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. 2026 Jan 25;16:6199. doi: 10.1038/s41598-026-36947-8

Effectively detect doxycycline in variable sample fields through simple fluorescence strategy using nitrogen doped carbon dots as probe

Linlin Xu 1,#, Xiaoyan Lin 3,#, Ling Li 3, Rouying Cai 2,3, Ying Zheng 3, Jianyong Huang 3, Shaohuang Weng 3,✉, Chen Zheng 2,✉
PMCID: PMC12905241  PMID: 41582204

Abstract

The convenient and widespread implementation of doxycycline (DOXY) detection still requires the development of new methods to enable rapid detection across various environments. In this study, blue-emitting nitrogen-doped carbon dots (N-CDs) were successfully prepared using glutamine and anhydrous citric acid as primary materials. Correspondingly, a simple and sensitive fluorescence method for detecting DOXY was developed using N-CDs as probe. The fluorescence intensities of N-CDs were significantly quenched by variable concentrations of DOXY based on the internal filtration effect, which was verified through quantum efficiency correction experiment. Under the optimized testing conditions, the concentrations of DOXY showed a good linear relationship with fluorescence quenching efficiencies in the range of 0.1-35 μg/mL, with a detection limit (LOD) of 0.018 μg/mL, suggesting the good analytical performance. This method was successfully applied for the determination of DOXY in human plasma, dairy products and tap water samples, demonstrating satisfactory recoveries and reproducibility. The results highlight the promising potential of this approach for clinical monitoring of blood drug concentrations, detection of drug residues, and environmental sample analysis.

Supplementary Information

The online version contains supplementary material available at 10.1038/s41598-026-36947-8.

Keywords: Fluorescent strategy, Carbon dots, Sensitive detection, Doxycycline, Diverse sample matrices

Subject terms: Biological techniques, Chemistry

Introduction

Doxycycline (DOXY) is a broad-spectrum antibiotic with potent antibacterial activity against common bacteria such as Staphylococcus aureus, Streptococcus pneumoniae, and Escherichia coli1. Additionally, it exhibits inhibitory effects on typical pathogens including mycoplasma, chlamydia, rickettsia, and spirochetes, particularly in the treatment of drug-resistant Mycoplasma infections2. DOXY targets the 30S ribosomal subunit of Mycoplasma, inhibiting protein synthesis and prolonging peptide chains, which suppresses the growth and replication of drug-resistant strains, alleviates symptoms, shortens disease duration, and improves clinical outcomes2,3. However, DOXY may cause permanent tooth yellowing and poor enamel development4. For children, especially those under the age of 8, the risks and benefits must be meticulously balanced. Therefore, immediate monitoring of DOXY blood concentration is essential to ensure safe and effective use. Moreover, due to its high lipophilicity and permeability, DOXY is particularly effective in preventing bacterial infections and promoting increased animal food production. Because of its beneficial properties, DOXY has become a prevalent antibiotic choice in the fields of aquaculture and animal husbandry. Compared to other antibiotics, DOXY can penetrate deep tissues and even cross the blood-brain barrier or placental barrier5. While antibiotics are critical in the prevention and treatment of animal diseases, they also contribute to drug residues and the emergence of drug-resistant strains, which pose risks to human health and threaten the sustainable development of animal husbandry. Consequently, the need for convenient and accurate DOXY determination methods is significant for both agricultural production and food safety.

Currently, detection methods for DOXY encompass biosensors6, enzyme-linked immunosorbent assay (ELISA)7, ultraviolet spectrophotometry (UV)8, high-performance liquid chromatography (HPLC)9, capillary electrophoresis10, liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS)11,12, colorimetry13,14, and fiber sensors15. Although multiple methods are available, each has its own advantages and limitations. Most of these methods are plagued by time-consuming and labor-intensive analysis processes, as well as high technical demands on operators. Additionally, few methods offer both clinical drug concentration detection and the ability to quantify drug residues in food or environmental samples. Therefore, there is a pressing need to develop a detection technology that is both user-friendly and accurate, which will guide DOXY administration protocols for treating clinical mycoplasma infections and precision feeding systems in animal farms.

