Development of an eco-friendly fluorescent probe for mefenamic acid sensing in pharmaceuticals and biofluids.

Mefenamic acid, renowned for its analgesic properties, stands as a reliable choice for alleviating mild to moderate pain. However, its versatility extends beyond pain relief, with ongoing research unveiling its promising therapeutic potential across diverse domains. A straightforward, environmentally friendly, and sensitive spectrofluorometric technique has been developed for the precise quantification of the analgesic medication, mefenamic acid. This method relies on the immediate reduction of fluorescence emitted by a probe upon interaction with varying concentrations of the drug. The fluorescent probe utilized, N-phenyl-1-naphthylamine (NPNA), was synthesized in a single step, and the fluorescence intensities were measured at 480 nm using synchronous fluorescence spectroscopy with a wavelength difference of 200 nm. Temperature variations and lifetime studies indicated that the quenching process was static. The calibration curve exhibited linearity within the concentration range of 0.50-9.00 µg/mL, with a detection limit of 60.00 ng/mL. Various experimental parameters affecting the quenching process were meticulously examined and optimized. The proposed technique was successfully applied to determine mefenamic acid in pharmaceutical formulations, plasma, and urine, yielding excellent recoveries ranging from 98% to 100.5%. The greenness of the developed method was evaluated using three metrics: the Analytical Eco-scale, AGREE, and the Green Analytical Procedure Index.

In Situ Visualizing Carboxylesterase Activity in Type 2 Diabetes Mellitus Using an Activatable Endoplasmic Reticulum Targetable Proximity Labeling Far-Red Fluorescent Probe.

Carboxylesterase (CE), an enzyme widely present in organisms, is involved in various physiological and pathological processes. Changes in the levels of CEs in the liver may predict the presence of type 2 diabetes mellitus (T2DM). Here, a novel dicyanoisophorone (DCI)-based proximity-labeled far-red fluorescent probe DCI2F-Ac with endoplasmic reticulum targeting was proposed for real-time monitoring and imaging of the CEs activity. DCI2F-Ac featured very low cytotoxicity and biotoxicity and was highly selective and sensitive for CEs. Compared with traditional CEs probes, DCI2F-Ac was covalently anchored directly to CEs, thus effectively reducing the loss of in situ fluorescent signals due to diffusion. Through the "on-off" fluorescence signal readout, DCI2F-Ac was able to distinguish cell lines and screen for CEs inhibitors. In terms of endoplasmic reticulum (ER) stress, it was found that thapsigargin (Tg) induced upregulation of CEs levels but not tunicamycin (Tm), which was related to the calcium homeostasis of the ER. DCI2F-Ac could efficiently detect downregulated CEs in the livers of T2DM, and the therapeutic efficacy of metformin, acarbose, and a combination of these two drugs was assessed by tracking the fluctuation of CEs levels. The results showed that combining metformin and acarbose could restore CEs levels to near-normal levels with the best antidiabetic effect. Thus, the DCI2F-Ac probe provides a great opportunity to explore the untapped potential of CEs in liver metabolic disorders and drug efficacy assessment.

Red-emitting 4-methyl coumarin fused barbituric acid as an electrochemical sensor for catechol detection and probe for latent fingerprints.

Herein, we have reported a red-emitting 4-methyl coumarin fused barbituric acid azo dye (4-MCBA) synthesized by conventional method. Density functional theory (DFT) studies of tautomer compounds were done using (B3LYP) with a basis set of 6-31G(d,p). NLO analysis has shown that tautomer has mean first-order hyperpolarisabilities (ß) value of 1.8188 × 10-30 esu and 1.0470 × 10-30 esu for azo and hydrazone forms, respectively, which is approximately nine and five times greater than the magnitude of urea. 4-MCBA exhibited two absorption peaks in the range of 290-317 and 379-394 nm, and emission spectra were observed at 536 nm. CV study demonstrated that the modified 4-MCBA/MGC electrode exhibited excellent electrochemical sensitivity towards the detection of catechol and the detection limit is 9.39 µM under optimum conditions. The 4-MCBA employed as a fluorescent probe for the visualisation of LFPs on various surfaces exhibited Level-I to level-II LFPs, with low background interference.

Fluorescence lifetime multiplexing with fluorogen activating protein FAST variants.

