simplification of surface functional groups. At the same time, pyrogallic acid can also promote the formation of a surface chemical environment characterized by thiophene sulfur, expand the π-conjugated system of carbon dots, reconstruct the surface electronic states, and integrate the originally dispersed, excitation-dependent multiple surface emission centers into a single dominant luminescent center, thereby making the effective luminescent transition path inside the carbon dot tend to be singular. Therefore, regardless of the change in external excitation light energy, photogenerated carriers will eventually relax to the lowest energy and most stable surface or molecular state through a rapid internal conversion or energy transfer process, resulting in radiative recombination and producing a basically fixed emission peak with good non-excitation dependence.

[0009] Furthermore, in step 1, the inner liner of the reactor is made of polytetrafluoroethylene.

[0010] Furthermore, in step 3, the oven temperature is 180°C and the time is 6 hours.

[0011] Furthermore, in step 6, the filter head has a pore size of 0.22 μm.

[0012] Furthermore, the use of water bath heating assisted by ultrasonic treatment in step 2 can improve the dissolution rate and dispersion uniformity of pyrogallic acid, thiourea, and o-phenylenediamine in DMF solvent without prematurely initiating a violent carbonization reaction. This allows the precursor mixture to reach a fully dissolved and homogeneous state more quickly, thereby reducing local concentration differences and uneven local nucleation. At the same time, the synergistic effect of water bath heating and ultrasonic cavitation helps pyrogallic acid to play a more uniform role as a surface engineer and structure guide in the precursor system, promoting the formation of a more uniform and stable surface chemical environment during the subsequent solvothermal reaction, reducing stray surface states and non-radiative recombination defects, and helping to further integrate the originally dispersed excitation-dependent surface emission centers into a single dominant emission channel, thereby improving the stability and repeatability of the non-excitation-dependent emission characteristics of the obtained PTO-CDs.

[0013] Furthermore, in step 1, acetonitrile is added to the DMF solvent. On the one hand, acetonitrile, as a volatile auxiliary solvent, can form a synergistic solvent system with DMF, further improving the dispersion and mixing uniformity of pyrogallic acid, thiourea, and o-phenylenediamine during ultrasonic pretreatment, reducing local concentration differences and uneven local nucleation. On the other hand, a more uniform precursor mixing environment is conducive to pyrogallic acid playing a more complete role as a surface state engineer and structure directing agent in subsequent solvothermal reactions, promoting the formation of a uniform surface chemical environment characterized by thiophene sulfur, reducing stray surface states and high-energy nonradiative recombination defects, and promoting the simplification of surface functional groups. This further simplifies the effective luminescence transition path within the carbon dots, strengthens the single dominant luminescence channel, and is more conducive to obtaining PTO-CDs with stable emission peaks and more prominent non-excitation-dependent characteristics.

[0014] Furthermore, in step 7, the carbon dot solution is rapidly pre-frozen, and then freeze-dried after the sample is fully frozen. This allows for faster physical fixation of the original dispersion state and surface microenvironment of the carbon dots in the solution, reducing local concentration, particle migration, and agglomeration caused by the slow growth of ice crystals during freezing. This avoids excessive contact accumulation between carbon dots and redistribution of surface functional groups. At the same time, this operation helps to better maintain the uniform surface chemical environment, fewer stray surface states, and a single dominant luminescence channel formed after pyrogallic acid regulation. It also prevents the reintroduction of competitive luminescence sites or non-radiative recombination defects during drying, thereby helping to maintain the technical effects of stable PTO-CD emission peaks, outstanding excitation-independent characteristics, and good detection repeatability.

[0015] In another aspect, the present invention provides an application of non-excitation-dependent carbon dots for the detection of tetracycline hydrochloride.

[0016] Furthermore, when this non-excitation-dependent carbon dot is used for the detection of tetracycline hydrochloride, the tetracycline hydrochloride sample to be tested is mixed with the diluted non-excitation-dependent carbon dot solution, the fluorescence intensity at 570 nm is detected at an excitation wavelength of 470 nm, and the concentration of tetracycline hydrochloride in the sample to be tested is determined according to the correspondence between fluorescence intensity and tetracycline hydrochloride concentration.

