Supramolecular Assembly of TPE-Based Glycoclusters with Dicyanomethylene-4H-pyran (DM) Fluorescent Probes Improve Their Properties for Peroxynitrite Sensing and Cell Imaging

Two red-emitting dicyanomethylene-4H-pyran (DM) based fluorescent probes were designed and used for peroxynitrite (ONOO⁻) detection. Nevertheless, the aggregation-caused quenching effect diminished the fluorescence and restricted their further applications. To overcome this problem, tetraphenylethylene (TPE) based glycoclusters were used to self-assemble with these DM probes to obtain supramolecular water-soluble glyco-dots. This self-assembly strategy enhanced the fluorescence intensity, leading to an enhanced selectivity and activity of the resulting glyco-dot comparing to DM probes alone in PBS buffer. The glyco-dots also exhibited better results during fluorescence sensing of intracellular ONOO⁻ than the probes alone, thereby offering scope for the development of other similar supramolecular glyco-systems for chemical biological studies.     

Aza-BODIPY based probe for photoacoustic imaging of ONOO− in vivo

The effective detecting ONOO⁻ variations in vivo is of great importance to well understand the complex pathophysiological processes. We reported here a photoacoustic (PA) probe AZB-1 for imaging ONOO⁻ in vivo. AZB-1 showed an originally strong photoacoustic signal at 660 nm. And its PA signal can be turned off by shutting the ICT effect caused by the conjugated electron withdrawing group at 2-position of the aza-BODIPY core. Moreover, the probe was successfully employed to imaging ONOO⁻ variations in inflammatory mice models. Wisely utilized this strategy may serve as powerful platforms for the preparation of novel PA chemosensors.     

Recent Progress of Small-Molecule Ratiometric Fluorescent Probes for Peroxynitrite in Biological Systems

Peroxynitrite (ONOO⁻) as a major reactive oxygen species plays important roles in cellular signal transduction and homeostatic regulation. Precise detection of ONOO⁻ in biological systems is vital for exploring its physiological and pathological function. Among numerous detection methods, fluorescence imaging technology using fluorescent probes offers some advantages, including simple operation, high sensitivity and selectivity, as well as real-time and nondestructive detection. In particular, ratiometric fluorescent probes, in which the built-in calibration of the two emission bands prevents interference from the biological environment, have been extensively employed to monitor the fluctuation of bioactive species. In this review, we will discuss small-molecule ratiometric fluorescent probes for ONOO⁻ in live cells or in vivo, which involves chemical structures, response mechanisms, and biological applications. Moreover, the challenges and future prospects of ONOO⁻-responsive ratiometric fluorescent probe are also proposed.     

Simultaneous tracking of autophagy and oxidative stress during stroke with an ICT-TBET integrated ratiometric two-photon platform

Over recent years, fluorescent probes exhibiting simultaneous responses to multiple targets have been developed for in situ, real-time monitoring of cellular metabolism using two photon fluorescence sensing techniques due to numerous advantages including ease of operation, rapid reporting, high resolution, long visualization time and being non-invasive. However, due to interference from different fluorescence channels during simultaneous monitoring of multiple targets and the lack of ratiometric capability amongst the available probes, the accuracy in tracing metabolic processes has been restricted. With this research, using a through-bond energy transfer (TBET) mechanism, we designed a viscosity and peroxynitrite (ONOO⁻) mitochondria-targeting two-photon ratiometric fluorescent probe Mito-ONOO. Our results indicated that with decreasing levels of mitochondrial viscosity and increasing levels of ONOO⁻, the maximum of the emission wavelength of the probe shifted from 621 nm to 495 nm under 810 nm two-photon excitation. The baselines for the two emission peaks were significantly separated (Δλ = 126 nm), improving the resolution and reliability of bioimaging. Moreover, by ratiometric analysis during oxygen-glucose deprivation/reoxygenation (OGD/R, commonly used to simulate cell ischemia/reperfusion injury), the real-time visualization of the metabolic processes of autophagy and oxidative stress was possible. Our research indicated that during cellular oxygen-glucose deprivation/reoxygenation, cells produce ONOO⁻, causing cellular oxidative stress and cellular autophagy after 15 min, as such Mito-ONOO exhibits the potential for the monitoring and diagnosis of stroke, as well as providing insight into potential treatments, and drug design.