Fluorescence-based strategies have garnered attention ascribed to their sensitivity, simple operation, and minimal sample pretreatment requirements16–19. Several probes, such as sulfur quantum dots20, copper nanoclusters21, Eu(III)-functionalized metal-organic framework22, and bimetallic Ag/Au nanoclusters functionalized ZIF-8 framework23, have been established to detect DOXY through fluorescence approach. While, designing an economical fluorescent probe to achieve convenient and applicable fluorescence detection of DOXY in multiple sample systems remains a core requirement. Carbon dots (CDs) are a type of fluorescent carbon nanomaterials with a diameter less than 10 nm24, possessing water solubility, biodegradable with low toxicity, and stability, have received increasing interests in bioimaging, drug delivery, biosensing, electrochemical biosensing, and inflammatory treatment24–27. Additionally, the controllable and tunable fluorescence properties of CDs, which can be modulated by preparation methods and precursor materials, make them highly promising for drug analysis, as changes in their fluorescence intensity or state can be induced by external stimuli19,28–31. It is noteworthy that many CDs-based fluorescence sensing platforms have been applied for the detection of DOXY5,32–34, but so far there was limited methods that can simultaneously meet the monitoring of clinical drug therapy and the detection of animal products and environmental samples through simple operation procedures. Correspondingly, inspired by the advantages of CDs and the importance of developing widespread method for DOXY, in this work, a specific method was developed to prepare a kind of CDs with excellent luminescent properties and target properties based on the direct interaction between DOXY and CDs considering the absorbance characteristics of drugs and the adjustable surface properties of CDs. This kind of CDs (named as N-CDs herein) was prepared from anhydrous citric acid and glutamine through hydrothermal method. Then, a fluorescence method based on the internal filtering effect (IFE) between DOXY and N-CDs was fabricated for DOXY detection through simple operation, as shown in Fig. 1. It was successfully applied to the spiked recovery of DOXY in human plasma, dairy products, and tap water, all of which met the recovery standards. Furthermore, this strategy exhibited high reproducibility and reliability, demonstrating its valuable tool in the ongoing efforts to monitor and assess drug presence in diverse environments.

Fig. 1.

Fig. 1

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The synthesis of N-CDs and the fluorescence strategy of doxycycline detection with N-CDs as the probe.

Experimental section

Reagents and apparatus

Doxycycline (DOXY) was purchased from Aladdin Chemistry Co., Ltd. (Shanghai, China). Sodium hydroxide (NaOH), sodium chloride (NaCl), calcium chloride (CaCl2), sodium dihydrogen phosphate dihydrate (NaH2PO4∙2H2O), disodium hydrogen phosphate dodecahydrate (Na2HPO4∙12H2O), potassium chloride (KCl), L-glucose (L-Glu), fructose (Fru), sucrose (Suc), L-ascorbic acid (AA), L-cysteine (L-Cys), L-arginine (L-Arg), and lactose (LAC) were obtained from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). L-glutathione (GSH) was acquired from Beijing Bailingwei Technology Co., Ltd. (Beijing, China). Amoxicillin, ampicillin, bleomycin, gentamicin, cefixime, azithromycin, and doxorubicin were obtained from local hospital. Anhydrous citric acid and glutamine were purchased from Shanghai Aladdin Biochemical Technology Co., Ltd. (Shanghai, China). All reagents were used directly without further purification.

Transmission electron microscopy (TEM) images were performed on a FEI Talos F200S (Thermo Fisher Scientific, USA). Fluorescence spectra were recorded on a Cary Eclipse fluorescence spectrophotometer (Agilent Technologies, USA). Decay curves were measured on a FS5 fluorescence spectrometer (Edinburgh Instruments, Britain). Ultraviolet visible (UV–vis) absorption spectra were measured on a UV-2450 spectrophotometer (Shimadzu Corporation, Japan). Fourier Transform Infrared Spectroscopy (FTIR) was recorded with a NICOLET iS50 Infrared Spectroscopy (Thermo Fisher Scientific, USA). X-ray photoelectron spectra (XPS) spectra were obtained using PHI VersaProbe 4 scanning XPS (ULVAC-PHI, Japan).