In this paper, we propose a fluorescence-lifetime imaging microscopy (FLIM) multiplexing system based on the fluorogen-activating protein FAST. This genetically encoded fluorescent labeling platform employs FAST mutants that activate the same fluorogen but provide different fluorescence lifetimes for each specific protein-dye pair. All the proposed probes with varying lifetimes possess nearly identical and the smallest-in-class size, along with quite similar steady-state optical properties. In live mammalian cells, we target these chemogenetic tags to two intracellular structures simultaneously, where their fluorescence signals are clearly distinguished by FLIM. Due to the unique structure of certain fluorogens under study, their complexes with FAST mutants display a monophasic fluorescence decay, which may facilitate enhanced multiplexing efficiency by reducing signal cross-talks and providing optimal prerequisites for signal separation upon co-localized and/or spatially overlapped labeling.

Biomolecule-regulation of fluorescent probe signaling: Homogeneous rapid portable protease sensing in serum.

BACKGROUND: As is well documented, prostate cancer (PCa) being the second most prevalent cancer in men worldwide, emphasizing the importance of early diagnosis for prognosis. However, conventional prostate-specific antigen (PSA) testing lacks sufficient diagnostic efficiency due to its relatively low sensitivity and limited detection range. Mounting evidence suggests that matrix metalloproteinase 9 (MMP-9) expression increases with the aggressive behavior of PCa, highlighting the significance of detecting the serum level of MMP-9 in patients. Developing a non-immune rapid, portable MMP-9 detection strategy and investigating its representativeness of PCa serum markers hold considerable implications. RESULTS: Herein, our study developed a simple, homogeneous dual fluorescence and smartphone-assisted red-green-blue (RGB) visualization peptide sensor of MMP-9, utilizing cadmium telluride quantum dots (CdTe QDs) and calcein as signal reporters. The essence of our approach revolves around the proteolytic ability of MMP-9, exploiting the selective recognition of molecule-Cu2+ complexes with different molecular weights by CdTe QDs and calcein. Under optimized conditions, the limits of detection (LODs) for MMP-9 were 0.5 pg/mL and 6 pg/mL using fluorescence and RGB values readouts, respectively. Indeed, this strategy exhibited robust specificity and anti-interference ability. MMP-9 was quantified in 42 clinical serum samples via dual-fluorescence analysis, with 12 samples being visually identified with a smartphone. According to receiver operating characteristic curve (ROC) analysis, its sensitivity and specificity were 90 % and 100 %, respectively, with an area under curve (AUC) value of 0.903. SIGNIFICANCE AND NOVELTY: Of note, the results of the aforementioned analysis were highly consistent with the serum level of PSA, clinical color Doppler flow imaging (CDFI), and histopathological results. Therefore, this simple, rapid, homogeneous fluorescence and visualization strategy can reliably measure MMP-9 levels and exhibit promising potential in point-of-care testing (POCT) applications for PCa patients.

Research progress of fluorescent composites based on cyclodextrins: Preparation strategies, fluorescence properties and applications in sensing and bioimaging.

Fluorescence analysis has been regarded as one of the commonly used analytical methods because of its advantages of simple operation, fast response, low cost and high sensitivity. So far, various fluorescent probes, with noble metal nanoclusters, quantum dots, organic dyes and metal organic frameworks as representatives, have been widely reported. However, single fluorescent probe often suffers from some deficiencies, such as low quantum yield, poor chemical stability, low water solubility and toxicity. To overcome these disadvantages, the introduction of cyclodextrins into fluorescent probes has become a fascinating approach. This review (with 218 references) systematically covers the research progress of fluorescent composites based on cyclodextrins in recent years. Preparation strategies, fluorescence properties, response mechanisms and applications in sensing (ions, organic pollutants, bio-related molecules, temperature, pH) and bioimaging of fluorescent composites based on cyclodextrins are summarized in detail. Finally, the current challenges and future perspectives of these composites in relative research fields are discussed.

A near-infrared fluorescent probe for differentiating cancer cells from normal cells and early diagnosis of liver cirrhosis.