Ratiometric simultaneous tracking of autophagy and oxidative stress was achieved using an ICT-TBET integrated platform. Mito-ONOO exhibited excellent selectivity, good chemical stability, and non-overlapping ratiometric signals.     

A Simple Near-Infrared Fluorescent Probe for the Detection of Peroxynitrite

Herein, we report the evaluation and synthesis of a reaction based fluorescent probe DCM-Bpin for the detection of Peroxynitrite (ONOO−). DCM-Bpin exhibits selective fluorescence off-on response for ONOO⁻ over other reactive oxygen species, including H₂O₂. Moreover, DCM-Bpin is biocompatible and has been used to visualize exogenous ONOO⁻ in HeLa cells.     

A novel turn-on fluorescent probe for Hg2+ detection based on rhodamine B spirolactam derivative

ABSTRACT

In this work, a new turn-on fluorescent probe 1 for Hg²⁺ ions detection based on rhodamine B spirolactam was reported. Among tested metal ions, probe 1 shows high selectivity towards Hg²⁺ in the the mixture solution of methanol and 0.02 M HEPES buffer (V/V = 9:1, pH = 7.2). No absorption and emission band of probe 1 was observed in the range from 450 to 700 nm. While only addition of Hg²⁺ to probe 1 could lead to appearance of a new absorption band centered at 553 nm and a fluorescence emission band around 577 nm upon excitation at 520 nm. Moreover, it exhibits excellent linear relationship (R² = 0.9993) between fluorescence intensity at 577 nm and the concentration of Hg²⁺ from 1.6 to 32 μM. The sensing mechanism was proven to be spirolactam ring open induced by Hg²⁺ through ¹H NMR, MS, absorption and fluorescence spectra. In addition, probe 1 could detect Hg²⁺ in real water samples and on filter paper, which demonstrates its application in environment science.     

Thiomaleimide Functionalization for Selective Biological Fluorescence Detection of Peroxynitrite as Tested in HeLa and RAW 264.7 Cells

The role of fluorescent molecules in diagnosis, treatment as well as in biomedical research has great current medicinal significance and is the focus of concentrated effort across the scientific research spectrum. Related research continues to reveal new practical sensing systems that bear enhanced features for interfacing of substituted molecules with biological systems. As part of an effort to better understand chalcogenide systems, a new dithiomaleimide BODIPY ( BDP‐NGM ) probe has been designed, synthesized and characterized. The fluorescence of BDP‐NGM was quenched by the incorporation of [3,4‐bis (phenylthio)] on the maleimide‐4‐phenyl moiety which is, in turn, placed at the meso ‐position of the BODIPY system. The probe shows a turn‐on fluorescence response upon reaction with ONOO⁻; mass spectral evidence reveals peaks in agreement with products involving oxidation of the sulfur groups to sulfone groups. An about 18.0‐fold emission intensity enhancement was found. By comparison, the emission signal from another ROS/RNS, superoxide, gave a modest turn on signal (≈5.0‐fold). The reaction is complete within 10 min, judging from the monitoring of the turn‐on fluorescence process; the detection limit was found to be 0.4 μm . BDP‐NGM can be used for the detection of ONOO⁻ under both acidic and basic conditions. Live cell imaging showed that the current probe can be used for the selective detection of ONOO⁻ in living systems.     

Spirolactonized Si-rhodamine: a novel NIR fluorophore utilized as a platform to construct Si-rhodamine-based probes

Spirolactonized Si-rhodamine was prepared as a platform to construct Si-rhodamine-based probes by following the design strategy widely used in rhodamine systems. Among them, the reaction-based probe SiR-Hg was operated for NIR sensing and bioimaging of Hg(2+) in living cells based on the similar irreversible spirolactam ring-opening process to traditional rhodamine derivatives.     