Synthesis of N-CDs

Briefly, 1.0 g glutamine and 1.5 g anhydrous citric acid were dissolved in 18 mL ultrapure water. Then, the mixture was heated at 180 ℃ for continuous 24 h via a hydrothermal strategy. Then, the reaction solution was cooled for subsequent 48-hour purification through dialysis in a 500-1000 Da dialysis bag. Fresh water was replaced every two hours in the purified process. After the freeze-drying of dialyzed solution, N-CDs were weighed and dissolved in water to be 1.6 mg/mL. When not use, the solution of N-CDs was stored at 4 ℃.

Detecting DOXY using N-CDs as probe

The 50 μL of N-CDs (1.6 mg/mL) was individually incubated with standard DOXY from 0 to 120.0 μg/mL in 200 μL phosphate buffered saline (PBS, pH 6.5). The mixed solution was kept at 30 ℃ for 20 min. Then the fluorescence spectra were recorded in the 345-500 nm with 335 nm excitation. The quenching efficiency (QE) of (F0-F)/F was applied to quantify the DOXY. F and F0 were the fluorescence intensities of N-CDs with and without the presence of DOXY.

The specificity of this method was also investigated. The 20 μL of 300 μg/mL tetracycline, tigecycline, L-glutamic acid, L-cysteine, L-arginine, L-ascorbic acid, FeSO4, CaCl2, NaCl, sucrose, glucose, lactose, cefoperazone sodium and sulbactam sodium, amoxicillin, glutathione, polymyxin B, some common antibiotics, and 30 μg/mL DOXY were mixed with 50 μL N-CDs (1.6 mg/mL) for fluorescence assay, respectively. In addition, the above-mentioned interferences except tetracycline, tigecycline, and antibiotics were respectively added to the solution containing 50 μL N-CDs (1.6 mg/mL) and 30 μg/mL DOXY to compare the fluorescence response for evaluating the anti-interfering property of this strategy.

Detecting DOXY in real samples

Approved by the Ethics Committee of Fujian Medical Union Hospital (No. 2023KY129, Fuzhou, China), the human blood samples obtained from volunteers with informed consent were utilized in this work. All blood samples were obtained and carried out in accordance with relevant guidelines and regulations by the ethics requirements. The spiked standard recoveries of DOXY were evaluated in human plasma, milk, formula, and tap water, respectively. To perform this evaluation, in a total 200 μL volume, using pH 6.5 PBS as a diluent, and different concentrations (18.0, 24.0, 30.0 μg/mL) of DOXY were individually spiked into 10% human plasma from healthy volunteers with informed consent, 10% milk (SC10562012300296), 20 mg/mL formula (SC10523062450028) and tap water (taken from Fujian Medical University Qishan Campus). The variable fluorescence intensities were measured using the testing procedure of the standard DOXY, which were subsequently utilized to assess the recovery of DOXY in actual samples.

Results and discussion

Characterization of N-CDs

TEM (Fig. 2A) revealed that the prepared N-CDs were regular quasi-spherical structures with good uniformity and dispersibility. The measured diameters of N-CDs distributed in the range of 2.03-7.71 nm with the average size of 4.73 nm. The high-resolution TEM (Fig. 2A) demonstrated that N-CDs had the appearance of a prominent lattice fringe like distinct graphene characterized by the lattice spacing of 0.21 nm35. The relative quantum yield of N-CDs (QYN-CDs) was quantified as 16.5% (Fig. 2B) using quinine sulfate (QYQS = 54.6%) as a standard reference, indicating that the N-CDs possessed an acceptable level of luminescent ability.

Fig. 2.

Fig. 2

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Characterization of N-CDs. (A) The TEM, HRTEM (upper right) and particle size distribution chart (lower right) of N-CDs. (B) The relative quantum yield of N-CDs using quinine in 0.1 M H2SO4 as reference. (C) The fluorescence emission spectra of N-CDs under excitation at 290-360 nm. (D) The UV-visible absorption spectra and fluorescence spectra of N-CDs.

