Turn-on Response Research Articles

A naphthalimide-based fluorescent probe with mitochondria targeting for GSH sensing and cancer cell recognition

GSH is an important biological thiol which involves many biological processes, and also an biomarker for cancer. So it's urgent to develop fluorescent probes with excellent sensitivity and high selectivity for GSH detection. Herein, we develop a new GSH-responsive fluorescent probe TPP-Naph-SS-Fc based on PET (photoinduced electron transfer) mechanism. TPP-Naph-SS-Fc contains naphthalimide fluorephore, the ferrocene quencher, the disulfide cleavage bond and the triphenylphosphine mitochondrial localizer. It shows a fluorescence turn-on response toward GSH for cleavage the disulfide bond, which result in the blocking of PET process. Especially, this probe can localize in mitochondria and make real-time imaging of GSH, also demonstrate the ability to distinguish between cancerous and normal cells.

A novel α-ketoamide reactivity-based two-photon fluorogenic probe for visualizing peroxynitrite in Parkinson’s disease models

Peroxynitrite (ONOO[Formula: see text] contributes to oxidative stress and neurodegeneration in Parkinson’s disease (PD). Developing a peroxynitrite probe would enable in situ visualization of the overwhelming ONOO− flux and understanding of the ONOO− stress-induced neuropathology of PD. Herein, a novel [Formula: see text]-ketoamide-based fluorogenic probe ( DFlu) was designed for ONOO− monitoring in multiple PD models. The results demonstrated that DFlu exhibits a fluorescence turn-on response to ONOO− with high specificity and sensitivity. The efficacy of DFlu for intracellular ONOO− imaging was demonstrated systematically. The results showed that DFlu can successfully visualize endogenous and exogenous ONOO− in cells derived from chemical and biochemical routes. More importantly, the two-photon excitation ability of DFlu has been well demonstrated by monitoring exogenous/endogenous ONOO− production and scavenging in live zebrafish PD models. This work provides a reliable and promising [Formula: see text]-ketoamide-based optical tool for identifying variations of ONOO− in PD models.

Covalent post-synthetic modified metal-organic framework UIO-66-NH2-HNA for selective and sensitive turn-on detection of acetylacetone, S2−, and PO43−

A fluorescent probe named UIO-66-NH2-HNA was prepared by covalent post-synthetic modification of UIO-66-NH2. Experiments show that the fluorescence and structure stability of UIO-66-NH2-HNA can be maintained in a wide pH range of 1–12. UIO-66-NH2-HNA can act as a turn-on fluorescent probe to detect not only acetylacetone (acac), but also S2− and PO43−, and the detection limits are as low as 0.25, 0.083 and 0.069 μM, respectively. Further study on the mechanisms showed that the sensing mechanism of UIO-66-NH2-HNA for acac could be ascribed to donor–acceptor electron-transfer, and the detection mechanism for S2− and PO43− could be attributed to the framework collapse caused by the increase of pH. It is worth mentioned that UIO-66-NH2-HNA is the first MOF probe based on covalent post-synthetic modification of aldimine condensation reaction for the detection of acac by turn-on response.

Unique development of a new dual application probe for selective detection of antiparallel G-quadruplex sequences.

G-Quadruplex (G4) structures play vital roles in many biological processes; consequently, they have been implicated in various human diseases like cancer, Alzheimer's disease etc. The selective detection of G4 DNA structures is of great interest for understanding their roles and biological functions. Hence, development of multifunctional fluorescent probes is indeed essential. In this investigation, we have synthesized a quinolinium based dual application probe (QnMF) that presents molecular rotor properties. This dual application molecular rotor is able to detect selectively antiparallel G4 sequences (22AG in 100 mM NaCl) through a turn-on response over other G4 topologies. The QnMF also contains a distinct fluorine-19 that undergoes a significant chemical shift in response to microenvironmental changes around the molecule when bound to G4 structures. The probe QnMF exhibits significantly brighter fluorescence emissions in glycerol (ε × ϕ = 2800 cm-1 M-1) and relatively less brighter fluorescence emissions in methanol (ε × ϕ = 40.5 cm-1 M-1). The restricted rotation inherent property of the QnMF molecular rotor is responsible for brighter fluorescence and leads to enhancement in the fluorescence upon binding to the G4 structure. Overall, the probe's dual detection method makes it useful for monitoring the G4 structures that are abundant and plays a vital role in living organisms.

A two-photon fluorescent probe for formaldehyde detection and regeneration in living cells.