[0017] When the non-excitation-dependent carbon dots of this invention are applied to the detection of tetracycline hydrochloride, they can maintain a relatively stable emission peak position under different excitation conditions. Therefore, they can effectively avoid the detection error caused by emission peak drift of traditional excitation-dependent carbon dots, and improve the stability, repeatability and quantitative accuracy of the detection signal. At the same time, the non-excitation-dependent carbon dots have a clear and fixed detection window under 470 nm excitation and 570 nm emission conditions, which is more conducive to realizing rapid, sensitive and convenient detection of tetracycline hydrochloride.

[0018] The beneficial effects of this invention are: (1) The present invention uses pyrogallic acid, thiourea and o-phenylenediamine as raw materials to prepare non-excitation-dependent carbon dots by solvothermal method; pyrogallic acid can regulate the surface chemical environment of carbon dots, reduce stray surface states and non-radiative recombination defects, promote the formation of a single dominant luminescence channel, make the emission peak position of the obtained carbon dots stable and the fluorescence repeatability good, and have good optical stability and application consistency.

[0019] (2) When the non-excitation-dependent carbon dots are applied to the detection of tetracycline hydrochloride, the emission peak position is basically fixed, which avoids the detection error caused by peak position drift of traditional excitation-dependent carbon dots. This is beneficial to improve the stability of fluorescence signal, the repeatability of detection results and the accuracy of quantitative analysis, thereby realizing rapid, sensitive and convenient detection of tetracycline hydrochloride.

[0020] In summary, the present invention has good application prospects in the field of carbon dot preparation and application technology. Attached Figure Description

[0021] Figure 1 Topographic images of PTO-CDs: (a) TEM image, (b) high-resolution TEM image, (c) particle size distribution map, (d) XRD diffraction pattern; Figure 2 The structural characterization diagrams of PTO-CDs are as follows: (a) FT-IR spectrum, (b) XPS spectrum, (c) C 1s high-resolution spectrum, (d) O 1s high-resolution spectrum, (e) N 1s high-resolution spectrum, and (f) S 2p high-resolution spectrum. Figure 3 Topographic images of TO-CDs: (a) TEM image, (b) high-resolution TEM image, (c) particle size distribution map, (d) XRD diffraction pattern; Figure 4 The structural characterization diagrams of TO-CDs are as follows: (a) FT-IR spectrum, (b) XPS spectrum, (c) C 1s high-resolution spectrum, (d) O 1s high-resolution spectrum, (e) N 1s high-resolution spectrum, and (f) S 2p high-resolution spectrum. Figure 5 Optical properties of TO-CDs and PTO-CDs: (a) UV-Vis spectrum and fluorescence excitation spectrum of TO-CDs, (b) fluorescence emission spectrum of TO-CDs, (c) fluorescence two-dimensional topographic map of TO-CDs, (d) UV-Vis spectrum and fluorescence excitation spectrum of PTO-CDs, (e) fluorescence emission spectrum of PTO-CDs, (f) fluorescence two-dimensional topographic map of PTO-CDs. Figure 6 For the quantitative detection of tetracycline hydrochloride by PTO-CDs-based fluorescent probes: (a) fluorescence spectra of PTO-CDs with varying TCH concentrations (0–180 nmol / L), (b) F0–F as a function of TCH concentration. Detailed Implementation

[0022] To make the objectives, technical solutions, and advantages of this application clearer, the following detailed description is provided with reference to the accompanying drawings and embodiments. Example 1

[0023] This invention provides a method for preparing non-excitation-dependent carbon dot PTO-CDs using a solvothermal method. Specifically, 0.1 g of pyrogallol, 1.0 g of thiourea, and 1.0 g of o-phenylenediamine are weighed into a 25 mL high-pressure reactor with a polytetrafluoroethylene liner. 10 mL of DMF solvent is added, and the mixture is stirred to obtain a precursor mixture. The precursor mixture is then sonicated for 20 min until fully dissolved. The sealed high-pressure reactor is then placed in a 180 °C oven for a solvothermal reaction for 6 h. After the reaction, the mixture is allowed to cool naturally to room temperature and centrifuged at 11000 RPM for 15 min to remove solid impurities. The supernatant is then filtered three times using a 0.22 μm filter to obtain a carbon dot solution. Finally, the obtained carbon dot solution is dried in a freeze dryer to obtain non-excitation-dependent carbon dot PTO-CDs powder.