The optical characteristics of N-CDs were further studied. The maximum peaks of N-CDs exhibited a slight redshift from 389 nm to 409 nm under excitation from 290 nm to 360 nm (Fig. 2C). This result displayed a similar excitation-dependent luminescence behavior to that of other reported CDs29,36–38, which may be attributed to the certain degree of the variable sizes of N-CDs (Fig. 2A)38–41. Correspondingly, the maximum excitation and emission wavelengths of N-CDs were 335 nm and 396 nm (Fig. 2D), respectively. Furthermore, the UV-vis absorbance peaks of N-CDs were 345 nm and 270 nm (Fig. 2D), which were attributed to the n-π* transition of C=O and the π-π* transition of C=C, respectively42.

The molecular structure of N-CDs was characterized via XPS and FTIR (Fig. 3). The XPS spectrum (Fig. 3A) exhibited the characteristic peaks of C1s, N1s, and O1s at 284.49 eV, 399.49 eV, and 531.60 eV, respectively. The corresponding atomic percentages were 68.73% (C), 7.12% (N), and 24.15% (O). The high-resolution C1s XPS spectra (Fig. 3B) illustrated two fitting bands of C-C/C-H (284.8 eV) and C=O/O-C=O (288.7 eV)43. The high-resolution N1s XPS spectra (Fig. 3C) could be deconvoluted into the binding energies of 399.5 eV (pyrrolic N/graphitic N), 401.2 eV (-NH2) and 398.2 eV (pyridinic N/-CONH2)43–46. In addition, the two deconvoluted bands of O1s (Fig. 3D) at 531.5 eV (C=O/O-H) and 533.0 eV (C-O/O-C=O) were found in the high-resolution O1s XPS spectra47. XPS results confirmed the successful doping of N. As shown in the FTIR spectrum of N-CDs (Fig. 3E), the absorption peak between 3750 and 3000 cm-1 was ascribed to the overlapping stretching vibration of -OH and -NH248,49. The peak at 2905 cm-1 indicated the C-H stretching vibration of alkyl groups49. The peak at 1661 cm−1 was associated with the stretching vibration of C=O in the amide structure49. The peak at 1580 cm-1 was attributed to the aromatic C-H bending vibration and the peak at 1403 cm-1 was related to C-H bending vibration49. Furthermore, FTIR was also used to analyze and compare the functional groups of citric acid and glutamine as the raw materials for preparing N-CDs (Fig. 3F). For nitrogen source of glutamine, the characteristic absorption peaks of glutamine included -NH2 stretching vibration at 3664 cm-1, C-N stretching vibration at 2118 cm-1 and C-O stretching vibration at 1935 cm-1. For carbon source of citric acid, its characteristic absorption peaks included -OH stretching vibration at 3496 cm-1, saturated C-H stretching vibration at 2805 cm-1, C-O stretching vibration at 2097 cm-1, C=O stretching vibration at 1727 cm-1 and C-C stretching vibration at 1560 cm-1. The wide absorption band of N-CDs in the range of 3000-3750 cm-1 can be attributed to the binding peak formed by the overlap of the N-H stretching vibration peak and the -OH stretching vibration peak, and its intensity was higher than that of citric acid and glutamine, indicating that N-CDs were rich in amino and hydroxyl groups. From the above analysis, it could be concluded that simple hydrothermal treatment can successfully connect functional groups such as amino and hydroxyl groups from the raw materials to the surface of N-CDs. To sum up, combined with FTIR and XPS results, it was confirmed that N-CDs prepared by one-step hydrothermal method were successfully doped by N with the presence of -OH, -COOH and amide bonds to enhance the inherent dispersibility and stability of N-CDs in water.

Fig. 3.

Fig. 3

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The molecular characterization of N-CDs. The XPS survey (A), high-resolution XPS survey of C1s (B), N1s (C) and O1s (D). The FTIR spectra of N-CDs (E), and the glutamine and citric acid (F).