A two-photon excited fluorescent probe CMB-1 has been rationally developed for the detection and regeneration of formaldehyde based on a novel nucleophilic addition of a secondary amine to FA and subsequential alcoholysis reactivity mechanism. It enables a specific turn-on response towards formaldehyde and facilitates the monitoring of exogenous and endogenous formaldehyde in living cells via both one- and two-photon microscopy, with minimal influence on its native homeostasis and local concentration.

A turn-on fluorescent probe for selective detection of H2S in environmental samples and bio-imaging in human breast cancer cells.

A triphenylamine-benzothaizole-based turn-on fluorescent probe TPB-NO2 was designed and synthesized for tracking H2S in both environmental and biological samples depending upon the sensing strategy of thiolysis of 2,4-dinitrophenyl (DNP) ether. Due to PET (photoinduced electron transfer), occurring from donor triphenylamine moiety to acceptor DNP moiety, the probe TPB-NO2 itself is very weakly fluorescent and colorless in DMSO/H2O solution (1 : 1, v/v; 10 mM HEPES buffer, pH 7.4). But the addition of H2S leads to thiolysis of 2,4-dinitrophenyl ether to block the initial PET process and hence it exhibits naked eye detectable turn-on response with bright cyan fluorescence and intense brown color. Not only that, the probe exhibits excellent selectivity over other bio-thiols like Cysteine (Cys), homocysteine (Hcy), and glutathione (GSH), fast response time (<2 min), and high sensitivity with a detection limit of 9.81 nM. Moreover, to explore the practical applicability of our probe we employed it to monitor H2S successfully in environmental water samples, solid-state TLC strip study, Quantitative determination of H2S in eggs, and in the bioimaging of human breast cancer cells (MDA-MB 231).

Aggregation assisted turn-on response of ANS dye towards protamine

Formation of an electrostatically-driven ANS–Pr assembly for protamine and heparin sensing.

Glutathione displacement assay based on a fluorescent Au(I) complex.

Glutathione (GSH) is an essential molecule that plays a pivotal role in maintaining intracellular redox homeostasis, as well as other physiological processes. However, the chemical mechanisms underlying the GSH-induced processes remain insufficiently understood due to the lack of appropriate detection tools. Fluorescence GSH imaging can serve as a useful principle for the rapid, convenient, and non-destructive detection of GSH in living organisms. In this study, we developed a fluorescent GSH probe based on a linear, homoleptic Au(I) complex with two 1,3-diphenylbenzimidazolium carbene ligands. The Au(I) complex produced a fluorescence turn-on response to GSH. Fluorescence GSH signaling was characterized with a short response time of a few seconds. The rapid response was attributed to the displacement of the carbene ligand with GSH, which involved a labile inner-sphere coordination interaction. Finally, we demonstrated the biological utility of our GSH probe by unambiguously discriminating between different GSH levels in normal and senescent preadipocytes.

A mitochondria-targeted chemiluminescent probe for detection of hydrogen sulfide in cancer cells, human serum and in vivo.

Hydrogen sulfide (H2S) as a critical messenger molecule plays vital roles in regular cell function. However, abnormal levels of H2S, especially mitochondrial H2S, are directly correlated with the formation of pathological states including neurodegenerative diseases, cardiovascular disorders, and cancer. Thus, monitoring fluxes of mitochondrial H2S concentrations both in vitro and in vivo with high selectivity and sensitivity is crucial. In this direction, herein we developed the first ever example of a mitochondria-targeted and H2S-responsive new generation 1,2-dioxetane-based chemiluminescent probe (MCH). Chemiluminescent probes offer unique advantages compared to conventional fluorophores as they do not require external light irradiation to emit light. MCH exhibited a dramatic turn-on response in its luminescence signal upon reacting with H2S with high selectivity. It was used to detect H2S activity in different biological systems ranging from cancerous cells to human serum and tumor-bearing mice. We anticipate that MCH will pave the way for development of new organelle-targeted chemiluminescence agents towards imaging of different analytes in various biological models.

Real-time visualization of butyrylcholinesterase activity using a highly selective and sensitive chemiluminescent probe.

Butyrylcholinesterase (BChE), one of the critical human cholinesterases, plays crucial roles in numerous physiological and pathological processes. Accordingly, it is a striking and at the same time challenging target for bioimaging studies. Herein, we developed the first ever example of a 1,2-dixoetane-based chemiluminescent probe (BCC) for monitoring BChE activity in native biological contexts such as living cells and animals. BCC was initially shown to exhibit a highly selective and sensitive turn-on response in its luminescence signal upon reacting with BChE in aqueous solutions. Later, BCC was utilized to image endogenous BChE activity in normal and cancer cell lines. It was also shown through inhibition experiments that BChE can detect fluctuations of BChE levels successfully. In vivo imaging ability of BCC was demonstrated in healthy and tumor-bearing mice models. BCC enabled us to visualize the BChE activity in different regions of the body. Furthermore, it was successfully employed to monitor tumors derived from neuroblastoma cells with a very high signal to noise ratio. Thus, BCC appears as a highly promising chemiluminescent probe, which can be used to further understand the contribution of BChE to regular cellular processes and the formation of diseased states.