BACKGROUND: Cirrhosis represents the terminal stage of liver disease progression and timely intervention in a diseased liver can enhance the likelihood of recovery. Viscosity, a crucial parameter of the cellular microenvironment, is intricately linked to the advancement of cirrhosis. However, viscosity monitoring still faces significant challenges in achieving non-invasive and rapid early diagnosis of cirrhosis. Near-infrared (NIR) fluorescence imaging has the advantages of high sensitivity, non-destructive detection, and ignoring background fluorescence interference, plays an important role in diagnosing and treating various biological diseases. Hence, monitoring cellular viscosity changes with NIR fluorescence probe holds great significance in the early diagnosis of cirrhosis. RESULTS: In this study, the NIR fluorescence probe based on the intramolecular charge transfer (TICT) mechanism was developed for imaging applications in mouse model of liver cirrhosis. A molecular rotor-type viscosity-responsive probe was synthesized by linking dioxanthracene groups via carbon-carbon double bonds. The probe demonstrated remarkable sensitivity, high selectivity and photostability, with its responsiveness to viscosity largely unaffected by factors such as polarity, pH, and interfering ions. The probe could effectively detect various drug-induced changes in cellular viscosity, enabling the differentiation between normal cells and cancerous cells. Furthermore, the enhanced tissue penetration capabilities of probe facilitated its successful application in mouse model of liver cirrhosis, allowing for the assessment of liver disease severity based on fluorescence intensity and providing a powerful tool for early diagnosis of cirrhosis. SIGNIFICANCE: A NIR viscosity-sensitive fluorescent probe was specifically designed to effectively monitor alterations in cellular and organ viscosity, which could advance the understanding of the biological characteristics of cancer and provide theoretical support for the early diagnosis of cirrhosis. Overall, this probe held immense potential in monitoring viscosity-related conditions, expanding the range of biomedical tools available.

Tb3+ assisted dithioerythritol stabilized copper nanocluster with AIE behavior for ratiometric fluorescent determination of fluoroquinolones.

BACKGROUND: Fluoroquinolones (FQs) are widely used in livestock and poultry industry because of their satisfactory effects in preventing and treating bacterial infection. However, due to irrational use and poor biodegradability, FQs can easily remain in food animals and further enter the human body through the food chain. Therefore, accurate and sensitive detection of FQs residues in animal-origin food is significant. The traditional methods commonly used for FQs detection have some limitations. Ratiometric fluorescence detection technology has the advantages of fast, sensitive, self-correcting, and easy visualization. However, the reports on the use of ratiometric fluorescence probes for FQs detection are limited. RESULTS: In this work, a novel probe was proposed for ratiometric fluorescent analysis of FQs. In this probe, the fluorescence of dithioerythritol stabilized copper nanoclusters (DTE-Cu NCs) was significantly enhanced due to the Tb3+ triggered aggregation-induced emission effect. FQs bound Tb3+ in Tb3+/DTE-Cu NCs through carboxyl and carbonyl groups, so that Tb3+ was effectively sensitized to emit green fluorescence. However, the red fluorescence of DTE-Cu NCs was not interfered. The fluorescence of the probe transformed from red to green with the increase of FQs concentration. Using norfloxacin (NOR), difloxacin (DIF), and enrofloxacin (ENR) as FQs simulants, this probe showed a sensitive linear response ranged from 0.025 to 22.5 µM, with the limits of detection of 9.6 nM, 9.3 nM, and 7.7 nM. The application potential for FQs detection was verified via a standard addition assay of egg samples with the recovery rate of 90.4 %-114.7 %. SIGNIFICANT: The fluorescence probe based on Tb3+/DTE-Cu NCs is expected to realize the ratiometric fluorescence sensitive detection of FQs. The establishment of this simple, effective, and rapid detection platform opens up a new way for the detection of FQs residues in animal-origin foods, and also provides a new idea for the design of rapid detection platforms for other hazard factors.

A dual-signal triple-readout optical sensing platform for α-glucosidase and ß-glucosidase activity monitoring and inhibitor screening based on luminescent covalent organic framework.

BACKGROUND: As promising biomarkers of diabetes, α-glucosidase (α-Glu) and ß-glucosidase (ß-Glu) play a crucial role in the diagnosis and management of diseases. However, there is a scarcity of techniques available for simultaneously and sensitively detecting both enzymes. What's more, most of the approaches for detecting α-Glu and ß-Glu rely on a single-mode readout, which can be affected by multiple factors leading to inaccurate results. Hence, the simultaneous detection of the activity levels of both enzymes in a single sample utilizing multiple-readout sensing approaches is highly attractive. RESULTS: In this work, we constructed a facile sensing platform for the simultaneous determination of α-Glu and ß-Glu by utilizing a luminescent covalent organic framework (COF) as a fluorescent indicator. The enzymatic hydrolysis product common to both enzymes, p-nitrophenol (PNP), was found to affect the fluorometric signal through an inner filter effect on COF, enhance the colorimetric response by intensifying the absorption peak at 400 nm, and induce changes in RGB values when analyzed using a smartphone-based color recognition application. By combining fluorometric/colorimetric measurements with smartphone-assisted RGB mode, we achieved sensitive and accurate quantification of α-Glu and ß-Glu. The limits of detection for α-Glu were determined to be 0.8, 1.22, and 1.85 U/L, respectively. Similarly, the limits of detection for ß-Glu were 0.16, 0.42, and 0.53 U/L, respectively. SIGNIFICANCE: Application of the proposed sensing platform to clinical serum samples revealed significant differences in the two enzymes between healthy people and diabetic patients. Additionally, the proposed sensing method was successfully applied for the screening of α-Glu inhibitors and ß-Glu inhibitors, demonstrating its viability and prospective applications in the clinical management of diabetes as well as the discovery of antidiabetic medications.

