An activatable near-infrared fluorescent probe with large Stokes shift for visualizing peroxynitrite in Alzheimer's disease models.

Alzheimer's disease (AD), characterized by its incurable nature and prevalence among the elderly, has remained a focal point in medical research. Increasing evidence suggests that peroxynitrite (ONOO-) serves as a crucial biomarker for the diagnosis of AD. In this study, we present a novel, easily available, high-yield, and cost-effective near-infrared (NIR) fluorescent probe, CDCI-ONOO. This probe utilizes a coumarin-dicyanoisophorone conjugate as the fluorophore and diphenylphosphinic chloride as the recognition site, enabling the detection of ONOO- both in vitro and in vivo. Upon interaction with ONOO-, CDCI-ONOO exhibits a distinct maximum emission peak at 715 nm with a substantial Stokes shift of 184 nm. The probe demonstrates excellent selectivity and sensitivity (LOD = 144 nM), along with noticeable colorimetric and fluorescence changes after the reaction. Comprehensive analyses using high-performance liquid chromatography (HPLC), high-resolution mass spectrometry (HRMS), and density functional theory (DFT) calculations confirm that the reaction with ONOO- restores the initially quenched Intramolecular Charge Transfer (ICT), resulting in the formation of CDCI-OH, a product that emitting fluorescence in the near-infrared region. Furthermore, we demonstrated the successful application of CDCI-ONOO for ONOO- detection in neuronal cells and imaging of ONOO- in the brains of mice. These findings underscore the potential of CDCI-ONOO as a near-infrared fluorescent probe for in vivo ONOO- detection, offering a significant avenue for advancing our understanding of AD pathology and diagnosis.

A near-infrared fluorescent probe with a large Stokes shift for the detection and imaging of biothiols in vitro and in vivo.

In this study, a new near-infrared (NIR) fluorescent turn-on probe featuring a large Stokes shift (198 nm) was developed for the detection of biothiols. The probe was based on a dicyanoisophorone derivative serving as the fluorophore and a 2,4-dinitrobenzenesulfonyl (DNBS) group functioning as both a recognition site and a fluorescence quencher. In the absence of biothiols, the fluorescence of the probe was low due to the photoinduced electron transfer (PET) effect between the fluorophore and DNBS. Upon the presence of biothiols, the DNBS group underwent a nucleophilic aromatic substitution reaction with the sulfhydryl group of biothiols, leading to the release of the fluorophore and a notable emission peak at 668 nm. This developed probe exhibited exceptional selectivity and sensitivity to biothiols in solution, with an impressive detection limit of 28 nM for cysteine (Cys), 22 nM for homocysteine (Hcy), and 24 nM for glutathione (GSH). Furthermore, the probe demonstrated its applicability by successfully visualizing both endogenous and exogenous biothiols in living systems.

Cellulose-based yellow-green emitting carbon dots with large Stokes shift as effective "turn off-on" fluorescence platforms for Cr (VI) and AA dual efficacy detection.

BACKGROUND: Hexavalent chromium (Cr (VI)) is highly carcinogenic to humans. Ascorbic acid (AA) deficiency can be hazardous to health. And the dual-effect fluorescence detection of them is an important research topic. Carbon dots (CDs) based on cellulose are excellent candidates for the fluorescence probes due to their low cost and environmental friendliness. But most of them exhibit shortwave emission, small Stokes shift and poor fluorescence performance, all of which limit their use. Therefore, there is an urgent need for cellulose CDs with longer emission wavelengths and larger Stokes shifts in dual-effect fluorescence detection of Cr (VI) and AA. RESULTS: Under optimal conditions (180 °C, 12 h), we prepared cellulose-based nitrogen-doped carbon dots (N-CDs) by a simple one-step hydrothermal process, which display longer emission wavelengths (ex: 370 nm, em: 510 nm), larger Stokes shifts (140 nm) and high fluorescence quantum yield (QY: 19.27 %). The continuous "turn-off" and "turn-off-on" fluorescence detection platforms were constructed based on the internal filtering effect (IFE) between Cr6+ and N-CDs, and Cr6+ reduced to Cr3+ by AA at pH = 6. The platform has been successfully simultaneous detect Cr (VI) and AA with a wide range of 0.01-40 µM and 0.1-100 µM. And the lowest limits of detection (LOD) are 0.0303 µM and 0.072 µM, respectively. In the presence of some other metals, non-metal ions and water-soluble acids in the fruits, this fluorescent platform can demonstrate a high level of interference immunity. SIGNIFICANCE AND NOVELTY: This represents the first yellow-green cellulose-based N-CDs with large Stokes shift for dual-effect detection of Cr (VI) and AA in real water samples and fresh fruits. The fluorescence detection platform has the advantage of low volume detection. Less than 2 mL of sample is required for testing and results are available in <5 min. This method is rare and supply a novel idea for the quantitative monitoring of Cr (VI) and AA.

