Design Of Fluorescent Probes Research Articles

Redefining Molecular Probes for Monitoring Subcellular Environment: A Perspective.

The development of small-molecule fluorescent probes has revolutionized the monitoring of in vivo physicochemical parameters, offering unprecedented insights into biological processes. In this perspective, we critically examine recent advances and trends in the design and application of fluorescent probes for real-time in vivo monitoring of subcellular environments. Traditional concepts such as membrane potential, microviscosity, and micropolarity have been superseded by more biologically relevant parameters like membrane voltage, tension, and hydration, enhancing the accuracy of physiological assessments. This redefinition not only presents an evolved concept with broader applications in monitoring subcellular dynamics but also addresses the unmet needs of subcellular biology more effectively. We also highlight the limitations of commonly used probes in providing specific information about the redox environment, noting their nonspecificity to oxidants and the influence of various chemical interactions. These probes typically rely on free radical mechanisms and require metal catalysts to react with hydrogen peroxide. They include naphthalimide, fluorescein, BODIPY, rhodamine, cyanine cores to cover the UV-vis-near-infrared window. The motif of this perspective is to provide critical insights into trending fluorescent-based systems employed in real-time or in vivo physicochemical-responsive monitoring, thus aiming to inform and inspire further research in creating robust and efficient fluorescent probes for comprehensive in vivo monitoring applications.

Rational Design of a Tandem Activatable Fluorescent Probe for Differential Diagnosis and Therapeutic Assessment of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a formidable disease, distinguished by its high aggressiveness and dismal outcomes. Although leucine aminopeptidase (LAP) has been widely employed as a biomarker in biological imaging of HCC, it is still susceptible to interference from false-positive signals activated in injured liver tissues. In this study, based on the significant difference of GSH levels in alcohol-damaged liver tissues and tumor tissues, a dual-tandem activatable probe (PCLT) was designed for differential diagnosis and treatment guidance of HCC by near-infrared fluorescence (NIRF) imaging. This probe comprised a dual-locked hemicyanine dye decorated with a tetraethylene glycol chain and dual-recognition unit of glutathione (GSH) and LAP, which could be sequentially cleaved by GSH and LAP to restore its NIRF signal. PCLT excellently discriminated orthotopic HCC from ALI far earlier (7 days) than histological analysis (28 days) and exhibited higher specificity toward early orthotopic HCC than the single-locked probe (PCL). In addition, PCLT is capable of accurately delineating the tumor contour, assisting in surgical resection of HCC tumors under fluorescence visualization, and noninvasively assessing the antitumor effect of HCC chemotherapy during ferroptosis, thereby presenting promising clinical implications for clinical diagnosis and therapy of HCC.

Balancing Brightness and Photobasicity: Modulating Excited-State Proton Transfer Pathways in Push-Pull Fluorophores for Biological Two-Photon Imaging.

Push-pull fluorophores with donor-π-acceptor architectures are attractive scaffolds for the design of probes and labels for two-photon microscopy. Such fluorophores undergo a significant charge-delocalization in the excited state, which is essential for achieving a large two-photon absorption cross-section and brightness. The polarized excited state may, however, also facilitate excited-state proton transfer (ESPT) pathways that can interfere with the probe response. Herein, we employed steady-state and time-resolved spectroscopic studies to elucidate whether ESPT is responsible for the pH-dependent emission response of the Zn(II)-selective fluorescent probe chromis-1. Composed of a push-pull architecture with a pyridine ring as the acceptor, the chromis-1 fluorophore core acts as a photobase that promotes ESPT upon acidification. Although the pKa of the pyridine acceptor increases more than six orders of magnitude upon excitation, the photobasicity is not sufficient to deprotonate solvent water molecules under neutral conditions. Rather, the pH-dependent emission response is caused by the pendant bis-isonicotinic acid chelating group which upon protonation facilitates an excited-state intramolecular proton transfer to the pyridine acceptor. A simple permutation of the core pyridine nitrogen from the para- to the ortho-position relative to the thiazole substituent was sufficient to reduce the excited-state basicity by two orders of magnitude without compromising the two-photon excited brightness. These results highlight the importance of choosing the appropriate fluorophore core and chelating moiety for minimizing pH-dependent responses in the design of fluorescent probes for biological imaging.