Moreover, the optical stability of the N-CDs was assessed under various environmental conditions (Fig. S1). In the pH range of 3-12, the fluorescence intensity remained above 96% (Fig. S1A), indicating good pH stability. Additionally, the fluorescence intensity was stable under varying salt concentrations (0-1000 mM, Fig. S1B) and temperatures (5-90 ℃, Fig. S1C), with retention rates of above 95% and 98%, respectively. Under continuous excitation at 335 nm for 1 h (Fig. S1D) and storage at 4 ℃ for more than one month (Fig. S2), the fluorescence intensity of N-CDs remained stable, highlighting the excellent optical stability of N-CDs under various conditions.

The feasibility and mechanism of DOXY detection using N-CDs as probe

The effect of DOXY on the fluorescence intensity of N-CDs was thoroughly investigated. As illustrated in Fig. 4A, the addition of DOXY resulted in a noticeable quenching of the fluorescence emitted by the N-CDs. Furthermore, this quenching effect strengthened gradually with the increasing concentration of DOXY. This observation suggested that it was feasible to develop a reliable analytical method that utilized the reduction in fluorescence signals of N-CDs to quantify DOXY.

Fig. 4.

Fig. 4

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The feasibility and quenching mechanism of N-CDs by DOXY. (A) The effect of different concentrations (0, 5, 30, 60 μg/mL) of DOXY on the emission spectra of N-CDs. (B) Fluorescence lifetime of N-CDs and N-CDs+DOXY. (C) UV-vis absorbance spectra of N-CDs, DOXY and N-CDs+DOXY. (D) UV-vis absorbance spectrum of DOXY and fluorescence spectra of N-CDs. (E) The parameters of the cuvette with the thickness of the excitation beam (s = 0.10 cm), the distance between the edge of the excitation beam and the edge of cuvette (g = 0.40 cm), and the width (d = 1.00 cm). (F) The observed QE (black) and the corrected QE (red) of the IFE in N-CDs+DOXY with different concentrations of DOXY. The concentrations of N-CDs and DOXY in (B-D) were 400 μg/mL and 30 μg/mL, respectively.

The quenching mechanism of DOXY on N-CDs was preliminarily investigated. The fluorescence lifetime (Fig. 4B) of N-CDs remained nearly unchanged by DOXY, suggesting the absence of fluorescence resonance energy transfer (FRET) or dynamic quenching effect (DQE) between N-CDs and DOXY. By analyzing and comparing the UV-vis absorbance spectra of N-CDs, DOXY, and N-CDs+DOXY systems (Fig. 4C), the absorbance peak of N-CDs and DOXY remained unchanged in N-CDs+DOXY with only an increase in absorbance intensity caused by absorption superposition, suggesting that no new absorption peaks were found in N-CDs+DOXY. This result indicated that the interaction of N-CDs and DOXY in N-CDs+DOXY did not form new non-fluorescent substances, indicating that the fluorescence quenching of N-CDs by DOXY was not caused by static quenching effect (SQE)49.

The UV-vis spectrum of DOXY showed the characteristic absorption peaks of 275 nm and 347 nm (Fig. 4D). Generally, the adequate overlap of the absorption band of an absorber with the excitation and/or emission bands of a fluorophore may cause inner filter effect (IFE)29,49. Considering the maximum excitation wavelength of N-CDs at 335 nm, the QE corrections were made by the consideration of the dish geometry (Fig. 4E) and Equation S1 (supporting information)50. The absorbance and fluorescence intensities of N-CDs (Table S1) were achieved with the introduction of variable concentrations of DOXY. And the corrected fluorescent intensity (F) from Equation S1 differed greatly at each concentration of DOXY, which were significantly different from the observed QEs (Fig. 4F), indicating the occurrence of IFE of DOXY on N-CDs.

Optimization of operation conditions

The variable conditions of fluorescent assay may induce the variable degrees of molecular motion and molecular interaction of the testing system51,52. Thus, the experimental conditions of the preliminary established methods were needed to be optimized for evaluating the testing performance, as shown in Fig. S3. As depicted in Fig. S3A, the fluorescence intensity of N-CDs and their fluorescence response to DOXY were highest in PBS compared to Tris-HCl and HAc-NaAc buffers. Therefore, PBS was selected as the optimal buffer for DOXY detection. The effect of pH on the QE of DOXY was also investigated (Fig. S3B). Initially, as the pH increased, the QEs of DOXY to N-CDs decreased, then gradually increased, followed by a subsequent decrease. Notably, the highest QE was observed at pH 6.5, which was selected as the optimal pH for further experiments. In addition, the reaction temperature and time were optimized. Fig. S3C showed that the optimal QE of DOXY to N-CDs was attained at 30 ℃. Regarding the reaction time, as shown in Fig. S3D, the fluorescence QEs changed with increased times, and exhibited the maximum response at 20 min. Thus, the optimal experimental conditions for this strategy were determined to be 30 ℃, using PBS buffer at pH 6.5 for a reaction time of 20 min.