A Vector Nanoplatform for the Bioimaging of Deep-Seated Tumors.

Today, in preclinical studies, optical bioimaging based on luminescence and fluorescence is indispensable in studying the development of neoplastic transformations, the proliferative activity of the tumor, its metastatic potential, as well as the therapeutic effect of antitumor agents. In order to expand the capabilities of optical imaging, sensors based on the bioluminescence resonance energy transfer (BRET) mechanism and, therefore, independent of an external light source are being developed. A targeted nanoplatform based on HER2-specific liposomes whose internal environment contains a genetically encoded BRET sensor was developed in this study to visualize deep-seated tumors characterized by overexpression of human epidermal growth factor receptor type 2 (HER2). The BRET sensor is a hybrid protein consisting of the highly catalytic luciferase NanoLuc (an energy donor) and a LSSmKate1 red fluorescent protein with a large Stokes shift (an energy acceptor). During the bioimaging of disseminated intraperitoneal tumors formed by HER2-positive SKOV3.ip1cells of serous ovarian cystadenocarcinoma, it was shown that the developed system is applicable in detecting deep-seated tumors of a certain molecular profile. The developed system can become an efficient platform for optimizing preclinical studies of novel targeted drugs.

Naturally Derived Luminescent Material in Engineered Silk and Its Application as a Fluorescent Dye with a Large Stokes Shift and Sensing Capability.

Silkworms have provided valuable byproducts (spanning from high-quality textiles to health supplements) to humans for millennia. Despite their importance in sericultural economy and biotechnology, manifold possibilities inherent in the myriad natural or artificially generated silk varieties have been underestimated. In this paper, we report that the Yeonnokjam silk strain, which shows light-green color, contains quercetin fluorochrome (QueF) in sericin, and QueF can be used as a fluorescence dye with a large Stokes shift and high sensitivity to environmental temperature and pH, thus functioning as an environmental sensing material. A Stokes shift exceeding 180 nm, a quantum efficiency of 1.28%, and a rapid fluorescence decay of 0.67 ns are obtained, which are influenced by solvent polarities. Moreover, QueF can be used as a UV blocker as well, and its low cytotoxicity and biocompatibility further suggest promising prospects for diverse application in cosmetics and medical materials in the future.

A novel red-to-near-infrared AIE fluorescent probe for detection of Hg2+ with large Stokes shift in plant and living cells.

Due to the highly toxic nature of mercury ions to living organisms, accurately detecting Hg2+ in water samples and biological systems is of great significance. In this study, we designed and synthesized a novel red-to-near-infrared Aggregation-Induced Emission (AIE) fluorescent probe (named as DS) based Fluorene derivatives on specifically for Hg2+ detection. Probe DS can visually identify Hg2+ through an red-to-near-infrared fluorescence enhancement change, characterized by a large Stokes shift (130 nm) and AIE feature. This probe offers a fast response, high selectivity and sensitivity. The Hg2+-induced deprotection reaction of the thioketal mechanism was thoroughly investigated using nuclear magnetic resonance spectroscopy (NMR), mass spectrometry (MS) and density functional theory (DFT) calculation. Additionly, dynamic light scattering (DLS) results indicated that the aggregation states changes of the molecular play a crucial role in the AIE fluorescence response of probe DS toward Hg2+. The red-to-near-infrared response with AIE feature not only avoids the interference of auto-fluorescence signals in complex environments, but also reduces the fluorescence quenching caused by probe molecular aggregation. This makes probe DS highly suitable for high-quality imaging detection of Hg2+ in aqueous environments. Furthermore, probe DS demonstrates the capability for visual fluorescence detection of Hg2+ concentrations in water sample, plant roots and living cells.