A novel terpene-BODIPY conjugates based fluorescent probes: Synthesis, spectral properties, stability, penetration efficiency into bacterial, fungal and mammalian cells

The design of fluorescent probes based on biocompatible luminophores for medical diagnostics is one of the rapidly developing areas worldwide. Here, we report the synthesis of a novel BODIPYs containing a propanoic acid residue at the α-position of one of the pyrrole rings conjugated to (+)-myrtenol or thiotherpenoid. Both conjugates are quite photostable (t1/2 ∼ 40 h) and exhibit high fluorescence efficiency (φfl ∼ 77–90 %). The advantage of the newly synthesized terpene-BODIPY conjugates lies in their stability and absence of fluorescence quenching over a wide pH range (1.65–9.18). Moreover, the affinity of the conjugates for lipid biostructures was significantly improved compared to the original α-BODIPY carboxylic acid. As opposed to unconjugated BODIPY, which shows negligible activity against any cell type used in the study, the luminophores with either (+)-myrtenol or thioterpenoid effectively stained the membranes of Gram-positive bacteria, while Gram-negative bacteria were not stained. In eukaryotic cells, both conjugates readily stained organelles, but not the cell and nuclear membranes, suggesting these compounds as tools for differential staining of microbes or cell structures. Taken together, our data demonstrates that conjugation of BODIPY to terpenoids significantly improves the fluorescence abilities and modulates the selectivity of the conjugates against various cells.

Design and synthesis of naphthalimide-based CO fluorescent probes realizing tricolor detection and turn-on/off response

Carbon monoxide (CO) fluorescent probes have attracted tremendous attention attributed to their low detection limit, high specificity, simple operation and good biocompatibility. However, the relationship between the molecular structure and the fluorescence response signal still needs to be systematically elucidated for different applications. Herein, three different CO fluorescent probes based on naphthalimide derivative with blue emission (B-NIA), green emission (G-NIA), and near-infrared emission (R-NIA) were designed and synthesized. Three-primary colors and opposite turn-on or turn-off fluorescent responses to CO were achieved by regulating the conjugation system of fluorophores and the position of fluorescent recognition group. The change of fluorescence response and sensing mechanism for CO detection was studied by theory calculation and mass spectrometry analysis. All the B/G/R-NIA possessed high sensitivity for CO detection. Notably, the R-NIA emerged with a noticeable NIR fluorescence response to CO with a low detection limit (0.61 µM) and high selectivity and relative pH stability. In addition, the R-NIA showed particularly low cytotoxicity and has been successfully used to detect CO in living cells. These studies provided the theoretical reference and technical route to synthesize fluorescent probes with different emission wavelengths and opposite fluorescent responses to CO for various application scenarios

Rationally Designed G-Quadruplex Selective "Turn-On" NIR Fluorescent Probe with Large Stokes Shift for Nucleic Acid Research-Based Applications.

Guanine-rich DNA/RNA sequences can form Hoogsteen bonds to adopt noncanonical secondary structures called G-quadruplexes, and these have been associated with diverse cellular processes. There has been considerable research interest in the design of G4-interacting ligands for cellular probing of the G4 structure and understanding its associated biological function. Most of the fluorescent G4 ligands either do not have significant selectivity over other nucleic acid structures, have high Stokes shift, or are not in the near-infrared (NIR) region, which limits its cellular visualization. The current work involves the rational design and synthesis of NIR fluorescent probes comprising a (Z)-1-methyl-2-((3-methylbenzo[d]thiazol-2(3H)-ylidene)methyl)quinolin-1-ium scaffold. Among the designed molecules, 4a exhibited far-red fluorescence (λmax = 680 nm) with large Stokes shift (∼182 nm) upon selective binding to human telomeric G-quadruplexes. The dye 4a does not disturb the conformation and stability of G-quadruplexes, thereby making it suitable for nucleic acid research based applications. Interestingly, 4a showed remarkable selectivity over single- and double-stranded structures in contrast to a commercially available quadruplex binding probe, Thiazole orange (TO). The molecular docking studies indicate that 4a binds at the groove region of the telomeric DNA G-quadruplex through π-π stacking interactions with the quinoline and amine-substituted phenyl ring and with the phosphate backbone through anion-π interactions with the benzothiazole ring. The designed molecule 4a has interesting photophysical properties, cell permeability, and biocompatibility with minimal cytotoxicity. Fluorescence imaging studies in live HeLa cells showed that probe 4a binds to the transient population of the DNA G-quadruplex in the nucleus and RNA quadruplexes in the cytoplasm. In brief, G-quadruplex NIR fluorescent probe 4a with a higher signal/noise ratio has significant potential for cellular imaging studies and thus opens avenues to decipher the biological pathways for better understanding of G-quadruplex biology.