Reaction‐Based Fluorescent Probe for Detection of Endogenous Cyanide in Real Biological Samples

Herein, two compounds ( 1 a and 1 b ) were rationally constructed as novel reaction‐based fluorescent probes for CN^? by making use of the electron‐withdrawing ability of the cyano group that was formed from the sensing reaction. Notably, this design strategy was first employed for the development of fluorescent CN^? probes. The experimental details showed that probe 1 a exhibited a fluorescence turn‐on response to CN^?, whereas other anions, biological thiols, and hydrogen sulfide gave almost no interference. The detection limit of probe 1 a for CN^? was found to be 0.12 μM . The sensing reaction product of 1 a with CN^? was characterized by NMR spectroscopy and mass spectrometry. TD‐DFT calculations demonstrated that the formed cyano group drives the intramolecular charge transfer (ICT) process from coumarin dye to the cyano group and thus the original strong ICT from the coumarin dye to the 3‐position pyridyl vinyl ketone substituent is weakened, which results in recovery of coumarin fluorescence. The practical utility of 1 a was also examined. By fabricating paper strips, probe 1 a can be used as a simple tool to detect CN^? in field measurements. Moreover, probe 1 a has been successfully applied for quantitative detection of endogenous CN^? from cassava root.     

A novel ratiometric fluorescent probe based on coumarin derivative for the recognition of Al(III) and its application on test strips

A novel ratiometric probe (L) which was composed of chromone and coumarin moieties has been designed and synthesized for sensing Al³⁺ in EtOH in view of the internal charge transfer (ICT) mechanism. The free probe L exhibited a strong fluorescence emission at 477 nm, and the fluorescence emission here almost disappeared after adding Al³⁺ (10 equiv.) while a new peak appeared at 524 nm. This may be due to the enhancement of intramolecular electron transfer efficiency from donor to acceptor. In addition, this probe L could be form a 1:1 complex with Al³⁺, which could be explained by the ESI-MS spectra, and L had a low detection limit for Al³⁺ with a binding constant of 1.32 × 10⁴ M⁻¹. More importantly, L could be applied to a solid probe for rapid detection of Al³⁺ with a significant color change.     

A Fluorescent Probe for Al3+ Based on Calix[4]arene Containing Salicylaldehyde-Derived Schiff Base

Two novel fluorescent probes L1 and L2 for recognizing Al³⁺ have been prepared by cooperating salicylaldehyde-derived Schiff base groups onto the upper rim of the calix[4]arene. Of these probes, L2 is the most effective at recognizing Al³⁺, displaying a highly selective fluorescence “on” response with an emission wavelength at 478 nm and a Stokes shift of 88 nm. Additionally, probe L2 can form a 1 : 1 coordination complex with Al³⁺ with a binding constant of 2.6×10¹⁰ M⁻¹. Furthermore, its fluorescence intensity exhibits a good linear relationship with Al³⁺ concentration within the range of 2.0×10⁻⁵ M to 1.4×10⁻⁴ M, and the probe has a low detection limit of 4.36×10⁻⁷ M.     

Ultraviolet B Light-induced Nitric Oxide/Peroxynitrite Imbalance in Keratinocytes—Implications for Apoptosis and Necrosis

Elevation of nitric oxide (NO˙) can either promote or inhibit ultraviolet B light (UVB)-induced apoptosis. In this study, we determined real-time concentration of NO˙ and peroxynitrite (ONOO⁻) and their role in regulation of membrane integrity and apoptosis. Nanosensors (diameter 300–500 nm) were used for direct in situ simultaneous measurements of NO˙ and ONOO⁻ generated by UVB in cultured keratinocytes and mice epidermis. An exposure of keratinocytes to UVB immediately generated ONOO⁻ at maximal concentration of 190 nm followed by NO^˙ release with a maximal concentration of 91 nm . The kinetics of UVB-induced NO˙/ONOO⁻ was in contrast to cNOS agonist stimulated NO˙/ONOO⁻ from keratinocytes. After stimulating cNOS by calcium ionophore (CaI), NO˙ release from keratinocytes was followed by ONOO⁻ production. The [NO˙] to [ONOO⁻] ratio generated by UVB decreased below 0.5 indicating a serious imbalance between cytoprotective NO˙ and cytotoxic ONOO⁻—a main component of nitroxidative stress. The NO˙/ONOO⁻ imbalance increased membrane damage and cell apoptosis was partially reversed in the presence of free radical scavenger. The results suggest that UVB-induced and cNOS-produced NO˙ is rapidly scavenged by photolytically and enzymatically generated superoxide (O₂˙⁻) to produce high levels of ONOO⁻, which enhances oxidative injury and apoptosis of the irradiated cells.     