Design and synthesis of large Stokes shift DNA dyes with reduced genotoxicity.

Despite intensive search over the past decades, only a few small-molecule DNA fluorescent dyes were found with large Stokes shifts. These molecules, however, are often too toxic for widespread usage. Here, we designed DNA-specific fluorescent dyes rooted in benzimidazole architectures with a hitherto unexplored molecular framework based on thiazole-benzimidazole scaffolding. We further incorporated a pyrazole ring with an extended sidechain to prevent cell penetration. These novel benzimidazole derivatives were predicted by quantum calculations and subsequently validated to have large Stokes shifts ranging from 135 to 143 nm, with their emission colors changed from capri blue for the Hoechst reference compound to iguana green. These readily-synthesized compounds, which displayed improved DNA staining intensity and detection limits along with a complete loss of capability for cellular membrane permeation and negligible mutagenic effects as designed, offer a safer alternative to the existing high-performance small-molecule DNA fluorescent dyes.

A Red-Emission Fluorescent Probe with Large Stokes Shift for Detection of Viscosity in Living Cells and Tumor-Bearing Mice.

Abnormal viscosity is closely related to the occurrence of many diseases, such as cancer. Therefore, real-time detection of changes in viscosity in living cells is of great importance. Fluorescent molecular rotors play a critical role in detecting changes in cellular viscosity. Developing red emission viscosity probes with large Stokes shifts and high sensitivity and specificity remains an urgent and important topic. Herein, a novel viscosity-sensitive fluorescent probe (TCF-VIS1) with a large stokes shift and red emission was prepared based on the 2-dicyanomethylene-3-cyano-4,5,5-trimethyl-2,5-dihydrofuran (TCF) skeleton. Due to intramolecular rotation, the probe itself does not fluorescence at low viscosity. With the increase in viscosity, the rotation of TCF-VIS1 is limited, and its fluorescence is obviously enhanced. The probe has the advantages of simple preparation, large Stokes shift, good sensitivity and selectivity, and low cytotoxicity, which make it successfully used for viscosity detection in living cells. Moreover, TCF-VIS1 showed its potential for cancer diagnosis at the cell level and in tumor-bearing mice by detecting viscosity. Therefore, the probe is expected to enrich strategies for the detection of viscosity in biological systems and offer a potential tool for cancer diagnosis.

Golgi-targeted NIR fluorescent probe with large stokes shift for real-time monitoring of nitric oxide in depression model.

Depression is a debilitating mental illness that poses a serious threat to human health. Nitric Oxide (NO), as an important gasotransmitter, is closely associated with the pathogenesis of depressive disorders. Effective monitoring of NO fluctuation is beneficial for the diagnosis of depression and therapy assessment of antidepressants. Currently, there is a lack of effective methods for rapidly and sensitively identifying NO and elucidating its relationship with depression diseases. Herein, we developed a NIR dye TJ730-based fluorescent probe TJ730-Golgi-NO incorporating benzenesulfonamide as a Golgi-targeted moiety and the thiosemicarbazide group for NO detection. The probe exhibited turn-on fluorescence ability and a large Stokes shift of 158 nm, which shows high sensitivity, selectivity, and rapid response (<1 min) for NO detection. TJ730-Golgi-NO could detect exogenous and endogenous NO in cells stimulated by Glu and LPS, and target Golgi apparatus. Moreover, we disclose a significant increase of NO in the depression model and a weak fluorescence evidenced in the fluoxetine-treated depression mice. This study provides a competent tool for studying the function of NO and helping improve the effective treatment of depression diseases.

Elucidating photocycle in large Stokes shift red fluorescent proteins: Focus on mKeima.