Analytical performance

Considering the actual monitoring of DOXY in clinical blood samples and aquaculture environment samples, the selectivity of this procedure was initially assessed. Consequently, the possible effects of these common antibacterial drugs, common metal ions, and physiological substances were first investigated. As shown in Fig. 5A, although other tetracycline drugs played a certain impact on QEs like the effect of DOXY in this method, other common substances exhibited little QEs on N-CDs. In general, tetracycline drugs are not used simultaneously, indicating that this method has acceptable specificity. It has been reported that metal ions, especially for Ca2+, can form complexes with DOXY34. While the introduction of several metal ions played ignorable response of the detection of DOXY using N-CDs as probe (Fig. 5A), suggesting the selectivity and specificity of this method for DOXY from metal ions. The UV-vis spectrum of the complex from DOXY and metal ions like Ca2+ exhibited a wide absorbance peak but the kept absorbance degree at around 350 nm, which was like the absorbance degree of free DOXY34, suggesting that the IFE of the testing system was unaffected by introducing metal ions. Furthermore, when other substances were added to the system for detecting DOXY (Fig. 5B), it was found that the QEs remained stable regardless of the addition of interfering substances, also implying the anti-interference capacity of this strategy for DOXY detection. In addition, considering the potential coexistence of other common antibiotics including amoxicillin, ampicillin, and so on, the responses of N-CDs to DOXY and other antibiotics were compared, as shown in Fig. S4. Compared with the obvious response of N-CDs to DOXY, other antibiotics exhibited negligible QEs on N-CDs, further suggesting the satisfactory specificity of this method.

Fig. 5.

Fig. 5

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The detecting performance of DOXY using N-CDs as probe. (A) The QE of N-CDs in the presence of DOXY or various substances, respectively. (B) The QE of N-CDs+ DOXY in the presence of various substances. (C) The fluorescence emission spectra of N-CDs with variable concentrations of DOXY in PBS (pH 6.5). (D) The linear relationship between QE of N-CDs and DOXY in the range of 0.1-35 μg/mL in PBS (The error bar represented the standard deviation of three parallel determinations).

The fluorescence emission spectra of N-CDs treated with varying concentrations of DOXY were recorded under the optimal conditions. With the increased concentration of DOXY (CDOXY), the fluorescence intensities of N-CDs gradually decreased (Fig. 5C), and the QEs gradually rose and tended towards stability (inset of Fig. 5D). Furthermore, the QEs ((F0-F)/F0) were fitted well with variable CDOXY in the range of 0.1-35 μg/mL with the linear equation of y = 0.0105 CDOXY + 0.0526 (R2 = 0.9939) (Fig. 5D). Based on the 3σ/S (σ and S are the standard deviation of 10 parallel blank measurements and the slope of the linear fitting equation, respectively), the LOD was 0.018 μg/mL. The testing repeatability of this strategy was also evaluated. The 30 individual samples of 30 μg/mL DOXY were applied to test the repeatability. Six samples were measured daily for 5 consecutive days, as shown in Fig. S5. The response values were highly consistent in the 30 tests. The relative standard deviation (RSD) of the parallel operation was 1.63%. The low RSD suggested the high reproducibility and reliability of the measurement. At the same time, by comparing with other fluorescence methods for detecting DOXY (Table S2), the linear range and LOD of this strategy exhibited the comparable and even superior performance. Moreover, this developed method possessed the characteristic of facile operation.