Highly sensitive perylene tetra-(alkoxycarbonyl) based colorimetric and ratiometric probes for fluorine detection

Fluoride anion (F^?) affects environmental, biological, and chemical processes significantly. Therefore, its detection has received increasing attention, and sensitive, effective, and convenient probes for F^? detection need to be developed urgently. In this work, two perylene tetra-(alkoxycarbonyl) (PTAC) based colorimetric and ratiometric probes, P1 and P2, were developed for the detection of F^?. The interactions between F^? and these two probes were investigated by absorption, electrochemistry, ¹H NMR, and density functional methods. Both the two probes were complexed with F^? with a ratio of 1:1. The detection limits of P1 and P2 were 0.22 μM and 0.87 μM, respectively. It was worth noting that the absorption peak of P1 showed a 190 nm red shift when sensing F^?, and P1 is the largest red shift value reported in F^? probes based on PTAC derivatives. This phenomenon was resulted from the unique configuration and deprotonation of P1 that can promote the intramolecular charge transfer (ICT). This strategy provides an example for the development of other ion probes based on D-A type ICT mechanism.     

Development of a Pre‐assembled Through‐Bond Energy Transfer (TBET) Fluorescent Probe for Ratiometric Sensing of Anticancer Platinum(ll) Complexes

Fluorescence microscopy has emerged as an attractive technique to probe the intracellular processing of Pt‐based anticancer compounds. Herein, we reported the first through‐bond energy transfer (TBET) fluorescent probe NPR1 designed for sensitive detection and quantitation of Pt^II complexes. The novel TBET probe was successfully applied for ratiometric fluorescence imaging of anticancer Pt^II complexes such as cisplatin and JM118 in cells. Capitalizing on the ability of the probe to discriminate between Pt^II complexes and their Pt^IV derivatives, the probe was further applied to study the activation of Pt^IV prodrug complexes that are known to release active Pt^II species after intracellular reduction.     

Integration of the 1,2,3‐Triazole “Click” Motif as a Potent Signalling Element in Metal Ion Responsive Fluorescent Probes

In a systematic approach we synthesized a new series of fluorescent probes incorporating donor–acceptor (D‐A) substituted 1,2,3‐triazoles as conjugative π‐linkers between the alkali metal ion receptor N‐phenylaza‐[18]crown‐6 and different fluorophoric groups with different electron‐acceptor properties (4‐naphthalimide, meso‐phenyl‐BODIPY and 9‐anthracene) and investigated their performance in organic and aqueous environments (physiological conditions). In the charge‐transfer (CT) type probes 1 , 2 and 7 , the fluorescence is almost completely quenched by intramolecular CT (ICT) processes involving charge‐separated states. In the presence of Na⁺ and K⁺ ICT is interrupted, which resulted in a lighting‐up of the fluorescence in acetonitrile. Among the investigated fluoroionophores, compound 7 , which contains a 9‐anthracenyl moiety as the electron‐accepting fluorophore, is the only probe which retains light‐up features in water and works as a highly K⁺/Na⁺‐selective probe under simulated physiological conditions. Virtually decoupled BODIPY‐based 6 and photoinduced electron transfer (PET) type probes 3 – 5 , where the 10‐substituted anthracen‐9‐yl fluorophores are connected to the 1,2,3‐triazole through a methylene spacer, show strong ion‐induced fluorescence enhancement in acetonitrile, but not under physiological conditions. Electrochemical studies and theoretical calculations were used to assess and support the underlying mechanisms for the new ICT and PET 1,2,3‐triazole fluoroionophores.     

An off-on fluorescent sensor with high selectivity and sensitivity for Fe(III)

A new fluorescent derivative (1) containing coumarin exhibits Fe(III)-selective strong yellow-green fluorescence in ethanol. This compound could be used as an “off-on” chemosensor and allow the detection of Fe³⁺ by monitoring changes in absorption and fluorescence spectra. Upon addition of Fe³⁺, an overall emission change of 125-fold was observed. High selectivity and sensitivity were observed over other metal ions, mainly due to the spirolactam ring-opening power of Fe³⁺. The detection limit was as low as 5.6?ppb. Photo-induced electron transfer, coupled with intramolecular charge transfer are proposed to account for the observed spectral response.     

