Yuan-Qiang Sun

Simultaneous Fluorescence Sensing of Cys and GSH from Different Emission Channels

A chlorinated coumarin-hemicyanine dye with three potential reaction sites was exploited as fluorescent probe for biothiols. The Cys-induced substitution-rearrangement-cyclization, Hcy-induced substitution-rearrangement, and GSH-induced substitution-cyclizatioin cascades lead to the corresponding amino-coumarin, amino-coumarin-hemicyanine, thiol-coumarin with distinct photophysical properties, enabling Cys and GSH to be selectively detected from different emission channels at two different excitation wavelengths.

Simultaneous fluorescent imaging of Cys/Hcy and GSH from different emission channels

A 4-methoxythiophenol-substituted pyronin dye 1 was exploited as reaction-type fluorescent probe for biothiols Cys/Hcy and GSH. The probe itself is nonfluorescent due to the photoinduced electron transfer (PET) process. The Cys (or Hcy)-induced substitution–rearrangement cascade reaction and GSH-induced substitution reaction with the probe lead to the corresponding aminopyronin and thiopyronin dyes with distinct photophysical properties, enabling Cys/Hcy and GSH to be detected from visible and near-infrared (NIR) emission channels, respectively, in pure PB buffer with relatively fast kinetics and obvious fluorescence turn-on response. Assisted by laser scanning confocal microscope, we also demonstrated that probe 1 could simultaneously sense Cys/Hcy and GSH in B16 cells in multicolor imaging.

A Ratiometric Fluorescent Probe for Biological Signaling Molecule H2S: Fast Response and High Selectivity

Traditionally, hydrogen sulfide (H2S) was considered to be a toxic gas. Later, it was reported that the typical concentration of H2S in blood is in the range 10–100 mm, suggesting its connection with biological functions. Indeed, recent studies regard H2S as the third gaseous transmitter besides nitric oxide (NO) and carbon monoxide (CO). For example, H2S has been recognized to mediate a wide range of physiological effects, including regulation of cell growth, vasodilation, and angiogenesis, mediation of neurotransmission, inhibition of insulin signaling, and regulation of inflammation. Moreover, it acts as an antioxidant or scavenger of reactive oxygen species (ROS). Furthermore, studies have shown that its deregulation is correlated with the symptoms of Alzheimer s disease, Down s syndrome, diabetes, and liver cirrhosis. Although H2S has been recognized to be linked to various physiological and pathological functions, many of its underlying molecular events remain unknown. Therefore, efficient methods to sensitively and selectively detect H2S in living systems are urgently required. Fluorescence spectroscopy is a powerful tool for sensing and imaging trace amounts of samples because of its simplicity, sensitivity, real-time imaging, and especially its nondestructive detection of target biomolecules in live cells or tissues. Undoubtedly, our understanding of molecular function in cell biology has significantly benefited from advancements of fluorescent probes. Generally, the design of fluorescent probes for detection of H2S in living systems involves several substantial challenges: one is to attain sufficient selectivity over other biothiols, including reduced glutathione (GSH, present at levels of about 1–10 mm) and lcysteine (l-Cys, about 100 mm); the other is to achieve sufficient sensitivity, because the physiologically relevant H2S concentration has been reported to range from nanoto millimolar levels; and the third challenge is fast response under mild condition, because H2S is metabolized rapidly. Previously reported fluorescent probes for H2S are mainly focused on in vitro assay systems. Recently, fluorescent probes that can potentially be used for H2S detection in living systems have become available, and the corresponding design strategies are based on several significant characteristic properties of H2S, namely dual nucleophilicity, [8–10]

A fluorescent turn-on probe for bisulfite based on hydrogen bond-inhibited CN isomerization mechanism

A fluorescence turn-on probe for bisulfite has been developed by taking advantage of the specific reaction of bisulfite and aldehyde in combination with the hydrogen bond inhibited C=N isomerization mechanism. The practical value of this selective and sensitive fluorescent probe was confirmed by its application to detection of bisulfite in granulated sugar.

Colorimetric Detection of Thiols Using a Chromene Molecule

A new thiol-containing colorimetric probe has been developed by using a chromene derivative, 7-nitro-2,3-dihydro-1H-cyclopenta[b]chromen-1-one (1). The molecule exhibited high selectivity and sensitivity for detecting thiol species as cysteine, homocysteine, and glutathione in aqueous solution through a rapid visual color change from colorless to yellow.