Rational design of an ultrabright quinolinium-fused rhodamine turn-on fluorescent probe for highly sensitive detection of SO2 derivatives: Applications in food safety and bioimaging

Sulfur dioxide (SO2) is an essential signaling molecule involved in various physiological processes within living organisms. Bisulfite (HSO3−) possesses antioxidant, antimicrobial, and preservative properties, making it a common food additive. However, elevated levels of SO2 or excessive HSO3− intake can lead to a range of diseases, highlighting the importance of detecting SO2 and its derivatives (HSO3−/SO32−). This study presents a quinolinium-fused rhodamine fluorogenic probe (RQB-R) for ultrafast, highly selective, and sensitive detection of HSO3−. The probe operates via a dual-response mechanism, exhibiting a visible color change and a transition from nonemissive to intense red fluorescence upon interaction with HSO3−. The detection mechanism involves a 1,4-nucleophilic addition reaction of HSO3− at the 4-position of the quinolinium unit, which bypasses the photoinduced electron-transfer fluorescence quenching pathway and activates the intramolecular charge transfer mechanism, thereby enhancing fluorescence emission. Practical applications of the RQB-R probe include rapid quantification of HSO3− levels in sugar samples and integration into smartphone-assisted detection platforms. This method demonstrates excellent biocompatibility and enables visualization of both exogenous and endogenous HSO3− within MCF-7 cells, with a specific focus on targeting mitochondria.

Polysiloxane-Based Fluorescent Probes for Visualizing pH and Thiocyanate during Mitochondrial Autophagy.

Mitochondrial autophagy, known as mitophagy, is a vital cellular process that involves the selective degradation of damaged or dysfunctional mitochondria through autophagy, which is critical to the functional integrity of the entire mitochondrial network and determines the survival and death of cells. An abnormal pH may lead to an imbalance in mitochondrial homeostasis and the occurrence of mitochondrial autophagic acidification and dysfunction. SCN- is also an important anion in cellular metabolism, and its abnormal concentration may lead to mitochondrial damage. However, so far, there are few reports on the simultaneous realization of pH and SCN- detection in mitochondria. Therefore, to complement the blank in this area, we developed the polysiloxane-based fluorescent probe P0-CMN that is capable of simultaneously visualizing pH and SCN- fluctuation levels in mitochondria. The probe P0-CMN has the desired mitochondrial-targeting properties and sensitivity to pH and SCN-. It is able to simultaneously monitor pH and SCN- changes in mitochondria in a dual-channel mode. In addition, probe P0-CMN can visualize pH changes during mitochondrial autophagy. This work provides an effective strategy for the design of dual-responsive fluorescent probes and further broadens the application of polysiloxane fluorescent materials.

Cyano-determined mercury (II) ion selective fluorescence assay over polymeric carbon nitride

Selective response is the key index to evaluate the performance of polymeric carbon nitride (PCN)-based heavy metal ion fluorescence sensors. Herein, to explore the role of cyano groups on selectivity, four kinds of PCN, including PCN-Cl, PCN-Ac, PCN-B and PCN-K were prepared by the molten salt method of sodium chloride and sodium acetate, the reduction method of sodium borohydride and the etching method of potassium hydroxide, respectively. These PCNs exhibited different surface cyano characteristics, but all of them had significant blue emission under ultraviolet excitation. It is proved that the assistant of sodium chloride or potassium hydroxide is an effective method to prepare PCNs with abundant surface cyano group. A series of fluorescence quenching experiments of metal ions showed that the cyano-rich degree of PCN is closely related to its selective response to mercury (II) ions. PCN-Cl and PCN-K emerged good selective quenching of mercury (II) ions, which may be related to the soft acid-soft base strong interaction between mercury (II) ions and cyano groups. Both PCN-Cl and PCN-K fluorescent probes for mercury (II) ions had a linear range of 5 ∼ 50 μmol L−1, and PCN-Cl exhibited a lower detection limit of 0.38 μmol L−1. This work confirmed the selective fluorescence response of cyano-rich PCN to mercury (II) ions, proposed the mechanism of selective fluorescence quenching response of mercury (II) ions, and provided a new idea for the design of efficient and accurate PCN-based fluorescence probes.