Cellular and Intravital Imaging of NAD(P)H by a Red-Emitting Quinolinium-Based Fluorescent Probe that Features a Shift of Its Product from Mitochondria to the Nucleus.

NAD(P)H is a vital hydrogen donor and electron carrier involved in numerous biological processes. The development of small-molecule tools for intravital imaging of NAD(P)H is significant for further exploring their pathophysiological roles. Herein, we rationally designed a fluorescent probe NADH-R by a simple graft of pyridiniumylbutenenitrile on a 1-methylquinolinium moiety in the 3-position. Benefited from the reduction of quinolinium by NAD(P)H, this probe releases the free push-pull fluorophore NADH-RH, allowing a turn-on red-emitting fluorescence response together with an ultralow detection limit of 12 nM. Under the assistance of the probe, we first monitored exogenous and endogenous generation of NAD(P)H in living cells, subsequently observed dynamic changes of NAD(P)H levels in living cells under different metabolic perturbations, and finally visualized the declined NAD(P)H levels in live mouse brain in a stroke model. Unexpectedly, the time-dependent colocalization experiment revealed that the probe reacts with mitochondrial NAD(P)H, followed by a shift of its reduced product NADH-RH from mitochondria to the nucleus, highlighting that NADH-RH is a novel nucleus-directed dye scaffold, which would facilitate the development of nucleus-targeting fluorescent probes and drugs.

Recognition of d-Glucose in Water with Excellent Sensitivity, Selectivity, and Chiral Selectivity Using γ-Cyclodextrin and Fluorescent Boronic Acid Inclusion Complexes Having a Pseudo-diboronic Acid Moiety.

Fluorescence recognition of d-glucose in water with excellent sensitivity, selectivity, and chiral selectivity is desired because d-glucose is an essential component in biological and pathological processes. We report an innovative approach that exploits the 1:2 stoichiometric inclusion complexes of γ-cyclodextrin (γ-CyD) with two molecules of fluorescent monoboronic acid-based receptors, which form a pseudo-diboronic acid moiety as the recognition site for d-glucose in water. Two monoboronic acids (1F and 2N) were easily synthesized without heating or column purification. The 1:2 stoichiometric inclusion complexes (1F/γ-CyD and 2N/γ-CyD) were prepared in a mixture of dimethyl sulfoxide/water (2/98 in v/v) by mixing γ-CyD and the corresponding monoboronic acids. Both 1F/γ-CyD and 2N/γ-CyD exhibited strong turn-on response to d-glucose with excellent selectivity over nine other saccharides in the water-rich solvent at pH 7.4 owing to the ditopic recognition of d-glucose by the pseudo-diboronic acid moieties. The limits of detection of 1F/γ-CyD and 2N/γ-CyD for d-glucose were 1.1 and 1.8 μM, respectively, indicating the remarkable sensitivity for the detection of d-glucose at μM levels. 1F/γ-CyD and 2N/γ-CyD also demonstrated chiral-selective recognition of d-glucose, which is apparent from the 2.0- and 6.3-fold enhancement of fluorescence by the addition of d-glucose relative to l-glucose addition, owing to the chiral pseudo-diboronic acid moieties produced by the chiral γ-CyD cavity. To the best of our knowledge, 2N/γ-CyD has the highest d/l selectivity among hitherto reported fluorescent diboronic acid-based receptors.

Recognition mechanism of imidazo[1,5-α]pyridine-based fluorescence probe towards thiophenols with multi-mechanisms of PET and ESIPT