[0024] Figure 1 (a) is a transmission electron microscope (TEM) image of PTO-CDs, showing that PTO-CDs exhibit spherical nanostructure characteristics. Furthermore, Figure 1 (b) is a high-resolution transmission electron microscope (HRTEM) image of PTO-CDs. As can be seen from the figure, its lattice spacing is 0.218 nm, which is similar to the interplanar spacing of the graphene (100) crystal plane. Figure 1 (c) shows the particle size distribution of PTO-CDs, with an average diameter of approximately 2.25 nm. Figure 1 (d) is the X-ray diffraction (XRD) pattern of PTO-CDs, with a broad peak at about 23°, corresponding to a graphene-like structure

[100] , and no impurity peaks appear in the spectrum.

[0025] Figure 2 (a) shows the infrared spectrum of PTO-CDs, 3448 cm⁻¹. -1 2928 cm -1 and 1658 cm -1 The absorption peaks at 2053 cm⁻¹ correspond to the stretching vibrations of the OH / NH, CH, and C=O bonds, respectively; -1 1388 cm -1 1011 cm -1 and 663 cm -1 The absorption peaks at these locations are attributed to the stretching vibrations of the C≡C, CN, CO, and CS bonds, respectively. Furthermore, the structural characteristics of PTO-CDs were analyzed in detail using X-ray photoelectron spectroscopy (XPS), and the full XPS spectrum (…) was obtained. Figure 2(b) It can be clearly seen that the carbon point is mainly composed of four elements: C, O, N, and S, with atomic proportions of 85.18%, 13.39%, 1.43%, and 11.03%, respectively. The C 1s spectrum ( Figure 2 (c) further confirmed the presence of C=C bonds (284.80 eV), CC bonds (285.61 eV), CN / CN / CS bonds (286.63 eV), and C=O bonds (289.33 eV); O 1s spectrum ( Figure 2 (d) Contains C=O bonds (531.35 eV) and CO bonds (533.00 eV); N 1s spectrum ( Figure 2 (e) contains CN bonds (400.59 eV) and NH bonds (398.94 eV); S 2p spectrum ( Figure 2 (f) contains CS bonds (163.80 eV), thiophene sulfide S 2p1 / 2 bonds (165.09 eV), thiophene sulfide S 2p3 / 2 bonds (162.35 eV) and S=O bonds (168.69 eV).

[0026] Comparative Example This invention provides a comparative method for preparing thiourea-o-phenylenediamine carbon dots (TO-CDs) using a solvothermal method. Specifically, 1.0 g of thiourea and 1.0 g of o-phenylenediamine are weighed into a 25 mL high-pressure reactor with a polytetrafluoroethylene liner. 10 mL of DMF solvent is added, and the mixture is stirred to obtain a precursor mixture. Subsequently, the precursor mixture is sonicated for 20 min until fully dissolved. Then, the sealed high-pressure reactor is placed in an oven at 180 °C for a solvothermal reaction for 6 h. After the reaction, the mixture is naturally cooled to room temperature and centrifuged at 11000 RPM for 15 min to remove solid impurities. The supernatant is collected and filtered three times using a 0.22 μm filter to obtain a carbon dot solution. Finally, the obtained carbon dot solution is dried in a freeze dryer to obtain TO-CDs carbon dot powder.