Design and synthesis of a TICT-based red-emissive fluorescent probe for the rapid and selective detection of HSA in human biofluids and live cell imaging.

Here, we report the design and synthesis of a D⋯π⋯A-based fluorescent probe, (E)-4-(4-(dibutylamine)-2-hydroxystyryl)-1-methylquinolin-1-ium (DHMQ), which is nonfluorescent in ∼100% PBS buffer medium due to a twisted intra molecular charge transfer (TICT) phenomenon and it becomes highly fluorescent (∼149 fold) in the presence of human serum albumin (HSA), owing to the restriction of its intramolecular free rotation inside the hydrophobic binding cavity of HSA. The site-selective fluorescence displacement assay and molecular docking studies clearly reveal that DHMQ selectively binds at subdomain IB of HSA. The 3σ/slope method was adopted to determine the limit of detection (LOD) value, which was as low as 2.39 nM in ∼100% PBS medium, indicating its high sensitivity towards HSA. The low dissociation constant value [Kd = (1.066 ± 0.017) μM] suggests a strong complexation between the DHMQ and HSA. Importantly, it has been demonstrated that DHMQ is capable of detecting HSA in real human serum and urine samples and was found to be suitable for live cell imaging of HSA.

A novel dual-site fluorescent probe for the one-step detection of Cys and SO2 in living cells

BackgroundCysteine (Cys) is the major intracellular thiol and plays a key role in human pathology. Furthermore, endogenous sulfur dioxide (SO2) is produced in mammals. Abnormal levels of SO2 are commonly associated with a variety of respiratory, cardiovascular, and neurological diseases. Therefore, given their important role in life activities, it is significant to construct a fluorescent probe that can detection between Cys and SO2. ResultsWe have designed and synthesized a two-site fluorescent probe CUM with coumarin derivative and benzaldehyde molecules, which can detect and differentiate between Cys and SO2 through dual excitation wavelengths. Its carbon-carbon double bond reacts with Cys and undergoes a nucleophilic reaction, emitting green fluorescence at 520 nm, while SO32− reacts with benzaldehyde molecules in the probe CUM and undergoes a blue fluorescence at 460 nm. SO32− reacts with the benzaldehyde molecule of probe CUM and fluoresces blue at 460 nm. Thus, the probe CUM with two reaction sites can distinguish between Cys and SO2 and shows good selectivity and fast reaction speed. In addition, we successfully utilized probe CUM to image Cys and SO2 in human breast cancer cells (MDA-MB-231). SignificanceThis work provides an effective method for the molecular design of coumarin-based fluorescent probes. Probe CUM as a promising and reliable tool for the meticulous discrimination and quantification of Cys and SO2 in diverse biological matrices, thereby opening up new avenues for various biological systems.

‘Turn On’ fluorescent imine linked 1,2,3-triazole based chemosensor for detection of mercuric ions

The exorbitant use or erroneous management of metal ions has been incurred with catastrophic pollution of our ecological system, particularly in soil and water bodies, resulting in significant repercussions. Hence, precise identification and analysis of heavy-metal ions in intricate hydrological environments is imperative. Therefore, an imine conjugated 1,2,3-triazole derivative VPT was synthesized, characterized via FTIR, NMR (1H &13C), LCMS spectroscopic techniques and was employed for the exclusive identification of Hg2+ ions in the presence of various other metal ions via UV–Visible and fluorescence spectroscopy. The sensor VPT exhibited binding constant (Ka) value of 2.93 × 104 M−1 and detection limit value of 0.5 × 10−9 M in the ACN:H2O::4:1 medium. The stoichiometric ratio was determined using Job’s plot, which revealed a (1:1) ratio of VPT with Hg2+ ions. In addition, time-dependent, temperature-dependent, and pH titration studies were conducted via U.V-Visible spectroscopy to expand the sensor’s applicability. The viable configurations of probe VPT and its 1:1 stoichiometric with Hg2+ ion were optimized using the B3LYP/6-311G++(d,p)/LANL2DZ levels of theory in the Gaussian 09 program respectively and this semi-empirical quantum mechanics approach demonstrated a reduction in the energy gap confirming the stable formation of [VPT-Hg2+] complex. These insights facilitate the design offluorescent probe VPT for detecting Hg2+ in both environmental and biological context.