The fluorescent turn-on response mechanism of an imidazo [1,5-α]pyridine-based probe MIPY-DNP for the detection of thiophenols (PhSH) has been theoretically studied. The first excited singlet (S1) state of probe MIPY-DNP is a dark state accompanying with complete charge transfer from the imidazo [1,5-α]pyridine group to the 2, 4-dinitrophenyl moiety, which implies the occurrence of the nonradiative photo-induced electron transfer (PET) process and explains the fluorescence quenching. The injection of PhSH induces the thiolysis of 2,4-dinitrophenyl ether group in the probe MIPY-DNP, and then the enol form of product MIPY-OH is formed. Due to lower potential barrier (6.72 kcal/mol) obtained by potential energy curves (PECs) in S1 state, the enol form of product MIPY-OH is converted into the keto form through the excited-state intramolecular proton transfer (ESIPT) process. Subsequently, the keto form of MIPY-OH emits strong fluorescence at 484 nm, which is consistent with the fluorescence observed in the experiment (478 nm). The frontier molecular orbitals (FMOs) and hole-electron analysis indicate that the electron transfer and distribution properties of the excited state, which further provides a reasonable explanation for the fluorescence quenching of probe MIPY-DNP and strong fluorescence of product MIPY-OH. Theoretical calculations gives a detailed detection mechanism of the probe MIPY-DNP towards PhSH based on the multi-mechanisms of PET and ESIPT.

Folic acid incorporated nitrogen-doped carbon dots as a turn-on fluorescence probe for homocysteine detection.

This study describes the development of a low-cost fluorescence assay for detecting homocysteine (Hcy) without the interference of cysteine and glutathione using carbon quantum dots. Herein nitrogen-doped carbon quantum dots (NCDs) were synthesized from citric acid as the carbon source and urea as the dopant using a one-pot microwave-assisted method. The obtained NCDs were incorporated with folic acid (FA) by the direct ex situ addition method and were used as a fluorescence probe to detect Hcy. The probe exhibited a fluorescence turn-on response with increased Hcy concentration up to 50 μM with a limit of detection of 2.276 μM. The point of care detection of Hcy using the probe was also tested with a paper-based assay strip.

A near infra-red emitting supramolecular dye-polymer assembly as promising platform for protamine sensing

Detection and quantification of bioanalytes using near infra-red (NIR) probes provide several advantages, viz, negligible scattering of light, minimum interference from the background matrices, deep sample penetration, etc. Herein, we report a supramolecular dye-polymer complex, comprising of mono-cationic LDS-798 dye and multi-anionic polystyrene sulfonate (PSS), as a promising NIR emitting turn-on probe for protamine (Pr) sensing. The supramolecular complex displays moderate fluorescence enhancement for LDS-798 due to its binding onto PSS surface. Strikingly, in the presence of Pr, this complex displays further enhancement in its emission intensity, which is also substantiated by a significant increase in the excited-state lifetime of LDS-798 dye. This turn-on response is found to be highly selective towards Pr, which is attributed to the structural change taking place for the anionic PSS on its interaction with highly cationic Pr molecule. Such a conformational change creates several cavities into the modified PSS structure, which can suitably encapsulate LDS-798 dye via strong hydrophobic interaction. Accordingly, there is large decrease in the radiationless decay processes for the excited dye molecules, making them strongly emissive in nature. Furthermore, LDS-798−PSS complex is found to be quite robust against changing temperature and ionic strength of the medium, indicating its possible use as a fluorescent probe over a wide range of temperature and salinity.

Development of novel fluorescent probes to detect and quantify specific reactive oxygen species

Hydrogen peroxide (H2O2) plays important roles in redox signaling and oxidative stress, and its dynamic concentration is critical to human health and diseases. Here we report the synthesis of a novel fluorescent probe based on pyridine-coumarin core 3-(2-pyridyl)-7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)chromen-2-one (P182) for quantitative measurement of H2O2. This fluorometric probe displays a fluorescence turn-on response in the process of deprotection of arylboronate to phenol in the presence of H2O2. It could offer good performances in terms of sensitivity and response time. Moreover, cytotoxicity studies on various model cell lines proved the non-toxic activity of the tested sensor allowing its use in vivo studies. This study provides research on molecular fluorescence probe for the detection of H2O2.

Anion-selective “Turn-on” two color phosphorescent probes based on “Pd-Pd” interaction of a series of cyclometallated Palladium (II) complexes induced by a self-assembly in aqueous solution

Herein, three pairs of cationic cyclometallated palladium (II) complexes with different types of C^N ligands, which is non-phosphorescent in aqueous solution, interestingly, they can be utilized as turn-on blue phosphorescent probes selectively for ClO−, HSO3− and CO32−, and turn-on green phosphorescent probe for HSO3− in aqueous solution. These different phosphorescent turn-on responses of Pd(II) complexes could be attributed to the degree of coordination and electrostatic interaction between them with specific anion. It suggests that the selectivity towards specific anion of these cyclometallated Pd(II) complexes can be further improved by rationally tuning the structure and enhancing aromaticity of C^N ligand. Our study reveals that these specific species of anions can effectively induce self-assembly of Pd(II) compounds with different C^N ligand based on PdPd interaction, the aggregation and morphology of palladium complex with anion in aqueous media was also investigated by various means of 1H NMR, UV/Vis, fluorescence spectra, and dynamic light scattering (DLS) analysis. Moreover, transmission electron microscopy (TEM) reveals that nanowires with increased length of diameters of Pd complexes can form in aqueous solution in presence of anions with different high concentration. Furthermore, the cellular uptake and location of Pd2a was also investigated by confocal imaging for the first time. DFT calculation of monomer and dimer of Pd2a was also performed, which is helpful to explain the turn on phosphorescent effect during self-assembly process.