Glutathione-activated biotin-targeted dual-modal imaging probe with improved PDT/PTT synergistic therapy.

BACKGROUND: Glutathione (GSH), a highly abundant thiol compound within cells, plays a critical role in physiological processes and exhibits close correlation with cancer. Among molecular imaging technologies, most probes have relatively short emission wavelengths and lack photoacoustic imaging (PA) capability, resulting in the inability to obtain tissue images with high penetration depth. The presence of GSH in the tumor microenvironment neutralizes ROS, diminishing the therapeutic effect of PDT, thus resulting in often unsatisfactory therapeutic efficacy. Therefore, it is imperative to develop a dual-modal probe for the detection of GSH and the diagnosis and treatment of cancer. RESULTS: In this study, we synthesized a novel dual-modal probe, Cy-Bio-GSH, utilizing near-infrared fluorescence (NIRF) and photoacoustic (PA) imaging techniques for GSH detection. The probe integrates cyanine dye as the fluorophore, nitroazobenzene as the recognition moiety, and biotin as the tumor-targeting moiety. Upon reacting with GSH, the probe emits NIR fluorescence at 820 nm and generates a PA signal. Significantly, this reaction activates the photodynamic and photothermal properties of the probe. By depleting GSH and employing a synergistic photothermal therapy (PTT) treatment, the therapeutic efficacy of photodynamic therapy (PDT) is remarkably enhanced. In-vivo experiments confirm the capability of the probe to detect GSH via NIRF and PA imaging. Notably, the combined tumor-targeting ability and PDT/PTT synergistic therapy enhance therapeutic outcomes for tumors and facilitate their ablation. SIGNIFICANCE: A novel tumor-targeting and dual-modal imaging probe (Cy-Bio-GSH) is synthesized, exhibiting remarkable sensitivity and selectivity to GSH, enabling the visualization of GSH in cells and the differentiation between normal and cancer cells. Cy-Bio-GSH enhances PDT/PTT with effective killing of cancer cells and makes the ablation of tumors in mice. This work represents the first tumor-targeting probe for GSH detection, and provides crucial tool for cancer diagnosis and treatment by dual-modal imaging with improved PDT/PTT synergistic therapy.

A novel mathematical template for developing fDOM probe fluorescence signal correction models for freshwaters.

The reliable application of field deployable fluorescent dissolved organic matter (fDOM) probes is hindered by several influencing factors which need to be compensated. This manuscript describes the corrections of temperature, pH, turbidity and inner filter effect on fluorescence signal of a commercial fDOM probe (fDOMs). For this, Australian waters with wide ranging qualities were selected, e.g. dissolved organic carbon (DOC) ranging from ∼1 to ∼30 mg/L, specific UV absorbance at 254 nm from ∼1 to ∼6 L/m/mg and turbidity from ∼1 to ∼ 350 FNU. Laboratory-based model calibration experiments (MCEs) were performed. A model template was developed and used for the development of the correction models. For each factor, data generated through MCEs were used to determine model coefficient (α) values by fitting the generated model to the experimental data. Four discrete factor models were generated by determination of a factor-specific α value. The α values derived for each water of the MCEs subset were consistent for each factor model. This indicated generic nature of the four α values across wide-ranging water qualities. High correlation between fDOMs and DOC were achieved after applying the four-factor compensation models to new data (r, 0.96, p < 0.05). Also, average biases (and %) between DOC predicted through fDOMs and actual DOC were decreased by applying the four-factor compensation model (from 3.54 (60.9%) to 1.28 (16.7%) mg/L DOC). These correction models were incorporated into a Microsoft EXCEL-based software termed EXOf-Correct for ready-to-use applications.