Large Stokes shift red fluorescent proteins (LSS-RFPs) are genetically encoded and exhibit a significant difference of a few hundreds of nanometers between their excitation and emission peak maxima (i.e., the Stokes shift). These LSS-RFPs (absorbing blue light and emitting red light) feature a unique photocycle responsible for their significant Stokes shift. The photocycle associated with this LSS characteristic in certain RFPs is quite perplexing, hinting at the complex nature of excited-state photophysics. This article provides a brief review on the fundamental mechanisms governing the photocycle of various LSS-RFPs, followed by a discussion on experimental results on mKeima emphasizing its relaxation pathways which garnered attention due to its >200 nm Stokes shift. Corroborating steady-state spectroscopy with computational studies, four different forms of chromophore of mKeima contributing to the cis-trans conformers of the neutral and anionic forms were identified in a recent study. Furthering these findings, in this account a detailed discussion on the photocycle of mKeima, which encompasses sequential excited-state isomerization, proton transfer, and subsequent structural reorganization involving three isomers, leading to an intriguing temperature and pH-dependent dual fluorescence, is explored using broadband femtosecond transient absorption spectroscopy.

Near-infrared Rhodols-based fluorescent probe with large Stokes shift for tracking of H2S in food spoilage and living cells.

Hydrogen sulfide (H2S), as a biomarker signaling gas, is not only susceptible to food spoilage, but also plays a key function in many biological processes. In this work, an activated near infrared (NIR) H2S fluorescent probe was designed and synthesized with quinoline-conjugated Rhodols dye as fluorophore skeleton and a dinitrophenyl group as the responsive moiety. Due to the quenching effect of dinitrophenyl group and the closed-loop structure of Rhodols fluorophore, probe itself has a very weak absorption and fluorescence background signal. After the H2S-induced thiolysis reaction, the probe exhibits a remarkable colormetric change and NIR fluorescent enhancement response at 716 nm with large Stokes shift (116 nm), and possesses high sensing selectivity and sensitivity with a low detection limits of 330 nM. The response mechanism is systematically characterized by 1H NMR, MS and DFT calculations. The colorimetric change allows the probe to be used as a test strips to detect H2S in food spoilage, while NIR fluorescent response helps the probe monitor intracellular H2S.

A Reaction-based Ratiometric Fluorescent Probe with Large STOKES Shift for Cu2+ in Neat Aqueous Solution.

A novel reaction-based ratiometric fluorescent probe 1 for Cu2+ using picolinate as the reaction site and hemicyanine as the fluorophore was developed. 1 displayed maximum absorption peak at 355 nm and fluorescence emission peak at 500 nm, with large Stokes shift of 145 nm. Upon reaction with Cu2+, the maximum absorption and fluorescence emission peaks red-shifted to 390 nm and 570 nm respectively, owing to Cu2+-induced hydrolysis of the picolinate moiety in 1. Meanwhile, the solution of 1 turned from green to orange under a 365 nm UV lamp. 1 not only could detect Cu2+ ratiometrically by the ratios of both absorbance (A390 nm/A355 nm) and fluorescence intensity (F570 nm/F500 nm), but also displayed large Stokes shift, fast response, high sensitivity and excellent selectivity over other metal ions in neat aqueous solution.

A near-infrared fluorescence probe with large Stokes shift for selectively monitoring nitroreductase in living cells and mouse tumor models.

Hypoxia is commonly regarded as a typical feature of solid tumors, which originates from the insufficient supply of oxygen. Herein, the development of an efficient method for assessing hypoxia levels in tumors is strongly desirable. Nitroreductase (NTR) is an overexpressed reductase in the solid tumors, has been served as a potential biomarker to evaluate the degrees of hypoxia. In this work, we elaborately synthesized a new near-infrared (NIR) fluorescence probe (MR) to monitor NTR activity for assessment of hypoxia levels in living cells and in tumors. Upon exposure of NTR, the nitro-unit of MR could be selectively reduced to amino-moiety with the help of nicotinamide adenine dinucleotide. Moreover, the obtained fluorophore emitted a prominent NIR fluorescence, because it possessed a classical "push-pull" structure. The MR displayed several distinguished characters toward NTR, including intense NIR fluorescent signals, large Stokes shift, high selectivity and low limit of detection (46 ng/mL). Furthermore, cellular confocal fluorescence imaging results validated that the MR had potential of detecting NTR levels in hypoxic cells. Significantly, using the MR, the elevated of NTR levels were successfully visualized in the tumor-bearing mouse models. Therefore, this detecting platform based on this probe may be tactfully constructed for monitoring the variations of NTR and estimating the degrees of hypoxia in tumors.