A turn-off xanthene-based fluorescent probe for detection of cysteine and its practical application in bioimaging and food samples

BackgroundCys, as an essential amino acid that can be ingested from daily food, plays an important role in maintaining the oxidative balance in cells. Abnormal Cys levels in organisms will lead to various diseases. Therefore, it is of great significance to construct a fluorescent probe that can detect Cys levels in food and biological systems. ResultsHere, a turn-off type probe TA had been successfully synthesized, which attached diethylamine as the strong electron donor, acrylate as the weak electron donor, and xanthene as the π-bridge. TA showed wonderful selectivity, low detection limit, good photostability and well live-cell compatibility for Cys by reducing acrylate group to hydroxyl group of TAOH. The reaction mechanism was demonstrated by 1H NMR, ESI-MS spectra, pH-dependent response experiments, and DFT calculations. Importantly, the reason why TAOH exhibited no fluorescence was the disappearance of the ICT effect in the molecule due to the dominant existence of spirocyclic state of TAOH. In addition, the probe can be used not only for the imaging detection of Cys in A549 cells and zebrafish, but also for the detection of Cys levels in food samples. SignificanceThis work provides a new idea for the design of Cys fluorescent probe, which may be beneficial to the comprehension of the potential mechanism of novel fluorescent probe.

Reversible Fluorescent Probes for Dynamic Imaging of Liver Ischemia-Reperfusion Injury.

ConspectusHepatic ischemia-reperfusion injury (HIRI) is an inevitable complication of clinical surgeries such as liver resection or transplantation, often resulting in postoperative liver dysfunction, hepatic failure in up to 13% of postresection patients, and early graft failure in 11-18% of liver transplantation patients. HIRI involves a series of biochemical events triggered by abnormal alterations in multiple biomarkers, characterized by short lifespans, dynamic changes, subcellular regional distribution, and multicollaborative regulation. However, traditional diagnosis, including serology, imaging, and liver puncture biopsy, suffers from low sensitivity, poor resolution, and hysteresis, which hinder effective monitoring of HIRI markers. Thus, to address the unique properties of HIRI markers, there is a pressing demand for developing novel detection strategies that are highly selective, transiently responsive, dynamically reversible, subcellular organelle-targeted, and capable of simultaneous multicomponent analysis.Optical probe-based fluorescence imaging is a powerful tool for real-time monitoring of biomarkers with the advantages of high sensitivity, noninvasiveness, rapid analysis, and high-fidelity acquisition of spatiotemporal information on signaling molecules compared with conventional methods. Moreover, with the growing demand for continuous monitoring of biomarkers, probes with reversible detection features are receiving more and more attention. Importantly, reversible probes can not only monitor fluctuations in marker concentrations but also distinguish between transient bursts of markers during physiological events and long-term sustained increases in pathological marker levels. This can effectively avoid false-positive test results, and in addition, reversible probes can be reutilized with green and economical features. Therefore, our team has employed various effective methods to design reversible optical probes for HIRI. We proposed reversible recognition strategies based on specific reactions or interactions to detect dynamic changes in markers. Given the biomarkers' unique signaling in subcellular organelles and the synergistic regulatory properties of multiple markers for HIRI, bifunctional reversible detection strategies are exploited, including organelle-targeted reversible and multicomponent simultaneous detection. With these strategies, we have tailored a variety of high-fidelity fluorescent probes for a series of HIRI markers, including reactive oxygen/nitrogen species (O2•- and ONOO-), ATP, protein (Keap1), mitochondrial DNA, etc. Utilizing the probes, the in situ dynamic imaging detection of the HIRI markers was successfully achieved. While performing the precise examination of the earlier occurrence of HIRI disease and visualizing the real-time monitoring of the disease process, we have also further elucidated the HIRI-associated signaling pathways. It is envisioned that our summarized work will inspire the design of future reversible fluorescent probes and help to improve the clinical diagnosis and therapeutic efficiency of these diseases.