Detection of DOXY in actual samples

Considering the practical applications of DOXY in clinical pharmacy and animal husbandry, different actual samples were selected to assess the reliability and applicability of the established approach. Firstly, the determination of DOXY in human plasma samples was conducted (Table 1). The spiked recoveries of 18, 24, 30 μg/mL of DOXY in human plasma samples ranged from 90.29% to 107.13%, with RSD ranging from 0.18% to 0.55%. The comparative results of added DOXY in human plasma were measured using standard HPLC. The recovery rates of 18, 24, 30 μg/mL of DOXY in human plasma were 92.12%, 91.92% and 94.37% using HPLC, respectively. These results validated the potential of this method for drug monitoring in clinical blood samples.

Table 1.

Results of DOXY in human plasma samples using this method (n = 3).

Samples Spiked (μg/mL) Found (μg/mL) Recovery (%) RSD (n = 3, %)
Human plasma 1 18 18.31 101.71 0.23
24 24.37 101.55 0.18
30 31.61 105.38 0.42
Human plasma 2 18 19.28 107.13 0.18
24 24.23 100.96 0.34
30 31.63 105.44 0.15
Human plasma 3 18 18.56 103.09 0.55
24 24.57 102.36 0.21
30 27.08 90.290 0.45
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Considering that multiple tetracycline drugs may be used in food and environment conditions, combined with the similar absorption properties of tetracycline drugs, DOXY was used as the representative tetracycline to assess the practical application of this method to evaluate the feasibility of determining tetracycline drugs in food and environmental samples. As shown in Table S3, the spiked recoveries of DOXY at different concentrations in milk samples were 106.61%, 100.89%, and 98.61%, respectively, and the RSD did not exceed 1%. Correspondingly, the spiked recoveries of DOXY in milk powder (Table S4) were 108.10%, 103.38%, and 106.93%, respectively, with the RSD less than 1%. Hence, the spiked recoveries and RSD results of DOXY in dairy products confirmed favorable detection accuracy of this method in dairy products, especially for milk and formula. Furthermore, the spiked recoveries of DOXY at different concentrations in tap water (Table S5) ranged from 94.24% to 102.37% with the RSD less than 2.1%. The comparative result of spiked DOXY in tap water was testing using standard HPLC, the obtained recovery of 108.69% implied the acceptable accuracy of the developed fluorescence method using N-CDs as probe. The above results indicated that the method established in this study had considerable potential for qualitative and quantitative analysis of doxycycline and even the total tetracycline drugs in environmental samples.

Conclusion

In summary, N-CDs were successfully prepared via a simple, and efficient one-step hydrothermal method using glutamine and citric acid as sources. N-CDs featured the functional groups of hydroxyls (-OH) and amino (-NH2), which exhibited excellent dispersibility and stability in water, with a fluorescence quantum yield of 16.50%. Based on inner filter effect (IFE), a fast, convenient, and accurate detection platform for DOXY was developed using N-CDs as probe with good analytical performance of a detection limit of 0.018 μg/mL. By examining the reproducibility and spiked recoveries of DOXY in human plasma, dairy products, and tap water systems, this method demonstrated high reproducibility and reliability, and exhibited significant potential application value in the detection of DOXY in clinical samples, drug residue control, and environmental samples.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary Material 1 (505.9KB, docx)

Acknowledgements

This work was supported by the Joint Funds for the Innovation of Science and Technology, Fujian Province (2021Y9007), and the National Science Foundation of Fujian Province (2023Y0021).

Author contributions

L. X., X. L. and L. L. wrote the main manuscript text, R. C. and Y. Z. discussed the manuscript, J. H. prepared figures 1,S. W. and C. Z. supervised and reviewed the manuscript. All authors reviewed the manuscript.

Data availability

The datasets used and/or analyzed during the current study available from the corresponding author on reasonable request.

Declarations

Competing interests

The authors declare no competing interests.

Footnotes

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

These authors contributed equally: Linlin Xu and Xiaoyan Lin.

Contributor Information

Shaohuang Weng, Email: shweng@fjmu.edu.cn.

Chen Zheng, Email: 20903155@qq.com.

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Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

Supplementary Material 1 (505.9KB, docx)

Data Availability Statement

The datasets used and/or analyzed during the current study available from the corresponding author on reasonable request.

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