Dicyanoisophorone-based near-infrared fluorescent probe with large Stokes shift for the monitoring and bioimaging of hypochlorite.

Hypochlorite (ClO-), as one of reactive oxygen species (ROS), is closely linked to various illnesses and is essential for the proper functioning of immune system. Hence, monitoring and assessing ClO- levels in organisms are extremely important for the clinical diagnosis of ClO--related disorders. In this study, a novel ClO--selective fluorescent probe, DCP-ClO, was synthesized with dicyanoisophorone-xanthene unit as parent fluorophore, which displayed excellent selectivity towards ClO-, near-infrared emission (755 nm), large Stokes shift (100 nm), real-time response to ClO-, high sensitivity (LOD = 3.95 × 10-8 M), and low cytotoxicity. The recognition mechanism of DCP-ClO towards ClO- was confirmed to be a typical ICT process by HPLC-MS, HR-MS, 1H NMR and theoretical calculations. Meanwhile, DCP-ClO demonstrated remarkable efficacy in monitoring ClO- levels in water samples and eye-catching ability in imaging endogenous/exogenous ClO- in living organisms, which verified its potential as a powerful tool for the recognition of ClO- in complex biological systems.

Organic Fluorophores with Large Stokes Shift for the Visualization of Rapid Protein and Nucleic Acid Assays.

Organic small-molecule fluorophores, characterized by flexible chemical structure and adjustable optical performance, have shown tremendous potential in biosensing. However, classical organic fluorophore motifs feature large overlap between excitation and emission spectra, leading to the requirement of advanced optical set up to filter desired signal, which limits their application in scenarios with simple settings. Here, a series of wavelength-tunable small-molecule fluorescent dyes (PTs) bearing simple organic moieties have been developed, which exhibit Stokes shift up to 262 nm, molar extinction coefficients ranged 30,000-100,000 M-1 cm-1, with quantum yields up to 54.8 %. Furthermore, these dyes were formulated into fluorescent nanoparticles (PT-NPs), and applied in lateral flow assay (LFA). Consequently, limit of detection for SARS-CoV-2 nucleocapsid protein reached 20 fM with naked eye, a 100-fold improvement in sensitivity compared to the pM detection level for colloidal gold-based LFA. Besides, combined with loop-mediated isothermal amplification (LAMP), the LFA system achieved the visualization of single copy level nucleic acid detection for monkeypox (Mpox).

A Coumarin-Hemicyanine Deep Red Dye with a Large Stokes Shift for the Fluorescence Detection and Naked-Eye Recognition of Cyanide.

In this study, we synthesized a coumarin-hemicyanine-based deep red fluorescent dye that exhibits an intramolecular charge transfer (ICT). The probe had a large Stokes shift of 287 nm and a large molar absorption coefficient (ε = 7.5 × 105 L·mol-1·cm-1) and is best described as a deep red luminescent fluorescent probe with λem = 667 nm. The color of probe W changed significantly when it encountered cyanide ions (CN-). The absorption peak (585 nm) decreased gradually, and the absorption peak (428 nm) increased gradually, so that cyanide (CN-) could be identified by the naked eye. Moreover, an obvious fluorescence change was evident before and after the reaction under irradiation using 365 nm UV light. The maximum emission peak (667 nm) decreased gradually, whilst the emission peak (495 nm) increased gradually, which allowed for the proportional fluorescence detection of cyanide (CN-). Using fluorescence spectrometry, the fluorescent probe W could linearly detect CN- over the concentration range of 1-9 µM (R2 = 9913, RSD = 0.534) with a detection limit of 0.24 µM. Using UV-Vis spectrophotometry, the linear detection range for CN- was found to be 1-27 µM (R2 = 0.99583, RSD = 0.675) with a detection limit of 0.13 µM. The sensing mechanism was confirmed by 1H NMR spectroscopic titrations, 13C NMR spectroscopy, X-ray crystallographic analysis and HRMS. The recognition and detection of CN- by probe W was characterized by a rapid response, high selectivity, and high sensitivity. Therefore, this probe provides a convenient, effective and economical method for synthesizing and detecting cyanide efficiently and sensitively.

Near-Infrared Fluorescence Probe for Monoamine Oxidase A with a Large Stokes Shift for Intraoperative Navigation.