Novel benzothiazole-based fluorescent probe for efficient detection of Cu2+/S2− and Zn2+ and its applicability in cell imaging

BackgroundIn recent years, environmental pollution has been increasing due to the excessive emission of toxic ions, which has caused serious harm to human health and ecological environment. There are various methods for detecting Cu2+, S2− and Zn2+, but the traditional ion detection methods have obvious disadvantages, such as poor selectivity and long detection time. Therefore, it is still crucial to develop simple, efficient and rapid detection methods. ResultsA fluorescent probe based on benzothiazole, (E)-N'-(3-(benzo[d]thiazol-2-yl)-2-hydroxy-5-methylbenzylidene)-3,4,5-tris(benzyloxy)benzohydrazide (BT), was designed and synthesized. It was characterized using ESI-MS, 1H NMR, and 13C NMR. BT can be used as a chemosensor to detect Cu2+, S2− and Zn2+ in CH3CN/H2O (7:3, v/v, pH = 7.4, HEPES buffer: 0.1 M), with detection limits of 0.301 μM, 0.017 μM, and 0.535 μM, respectively. At an excitation wavelength of 320 nm, BT exhibits an “on-off-on” response to Cu2+/S2− and enhanced fluorescence response to Zn2+, with a change in fluorescence color from orange to green. The coordination ratio of ions to the probe was determined to be 1:1 through Job's plot and hydrogen spectral titration. The recognition mechanism was discussed in conjunction with theoretical calculations. Furthermore, the probe has been successfully used in test strips and medical swabs colorimetry, as well as live cell imaging. SignificanceThe probe BT lays the foundation for the design and synthesis of multifunctional fluorescent probes. As a portable detection method, probe BT was used to detect Cu2+, S2− and Zn2+ on strips. Furthermore, the probe was applied to biological cells to detect target ions with low cytotoxicity and excellent cell permeability. This indicating that it can be used as a potential candidate for tracking Cu2+ and S2− in clinical diagnostics and biological systems.

Sensing mechanism of the benzo-bodipy based fluorescent probe for Hypochlorous acid detection: Invalidity of photoinduced electron transfer

In vivo real-time detection of hypochlorous acid (HClO) in biological systems plays a crucial role in diagnosing immune-related diseases. Experimentally, a benzo-bodipy probe based on the photo-induced electron transfer (PeT) sensing mechanism has been developed for live fluorescence imaging. However, there have been no theoretical studies conducted to substantiate the precision of the sensing mechanism. This paper employs density functional theory (DFT) and time-dependent density functional theory (TDDFT) methods to investigate the fluorescence detection mechanism of benzo-bodipy derivatives (BBy-T and BBy-TO), proposing a detection approach based on dark nπ* state quenching. The study reveals that the fluorescence quenching mechanism of BBy-T is primarily regulated by a thiomorpholine moiety, involving a dark nπ* state transition non-radiatively. Furthermore, this paper explains the fluorescence enhancement observed in BBy-TO. Theoretical investigations demonstrate, based on frontier molecular orbitals (FMOs) and hole-electron analysis, that the fluorescence enhancement for BBy-TO is not governed by the previously proposed intramolecular charge transfer (ICT) mechanism in experiments but rather follows a locally excited (LE) ππ* pattern. This work offers new insights for the design of novel fluorescence probes based on bodipy and benzo derivatives, expanding the understanding of their fluorescence properties.