Smartphone assisted fluorescent-colorimetric probe for bismuth (III) ion and potential applications

A new dual fluorescent-colorimetric probe (E)-4-methyl-N'-((2,3,6,7-tetrahydro-1H,5H-pyrido[3,2,1-ij]quinolin-9-yl)methylene)benzenesulfonohydrazide (L) was synthesized by condensation reaction of julolidine carboxaldehyde and p-toluene sulfonyl hydrazide to get the probe (L) for recognition of bismuth ions (Bi3+). The structural conformation of L was confirmed through 1H NMR, IR, and mass spectral analysis. The probe L showed visual color change and turn-on emission upon interaction with Bi3+ in H2O:CH3CN (7:3, v/v) media over other divalent and trivalent metal ions. The probe displays selective ‘turn-on’ emission for Bi3+via chelation enhanced fluorescence (CHEF) and > CN isomerization mechanism. The experimental analysis of Job’s plot showed a 2:1 stoichiometric ratio of L-Bi3+ complex formation and this was further verified with theoretical studies. The detection limit by spectrophotometric and spectrofluorometric were found to be 170 nM and 6.39 nM, respectively. The binding mechanism was verified by 1H NMR, ESI-mass, and theoretical studies. Furthermore, the selective colour change of L with Bi3+ was integrated with a smartphone to monitor the change in the RGB tool and test paper kit. The turn-on response of the probe with Bi3+ was used to capture by the fluorescence imaging technique in live Human Bone Osteosarcoma cell line (HOS) cells. Overall, probe L demonstrates a promising potential for the detection of Bi3+ ions in the protic environment and this is the first report of a fluorescent probe used in HOS cell live imaging.

PH tolerant metal ion controlled luminescence behaviour of supramolecular assembly and its application in bioimaging and supramolecular logic gate

Developing intelligent supramolecular assemblies endowed with highly tunable optical properties has been a challenge. Herein, we demonstrate pH tolerant metal ion controlled highly tunable optical properties of a fluorescence quenched supramolecular assembly composed of sulfated-β-cyclodextrin (SβCD) templated acridine orange (AO+) aggregates by using an external stimulus, aluminium (Al3+), a well-known neurotoxic metal ion. The addition of Al3+ ion to the AO+-SβCD complex provides a strong screening effect, resisting the electrostatic interaction between the AO+ and SβCD host, which triggers significant changes in the absorption and fluorescence characteristics of the AO+-SβCD complex along with a favorable fluorescence turn-On response. Additionally, the fluorescence On-Off-On switching of AO+ activated by SβCD and Al3+ ion was successfully utilised to construct an uncommon supramolecular-based two-input IMPLICATION logic gate. Finally, we could also monitor the fluorescence On-Off-On switching events of AO+ in more complex live MCF-7 cancer cells, suggesting the potential of the current system in real-time monitoring of in-vivo biosensing and drug delivery applications.

A flavonol-derived fluorescent probe for highly specific and sensitive detection of hydrazine in actual environmental samples and living zebrafish

Hydrazine (N2H4) is a significant chemical reagent and widely applied in industrial field, which can bring potential risk to environmental safety and human health due to its high toxicity and potential carcinogenicity. In this paper, a flavonol-derived fluorescent probe named TB-N2H4 was rationally developed for detecting N2H4 based on the excited intramolecular proton transfer (ESIPT) principle. TB-N2H4 exhibited a remarkable fluorescence turn-on response toward N2H4 with a large Stokes shift of 191 nm. Moreover, TB-N2H4 could selectively recognize N2H4 over other competitive analytes, and displayed high sensitivity toward N2H4 with a low detection limit of 0.117 μM. The sensing mechanism of the probe TB-N2H4 for N2H4 was confirmed by theoretical calculation and HRMS analysis. This probe was able to quantitatively determine N2H4 in environmental water and soil samples. Additionally, TB-N2H4 was also successfully utilized for real-time tracking of the distribution of N2H4 in living zebrafish.