Monoamine oxidase A (MAO-A) is a dimeric flavoprotein that is found in the mitochondrial membrane. Currently, there is a lack of near-infrared fluorescent probes (NIR-FPs) with good specificity and high sensitivity for detecting MAO-A, making it difficult to accurately recognize and image cells in vitro and in vivo. In this study, the NIR-FP DDM-NH2 was designed and synthesized in order to detect MAO-A specifically in live biological systems. The probe comprised two functional components: dicyanoisophosphone as an NIR dye precursor and alanine as a recognition moiety. After identifying MAO-A, the probe exhibited an NIR emission peak at 770 nm with a significant Stokes shift (180 nm), 11-fold response factor, low detection limit of 99.7 nM, and considerably higher affinity toward MAO-A than that toward MAO-B, indicating high sensitivity. In addition, DDM-NH2 was effective when applied to the image-based assessment of MAO-A activity in HeLa cells, zebrafish, and tumor-bearing mice, demonstrating great potential for visualization-based research and MAO-A application in vivo.

Nitrogen, sulfur-doped carbon quantum dots with large Stokes shift for real-time monitoring of pH in living cells.

Construction of carbon quantum dots-based (CQDs) fluorescent probes for real-time monitoring pH in cells is still unsatisfied. Here, we propose the synthesis of nitrogen, sulfur-doped CQDs (N,S-CQDs) using one-pot hydrothermal treatment, and serve it as fluorescent probes to realize the real-time sensing of intracellular pH. These pH-responsive N,S-CQDs were proved exhibited a diversity of admirable properties, including great photostability, nontoxicity, favorable biocompatibility, and high selectivity. Particularly, due to the doping of nitrogen and sulfur, N,S-CQDs possessed long-wavelength emission and large Stokes Shift (190 nm), which could avoid self-absorption of tissue to realize high contrast and resolution bioimaging. The response of the probes to pH showed a good linear in range of 0.93-7.00 with coefficient of determination of 0.9956. Moreover, with advantages of high signal-to-noise ratio and stability against photobleaching, the as-prepared N,S-CQDs were successfully applied to monitor pH in living cells via bioimaging. All findings suggest that N,S-CQDs have significant potential for practical application for sensing and visualizing pH fluctuation in living systems.

Development of europium(III) complex fluorescent probe for hydrogen sulfide detection and its application in water samples.

Hydrogen sulfide (H2 S) is a crucial endogenous signaling component in organisms that is involved in redox homeostasis and numerous biological processes. Modern medical research has confirmed that hydrogen sulfide plays an important role in the pathogenesis of many diseases. Herein, a fluorescent probe Eu(ttbd)3 abt based on europium(III) complex was designed and synthesized for the detection of H2 S. Eu(ttbd)3 abt exhibited significant quenching for H2 S at long emission wavelength (625 nm), with rapid detection ability (less than 2 min), high sensitivity [limit of detection (LOD) = 0.41 µM], and massive Stokes shift (300 nm). Additionally, this probe showed superior selectivity for H2 S despite the presence of other possible interference species such as biothiols. Furthermore, the probe Eu(ttbd)3 abt was successfully applied to detect H2 S in water samples.

A Reaction-Based ESIPT Fluorescent Probe for the Detection of Hg2+ with Large Stokes Shift.

A novel reaction-based fluorescent probe 1 for Hg2+ was designed and synthesized. 1 was almost nonfluoresent due to inhibition of the ESIPT process between hydroxy group and imid carbonyl oxygen by diphenylphosphinothioate group. After reacting with Hg2+, the fluorescence intensity of 1 exhibited significant enhancement owing to recovery of the ESIPT process via Hg2+-promoted desulfurization-hydrolysis of the diphenylphosphinothioate moiety and cleavage of the P-O bond. 1 not only showed rapid response, high sensitivity, excellent selectivity for Hg2+ over other metal ions, but also could detect Hg2+ with large Stokes shift (165 nm), which was attributed to the ESIPT process. Moreover, the reaction mechanism was fully validated by absorption spectra, fluorescence spectra, fluorescence color as well as ESI-MS analysis. 1 is the reaction-based ESIPT fluorescent probe for the detection of Hg2+ with large Stokes shift, rapid response, high sensitivity and selectivity.