A Mitochondria-Targetable Fluorescent Probe for Dual-Channel NO Imaging Assisted by Intracellular Cysteine and Glutathione

A mitochondria-specific fluorescent probe for NO (1) was synthesized by the direct conjugation of a pyronin dye with one of the amino groups of o-phenylenediamino (OPD). The probe could selectively detect NO over dehydroascorbic acid (DHA), ascorbic acid (AA), and methylglyoxal (MGO) as well as the reactive oxygen/nitrogen species (ROS/RNS) with the significant off-on response due to the production of a red-emission triazole 2. In the presence of cysteine/glutathione (Cys/GSH), 2 could be further transformed into a green-emission aminopyronin 4 and a red-emission thiopyronin 5, respectively. Assisted by intracellular Cys and GSH, the probe demonstrated its potential to monitor mitochondrial NO in a dual-channel mode.

Construction of a Selective Fluorescent Probe for GSH Based on a Chloro-Functionalized Coumarin-enone Dye Platform†

Glutathione (GSH), the most abundant intracellular biothiol, protects cellular components from damage caused by free radicals and reactive oxygen species (ROS), and plays a crucial role in human pathologies. A fluorescent probe that can selectively sense intracellular GSH would be very valuable for understanding of its biological functions and mechanisms of diseases. In this work, a 3,4-dimethoxythiophenol-substituted coumarin-enone was exploited as a reaction-type fluorescent probe for GSH based on a chloro-functionalized coumarin-enone platform. In the probe, the 3,4-dimethoxythiophenol group functions not only as a fluorescence quencher through photoinduced electron transfer (PET) to ensure a low background fluorescence, but also as a reactive site for biothiols. The probe displays a dramatic fluorescence turn-on response toward GSH with the long-wavelength emission (600 nm) and significant Stokes shift (100 nm). The selectivity of the probe toward GSH over cysteine (Cys), homocysteine (Hcy), and other amino acids was demonstrated. Assisted by laser-scanning confocal microscopy, we have demonstrated that the probe could specifically sense GSH over Cys/Hcy in human renal cell carcinoma SiHa cells.

Naked-eye determination of oxalate anion in aqueous solution with copper ion and pyrocatechol violet

A novel strategy for the determination of oxalate anions was successfully established using a copper ion and pyrocatechol violet (PV) ensemble. The sensor ensemble can discriminate oxalate over other common anions including F(-), Cl(-), I(-), Br(-), HPO(4)(2-), PO(4)(3-), AcO(-), CO(3)(2-), SO(4)(2-), ClO(4)(-), P(2)O(7)(4-), S(2-) (deposited by Ag(+)), CN(-) (shielded by Fe(3+)) and can detect oxalate at low microgram levels in quasi-physiological aqueous solutions. The detection of the oxalate anion gives rise to a rapid observable visual color change from blue to yellow.

A carboxylic acid-functionalized coumarin-hemicyanine fluorescent dye and its application to construct a fluorescent probe for selective detection of cysteine over homocysteine and glutathione

A carboxylic acid-functionalized coumarin-hemicyanine near-infrared (NIR) dye 1 was exploited, which possesses good water solubility (more than 50 μM) and favorable photophysical properties, especially a large Stokes shift (around 90 nm), and has been proved to be a suitable imaging agent for targeting mitochondria. With the dye platform, fluorescent probe 2, a thioester derivative of 1, was constructed for biothiols. Probe 2 can react with cysteine (Cys) via the native-chemical-ligation (NCL) and cyclization cascade reactions to lead to coumarin 2-Cys. However, the reaction of 2 with homocysteine (Hcy) or glutathione (GSH) only stays at the stage of the initial transthioesterification reaction, producing coumarin-hemicyanines 2-Hcy or 2-GSH, due to an electrostatic interaction in 2-Hcy and an unstable macrocyclic transition state in 2-GSH, both inhibiting their subsequent S,N-acyl shift. Given the distinct photophysical properties between 2-Cys and 2-Hcy (or 2-GSH), probe 2 could highly selectively discriminate Cys from Hcy/GSH. Even in the presence of Hcy or GSH, probe 2 still works well for Cys due to the reversible transthioesterification and the irreversible S,N-acyl shift in the NCL reaction. The cell imaging assays revealed that probe 2 is cell permeable and could selectively image Cys in living cells.

D-Luciferin Analogues: a Multicolor Toolbox for Bioluminescence Imaging†

Colorful mixture: Three types of luciferin analogues, that is, alkylaminoluciferins, aminoselenoluciferin, and luciferins with a benzimidazole scaffold, have been reported. These analogues show excellent bioluminescent properties and great potential in bioluminescence imaging.