A new insight into the luminescence mechanism of a FRET-ICT-based ratiometric and colorimetric pH probe

Recently, a ratio fluorescent probe molecule BP was designed and synthesized to detect pH in living cells. However, few have conducted in-depth theoretical exploration of the detailed transfer process. In this paper, the luminescence mechanisms of BP and BPH were studied with the results of geometric optimization structures and molecular orbitals at the CAM-B3LYP/TZVP computational level. It was found that a fluorescence resonance energy transfer (FRET) process occurs in the excited state of BP molecules, and the final fluorescence emission is attributed to the intramolecular charge transfer (ICT) of the acceptor styryl pyridine ring. Meanwhile, the fluorescence emission of BPH molecules is due to the ICT process caused by the local excitation process of the acceptor group protonated styryl pyridine ring itself. Therefore, this luminescence is a FRET “ON-OFF” combined ICT process, not like the FRET “OFF-ON” mechanism which the experiment suspected. Our work illustrates a specific FRET combined ICT process of probes BP and BPH from a theoretical perspective which could provide theoretical guidance for the future design of similar fluorescent probes.

Visualizing detection of cysteine level under LPS-induced oxidative stress process using benzopyrylium-based mitochondrial-targeted NIR fluorescent probe

As an important scavenger of reactive oxygen species, cysteine (Cys) plays a crucial role in mitochondrial oxidative stress. Abnormal intracellular Cys concentration under oxidative stress leads to various diseases. Cys is also widely used in the food industry. Many vegetables and fruits are rich in Cys. Abnormal levels of Cys in the body can lead to a variety of diseases. Therefore, the design of fluorescent probe that can detect Cys quickly is particularly important in living organisms. In this work, a novel turn-on mitochondrial-targeted NIR fluorescent probe (SXNU-6) was designed for the high sensitivity and specificity of Cys based on benzopyrylium. Among them, acrylate group acted as recognition site and fluorescence quencher. With the addition of Cys, the fluorescence of SXNU-6 was gradually enhanced at 660 nm. The probe had good selectivity and high sensitivity, which could realize the detection of Cys in food samples (onion, garlic, tomato, lettuce, apple, pear, milk and bread). Also, SXNU-6 could detect both endogenous and exogenous Cys under LPS-induced oxidative stress process. This work provide a powerful tool for detecting Cys in food samples and biological systems.

Rational design of cascade reaction-assisted trinal-site fluorescent probe for simultaneous discrimination of Cys, Hcy, GSH, and H2S in living cells and zebrafish

The reactive sulfur species (RSS), in particular Cys, Hcy, GSH, and H2S, play an essential role in lots of critical physiological and pathological processes. To elucidate their complex interrelationships in signaling and metabolic pathways, suitable molecular tools to simultaneously distinguish Cys, Hcy, GSH, and H2S are urgently needed. Herein, we developed a novel cascade reaction-assisted trinal-site fluorescent probe BCS by connecting three functional fragments for simultaneously distinguishing Cys, Hcy, GSH, and H2S. The probe BCS exhibited red fluorescence in the absence of the four biological RSS. However, with the attendance of Cys, Hcy, GSH, and H2S, the probe showed different fluorescence patterns, including blue, blue/red, green/red, and no fluorescence, respectively. More importantly, based on the different RSS-induced fluorescence patterns and multi-color fluorescence imaging, probe BCS was used to discriminate and visualize all four biological RSS in both HepG2 cells and zebrafish. It is expected that probe BCS will have applications in the diagnosis and research of RSS-related diseases in the future.

Iterative Design of Near-Infrared Fluorescent Probes for Early Diagnosis of Alzheimer's Disease by Targeting Aβ Oligomers.

Amyloid-β oligomers (AβOs), crucial toxic proteins in early Alzheimer's disease (AD), precede the formation of Aβ plaques and cognitive impairment. In this context, we present our iterative process for developing novel near-infrared fluorescent (NIRF) probes specifically targeting AβOs, aimed at early AD diagnosis. An initial screening identified compound 18 as being highly selective for AβOs. Subsequent analysis revealed that compound 20 improved serum stability while retaining affinity for AβOs. The most promising iteration, compound 37, demonstrated exceptional qualities: a high affinity for AβOs, emission in the near-infrared region, and good biocompatibility. Significantly, ex vivo double staining indicated that compound 37 detected AβOs in AD mouse brain and in vivo imaging experiments showed that compound 37 could differentiate between 4-month-old AD mice and age-matched wild-type mice. Therefore, compound 37 has emerged as a valuable NIRF probe for early detection of AD and a useful tool in exploring AD's pathological mechanisms.