[0027] Figure 3 (a) is a transmission electron microscope image of TO-CDs, showing that TO-CDs exhibit spherical nanostructure characteristics. Furthermore, Figure 3 (b) is a high-resolution transmission electron microscope (HRTEM) image of TO-CDs. As can be seen from the figure, its lattice spacing is 0.233 nm, which is similar to the interplanar spacing of the graphene (100) crystal plane. Figure 3 (c) shows the particle size distribution of TO-CDs, with an average diameter of approximately 2.16 nm. Figure 3(d) is the X-ray diffraction (XRD) pattern of TO-CDs, with a broad peak at about 23°, corresponding to a graphene-like structure

[100] , and no impurity peaks appear in the spectrum.

[0028] Figure 4 (a) shows the infrared spectrum of TO-CDs, at 3142 cm⁻¹. -1 2815 cm -1 and 1662 cm -1 The absorption peaks at 2057 cm⁻¹ correspond to the stretching vibrations of the OH / NH, CH, and C=O bonds, respectively; -1 1404 cm -1 1099 cm -1 and 661cm -1 The absorption peaks at these locations are attributed to the stretching vibrations of the C≡C, CN, CO, and CS bonds, respectively. Furthermore, the structural characteristics of TO-CDs were analyzed in detail using X-ray photoelectron spectroscopy (XPS), and the full XPS spectrum (…) was obtained. Figure 4 (b) It can be clearly seen that the carbon dot is mainly composed of four elements: C, O, N, and S, with atomic proportions of 62.82%, 7.44%, 14.51%, and 15.23%, respectively. The C 1s spectrum ( Figure 4 (c) further confirmed the presence of CC / C=C bonds (284.48 eV), CN / CO / CS bonds (285.70 eV), and C=O bonds (288.98 eV); O 1s spectrum ( Figure 4 (d) contains C=O bonds (531.10 eV) and CO bonds (532.75 eV); N 1s spectrum ( Figure 4 (e) contains CN bonds (399.67 eV) and NH bonds (398.05 eV); S 2p spectrum ( Figure 4 (f) contains the CS bond (162.80 eV) and the S=O bond (168.01 eV).

[0029] Figure 5 (a) The liquid UV-Vis spectrum and fluorescence excitation spectrum of TO-CDs are shown. The absorption peak at 279 nm in the UV spectrum is caused by the π-π* transition of the C=C bond, and the absorption peak at 311 nm is caused by the n-π* transition of the surface functional groups. The excitation spectrum was measured at a monitoring wavelength of 573 nm. Figure 5(b) shows the emission spectra of TO-CDs at different excitation wavelengths. It can be seen that as the excitation wavelength changes from 340 nm to 490 nm, the emission center moves from 420 nm to 581 nm. When the excitation wavelength is 470 nm, the absorption peak at 573 nm reaches its maximum intensity, which indicates the excitation dependence of TO-CDs. Figure 5 (d) Shows the liquid UV-Vis spectrum and fluorescence excitation spectrum of PTO-CDs. The absorption peak at 278 nm in the UV spectrum is caused by the π-π* transition of the C=C bond, and the absorption peak at 311 nm is caused by the n-π* transition of the surface functional groups. The excitation spectrum was measured at a monitoring wavelength of 572 nm. Figure 5 (e) shows the emission spectra of PTO-CDs at different excitation wavelengths. It can be seen that as the excitation wavelength changes from 340 nm to 490 nm, the emission center remains almost constant at around 572 nm. When the excitation wavelength is 480 nm, the absorption peak at 572 nm reaches its maximum intensity, indicating that PTO-CDs do not exhibit excitation dependence. Meanwhile, Figure 5 (c) and Figure 5 (f) are two-dimensional fluorescence topographic maps of TO-CDs and PTO-CDs, respectively. TO-CDs show a tilted pattern, while PTO-CDs show a vertical pattern, which also indicates that the addition of pyrogallic acid almost eliminates the excitation dependence. Example 2

[0030] Based on Example 1, when ultrasonicating the precursor mixture, a water bath heating-assisted ultrasonication was used. The water bath heating temperature was controlled at 35–50°C, and the ultrasonication time was controlled at 20–30 min, allowing the precursor mixture to achieve more complete dissolution and uniform dispersion under gentle heating conditions. Through the synergistic effect of water bath heating and ultrasonic cavitation, the local concentration differences of pyrogallic acid, thiourea, and o-phenylenediamine in the DMF solvent can be further reduced. This is beneficial for forming a more uniform and stable surface chemical environment during the subsequent solvothermal reaction, reducing stray surface states and non-radiative recombination defects, thereby further strengthening the single dominant luminescence channel and improving the stability and repeatability of the excitation-dependent emission characteristics of the obtained PTO-CDs. Example 3