A Quencher-Based Blood-Autofluorescence-Suppression Strategy Enables the Quantification of Trace Analytes in Whole Blood

Precise quantification of trace components in whole blood via fluorescence is of great significance. However, the applicability of current fluorescent probes in whole blood is largely hindered by the strong blood autofluorescence. Here, we proposed a blood autofluorescence-suppressed sensing strategy to develop an activable fluorescent probe for quantification of trace analyte in whole blood. Based on inner filter effect, by screening fluorophores whose absorption overlapped with the emission of blood, a redshift BODIPY quencher with an absorption wavelength ranging from 600–700 nm was selected for its superior quenching efficiency and high brightness. Two 7-nitrobenzo[c] [1,2,5] oxadiazole ether groups were introduced onto the BODIPY skeleton for quenching its fluorescence and the response of H₂S, a gas signal molecule that can hardly be quantified because of its low concentration in whole blood. Such detection system shows a pretty low background signal and high signal-to-back ratio, the probe thus achieved the accurate quantification of endogenous H₂S in 20-fold dilution of whole blood samples, which is the first attempt of quantifying endogenous H₂S in whole blood. Moreover, this autofluorescence-suppressed sensing strategy could be expanded to other trace analytes detection in whole blood, which may accelerate the application of fluorescent probes in clinical blood test.     

Overcoming the Oxygen Dilemma in Photoredox Catalysis: Near-Infrared (NIR) Light-Triggered Peroxynitrite Generation for Antibacterial Applications

The peroxynitrite anion (ONOO⁻) is closely associated with many diseases and the creation of ONOO⁻ donors is an essential means of understanding its pathophysiological functions. However, it is challenging to develop ONOO⁻ donors due to the difficulties in simultaneously producing highly reactive and short-lived nitric oxide (NO) and superoxide anion (O₂⋅⁻). Here, we report a novel strategy for constructing ONOO⁻ donors by combining near-infrared (NIR)-mediated type I photosensitization and photoredox catalysis. The key design using a Nile blue analogue that can serve as both a type I photosensitizer and a metal-free photocatalyst. Intriguingly, the formation of O₂⋅⁻ via type I photosensitization avoids oxygen interference and instead activates nitrobenzofurazan-based NO donors via oxygen-tolerant NIR photoredox catalysis. The simultaneous release of O₂⋅⁻ and NO leads to ONOO⁻ release, showing both antibacterial and antibiofilm activities.     

A boronate-based ratiometric fluorescent probe for fast selective detection of peroxynitrite

Given that peroxynitrite (ONOO^?) is profoundly associated with health and diseases, a new fluorescent probe ABT was designed and synthesized for detection of ONOO^?. ABT manifested not only ratiometric fluorescence signals simultaneously in response to concentrations of ONOO^? (within 10?s), but high selectivity and sensitivity towards ONOO^? over other physiological relevant species (detection limit?=?26.3?nM). Moreover, ABT worked in a broad pH range with biological relevance. Thus, ABT could be used to quantitative detection of ONOO^? concentration and has the potential to efficiently monitor ONOO^? in living organisms.     

A colormetric and fluorescent chemosensor for adenosine-5′-triphosphate based on rhodamine derivative

A rhodamine spirolactam derivative (1) was developed as a colormetric and fluorescent chemosensor for adenosine-5′-triphosphate (ATP) via hydrogen bonds interaction. As far as we know, this is the first case to explore ATP-induced ring-opening of spirolactam in rhodamine derivatives. It exhibited a highly sensitive “turn-on” fluorescent response toward ATP with a 47-fold fluorescence intensity enhancement under 20 equiv. of ATP added. The chemosensor can be applied to the quantification of ATP with a linear range covering from 1.0 × 10⁻⁷ to 2.0 × 10⁻⁴ M and a detection limit of 2.5 × 10⁻⁸ M. The experiment results show that the response behavior of 1 toward ATP is pH independent in medium condition (pH 6.0–8.0). Most importantly, the novel chemosensor has well solved the problem of serious interferences from other nucleoside polyphosphates such as ADP and AMP generally met by previously reported typical fluorescent chemosensors for ATP. Moreover, the response of the chemosensor toward ATP is fast (response time less than 3 min). In addition, the chemosensor can be used for the fluorescence assay for protein kinase activity with satisfactory results. The chemosensor for ATP based on hydrogen bonds interaction provided a novel strategy for the design of colormetric and ratiometric fluorescent probes for other target anions with high sensitivity and selectivity.