Different fluorescent labels report distinct components of spHCN channel voltage sensor movement.

We used voltage clamp fluorometry to probe the movement of the S4 helix in the voltage-sensing domain of the sea urchin HCN channel (spHCN) expressed in Xenopus oocytes. We obtained markedly different fluorescence responses with either ALEXA-488 or MTS-TAMRA covalently linked to N-terminal Cys332 of the S4 helix. With hyperpolarizing steps, ALEXA-488 fluorescence increased rapidly, consistent with it reporting the initial inward movement of S4, as previously described. In contrast, MTS-TAMRA fluorescence increased more slowly and its early phase correlated with that of channel opening. Additionally, a slow fluorescence component that tracked the development of the mode shift, or channel hysteresis, could be resolved with both labels. We quantitated this component as an increased deactivation tail current delay with concomitantly longer activation periods and found it to depend strongly on the presence of K+ ions in the pore. Using collisional quenching experiments and structural predictions, we established that ALEXA-488 was more exposed to solvent than MTS-TAMRA. We propose that components of S4 movement during channel activation can be kinetically resolved using different fluorescent probes to reveal distinct biophysical properties. Our findings underscore the need to apply caution when interpreting voltage clamp fluorometry data and demonstrate the potential utility of different labels to interrogate distinct biophysical properties of voltage-gated membrane proteins.

The spectral phasor approach to resolving membrane order with environmentally sensitive dyes.

Hyperspectral imaging is a technique that captures a three-dimensional array of spectral information at each spatial location within a sample, enabling precise characterization and discrimination of biological structures, materials, and chemicals, based on their unique spectral features. Nowadays most commercially available confocal microscopes allow hyperspectral imaging measurements, providing a valuable source of spatially resolved spectroscopic data. Spectral phasor analysis quantitatively and graphically transforms the fluorescence spectra at each pixel of a hyperspectral image into points in a polar plot, offering a visual representation of the spectral characteristics of fluorophores within the sample. Combining the use of environmentally sensitive dyes with phasor analysis of hyperspectral images provides a powerful tool for measuring small changes in lateral membrane heterogeneity. Here, we focus on applications of spectral phasor analysis for the probe LAURDAN on model membranes to resolve packing and hydration. The method is broadly applicable to other dyes and to complex systems such as cell membranes.

Probing the structural evolution on the surface of cardiolipin vesicles with an amphiphilic second harmonic generation and fluorescence probe.

Investigating the influence of the ambient chemical environment on molecular behaviors in liposomes is crucial for understanding and manipulating cellular vitality as well as the capabilities of lipid drug carriers in various environments. Here, we designed and synthesized a second harmonic generation (SHG) and fluorescence probe molecule called Pyr-Py+-N+ (PPN), which possesses membrane-targeting capability. We employed PPN to investigate the response of lipid vesicles composed of cardiolipin to the presence of exogenous salt. The kinetic behaviors, including the adsorption and embedding of PPN on the surface of small unilamellar vesicles (SUVs) composed of cardiolipin, were analyzed. The response of the SUVs to the addition of NaCl was also monitored. A rapid decrease in vesicle size can be evidenced through the rapid drop in SHG emission originating from PPN located on the vesicle surface.

Assessing Seed Germination Response of Parasitic Plant Striga hermonthica with Small-Molecule Probes.

Seed germination of a parasitic plant Striga hermonthica is elicited by strigolactones which are exuded from roots of host plants. Here, we describe a high-throughput germination assay and a method for visualizing in vivo strigolactone receptor functions with a fluorogenic probe.

Confocal Imaging of Seeds.

In flowering plants, proper seed development is achieved through the constant interplay of fertilization products, embryo and endosperm, and maternal tissues. Understanding such a complex biological process requires microscopy techniques able to unveil the seed internal morphological structure. Seed thickness and relatively low permeability make conventional tissue staining techniques impractical unless combined with time-consuming dissecting methods. Here, we describe two techniques to imaging the three-dimensional structure of Arabidopsis seeds by confocal laser scanning microscopy. Both procedures, while differing in their time of execution and resolution, are based on cell wall staining of seed tissues with fluorescent dyes.

Measurement of Intracellular Ca2+ for Studying GPCR-Mediated Lipid Signaling.