[0031] Based on Example 2, acetonitrile was further added to the DMF solvent. Acetonitrile, as a volatile auxiliary solvent, was preferably added in an amount of 5% to 15% of the DMF volume. Taking 10 mL of DMF in Example 1 as an example, the amount of acetonitrile added was preferably 0.5 to 1.5 mL. Adding acetonitrile within the above range can form a synergistic solvent system with DMF, further improving the dispersion and mixing uniformity of pyrogallic acid, thiourea, and o-phenylenediamine under water bath heating and ultrasonic conditions, reducing local concentration differences and uneven local nucleation. At the same time, a more uniform precursor mixing environment is conducive to pyrogallic acid playing a more complete role as a surface state engineer and structure directing agent in subsequent solvothermal reactions, promoting the formation of a uniform surface chemical environment characterized by thiophene sulfide, reducing stray surface states and high-energy nonradiative recombination defects, and promoting the simplification of surface functional groups, thereby further strengthening the single dominant emission channel and improving the stability and repeatability of the non-excitation-dependent emission characteristics of the obtained PTO-CDs. Example 4

[0032] Based on Example 3, before drying the carbon dot solution, rapid pre-freezing is performed. The preferred rapid pre-freezing temperature is -60℃ to -90℃. After the sample is completely frozen, freeze-drying is then carried out. Rapid pre-freezing within the above temperature range can more quickly fix the original dispersion state and surface microenvironment of carbon dots in the solution, reduce local concentration, particle migration, and agglomeration caused by slow ice crystal growth during freezing, and avoid excessive contact accumulation between carbon dots and redistribution of surface functional groups. This is more conducive to maintaining the uniform surface chemical environment, fewer stray surface states, and a single dominant luminescence channel formed after pyrogallic acid regulation. It also prevents the reintroduction of competitive luminescence sites or non-radiative recombination defects during subsequent drying, thereby helping to maintain the technical effects of stable PTO-CD emission peaks, outstanding excitation-independent characteristics, and good detection repeatability. Example 5

[0033] This embodiment provides a method for non-excitation-dependent carbon dot detection of tetracycline hydrochloride (TCH). First, 4.809 mg of tetracycline hydrochloride is weighed into a 10 mL colorimetric tube, and distilled water is added to the mark to obtain a 1 mmol / L high-concentration tetracycline hydrochloride aqueous solution. Then, 1 mL of the above solution is taken into a 100 mL volumetric flask, and distilled water is added to the mark to obtain a 10 μmol / L medium-concentration tetracycline hydrochloride aqueous solution. Next, a 200 nmol / L tetracycline hydrochloride working stock solution is prepared using the above medium-concentration solution. Subsequently, a gradient concentration of tetracycline hydrochloride aqueous solutions (10 nmol / L, 20 nmol / L, 40 nmol / L, 60 nmol / L, 80 nmol / L, 100 nmol / L, 120 nmol / L, 140 nmol / L, 160 nmol / L, 180 nmol / L) is prepared using the working stock solution. The prepared PTO-CDs DMF solution was diluted 100 times to serve as the CDs stock solution. When testing, 2 mL of the CDs stock solution was taken and 2 mL of tetracycline hydrochloride aqueous solution of the above concentration was added to it. The fluorescence spectrum at 568 nm was recorded under 470 nm excitation.

[0034] like Figure 6 (a) and Figure 6 As shown in (b), the fluorescence intensity decreases as the concentration of tetracycline hydrochloride increases, and there is a good linear relationship between the relative PL intensity and the concentration of tetracycline hydrochloride (R0). 2 = 0.9903). The linear regression equations are (F-F0) = 5.56C + 47.71, where F0 and F represent the fluorescence intensity of PTO-CDs at 568 nm before and after the addition of TCH, respectively; and C is the concentration of TCH. In this study, the formula for calculating the detection limit in spectral analysis was adopted: . In the formula, represents the standard deviation of multiple measurements of the blank sample; K is the slope of the standard curve. The calculated limit of detection (LODs) for TCH is 3.51 nM.

[0035] In summary, this invention provides an excitation-independent carbon dot, its preparation method, and its application. This carbon dot is prepared from pyrogallic acid, thiourea, and o-phenylenediamine via a solvothermal method. The pyrogallic acid modulates the surface states and electronic structure of the carbon dot, creating a more uniform and stable surface chemical environment, reducing stray surface states and non-radiative recombination defects, thereby strengthening the single dominant emission channel. This results in carbon dots with stable emission peak positions, prominent excitation-independent characteristics, and good fluorescence repeatability. Furthermore, this invention utilizes process optimization measures such as water bath heating assisted by ultrasound, the addition of acetonitrile, and rapid pre-freezing followed by freeze-drying to further improve the homogeneity of precursor mixing and the consistency of product surface states, facilitating the stable acquisition of high-performance excitation-independent carbon dots. When applied to the detection of tetracycline hydrochloride, this carbon dot effectively avoids detection errors caused by emission peak drift in traditional excitation-independent carbon dots, improving the stability, repeatability, and quantitative analysis accuracy of the detection signal. It is more suitable for constructing a rapid, sensitive, convenient, and low-cost fluorescence detection system, thus possessing good practical value and application prospects.

[0036] The above description is merely a preferred embodiment of this application and is not intended to limit this application. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the scope of protection of this application.

Claims

1. A non-excitation-dependent carbon dot, characterized in that, The non-excitation-dependent carbon dots are prepared by a solvothermal method using pyrogallol, thiourea, and o-phenylenediamine as raw materials.

2. A method for preparing non-excitation-dependent carbon dots, characterized in that: Includes the following steps: Step 1: Place pyrogallol, thiourea and o-phenylenediamine in a reaction vessel, add DMF solvent, and mix to obtain a precursor mixture; Step 2: Perform ultrasonic treatment on the precursor mixture; Step 3: Place the sealed reactor in an oven to carry out the solvothermal reaction; Step 4: After the reaction is complete, cool to room temperature; Step 5: Centrifuge to remove solid impurities; Step 6: Filter the supernatant to obtain the carbon dot solution; Step 7: Dry the carbon dot solution to obtain carbon dot powder.

3. The method for preparing non-excitation-dependent carbon dots as described in claim 2, characterized in that: In step 1, the inner liner of the reactor is made of polytetrafluoroethylene.

4. The method for preparing non-excitation-dependent carbon dots as described in claim 2, characterized in that: In step 3, the oven temperature is 180℃ and the time is 6 hours.

5. The method for preparing non-excitation-dependent carbon dots as described in claim 2, characterized in that: In step 6, the filter head has a pore size of 0.22 μm.

6. The method for preparing non-excitation-dependent carbon dots as described in claim 2, characterized in that: In step 2, the ultrasound is performed using a water bath for heating.

7. The method for preparing non-excitation-dependent carbon dots as described in claim 6, characterized in that: In step 1, acetonitrile is added to the DMF solvent.

8. The method for preparing non-excitation-dependent carbon dots as described in claim 2, characterized in that: In step 7, the carbon dot solution is first rapidly pre-frozen, and then freeze-dried after the sample is fully frozen.

9. The application of the non-excitation-dependent carbon dot as described in claim 1, characterized in that: The non-excitation-dependent carbon dots are used for tetracycline hydrochloride detection.

10. The application of the non-excitation-dependent carbon dot as described in claim 9, characterized in that: The tetracycline hydrochloride sample to be tested was mixed with a diluted non-excitation-dependent carbon dot solution, and the fluorescence intensity at 570 nm was detected at an excitation wavelength of 470 nm. The concentration of tetracycline hydrochloride in the sample to be tested was determined based on the correspondence between the fluorescence intensity and the concentration of tetracycline hydrochloride.

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