Intracellular Ca2+ can be conveniently monitored by sensitive Ca2+ fluorescent dyes in live cells. The Gαq involved lipid signaling pathways and, thus, can be studied by intracellular Ca2+ imaging. Here we describe the protocols to measure intracellular Ca2+ for studying PEG2-EP1 activity in esophageal smooth muscle cells. The ratiometric Fura-2 imaging provides quantitative data, and the Fluo-4 confocal microscopic imaging has high-spatial resolution.

A smart hypochlorous acid fluorescent probe enabling Ibuprofen-release for osteoarthritis theranostics.

Background: Osteoarthritis (OA) standing as the most prevalent form of arthritis, closely associates with heightened levels of reactive oxygen species, particularly hypochlorous acid (HOCl). Although there are numerous probes available for detecting HOCl in the OA region, probes with dual functions of diagnostic and therapeutic capabilities are still significantly lacking. While this type of probe can reduce the time gap between diagnosis and treatment, which is clinically needed. Methods: We developed a fluorescent probe (DHU-CBA1) toward HOCl with theranostics functions through the release of methylene blue (MB) and ibuprofen (IBP) in this work. DHU-CBA1 can detect HOCl with high specificity and sensitivity, releasing MB and IBP with an impressive efficiency of ≥ 95% in vitro. Results: DHU-CBA1 exhibits good biosafety, enabling in vivo imaging of endogenous HOCl, along with reducing arthritis scores, improving synovitis and cartilage damage, and maintaining catabolic balance while alleviating senescence in cartilage. Conclusions: This study proposes a novel approach to enhance osteoarthritis therapy by releasing IBP via a smart HOCl-enabled fluorescent probe.

Dual benefits of NBD-Cl fluorogenic action and sample pretreatment (SALLE) technology in the assessment of anticoagulant medication: Dabigatran in pharmaceutical capsules and plasma samples.

Dabigatran (DBG), marketed as Pradaxa, is an anticoagulant medication prescribed for the treatment and mitigation of blood clots and to lower the risk of stroke in individuals with the heart condition known as atrial fibrillation. This medication is specifically indicated for preventing blood clots post hip or knee replacement surgeries and in patients with a prior history of clots. Compared to warfarin, dabigatran serves as a viable alternative that does not necessitate routine blood monitoring tests. The complimentary benefits associated with SALL (salting-out assisted liquid-liquid extraction) and the fluorogenic capabilities of benzofurazan. These methods were combined to provide an affordable and sensitive DBG assaying method. The spectral strength of the yellow luminous product was examined at 533.8 nm and by adjustment of a wavelength of 474.7 nm for excitation. To assess its linearity, the calibration chart was tested across a DBG concentration range of 30-500 ng/ml. Via accurate computation based on ICH, the detection limit (LD) was determined to be 9.5 ng/ml, and the strategy can quantify the DBG to a limit of 28 ng/ml. To ensure success, various crucial parameters for method implementation have been extensively studied and adapted. The validation of the strategy adhered to the policies outlined by ICH, affirming its precision in quantifying DBG in capsules. Furthermore, the inclusion of SALLE steps facilitated accurate monitoring of DBG in plasma samples, introducing a unique and advanced methodology for analyzing this compound in biological samples.

Sensitive and rapid determination of tetracycline antibiotic by carrot juice-derived carbon dots as a fluorescent probe.

The antibiotic tetracycline can be efficiently used as medicine for the deterrence of bacterial infections in humans, animals, and plants. However, the unprecedented use of tetracycline is of great concern owing to its low biodegradability, extensive usage, and adverse impacts on the environment and water quality. In this study, a sensitive spectrofluorometric method was proposed for the direct determination of tetracycline, based on biocompatible fluorescent carbon dots (CDs). The synthesis of CDs was performed by adopting a green hydrothermal procedure from carrot juice without requiring surface passivation or outflowing any environmentally hazardous waste. X-ray diffraction analysis and transmission electron microscopy revealed amorphous spherical-shaped CDs that exhibited blue emission under blue illumination. The fabricated fluorescent probe directly detected tetracycline in the concentration range of 4.00 × 10-6 to 1.55 × 10-5 mol L-1 with an LOD of 1.33 × 10-6 mol L-1. The performance of the probe was assessed in a tap water sample, with recovery values between 80.70 and 103.60%. The method's greenness was evaluated using the Analytical Green metric approach (AGREE) and confirmed to be within the green range. The developed method is facile, rapid, cost-effective, and offers a wide linear range and satisfactory selectivity, making it potentially suitable for determining tetracycline in water applications.