Highly selective two-photon fluorescent off-on probes for imaging tyrosinase activity in living cells and tissues.

A coumarin-based two-photon (TP) fluorescent off-on probe has been developed for detecting tyrosinase activity. High selectivity, sensitivity and biocompatibility enable the probes to successfully image tyrosinase activity in live cells and tissues using TP microscopy.

Rational Design of a Highly Selective Near-Infrared Two-Photon Fluorogenic Probe for Imaging Orthotopic Hepatocellular Carcinoma Chemotherapy.

Selective fluorescence imaging of biomarkers in vivo and in situ for evaluating orthotopic hepatocellular carcinoma (HCC) chemotherapy remains a great challenge due to current imaging agents suffering from the potential interferences of other hydrolases. Herein, we observed that carbamate unit showed a high selectivity toward the HCC-related biomarker carboxylesterase (CE) for evaluation of treatment. A near-infrared two-photon fluorescent probe was developed to not only specially image CE activity in vivo and in situ but also target orthotopic liver tumor after systemic administration. The in vivo signals of the probe correlate well with tumor apoptosis, making it possible to evaluate the status of treatment. The probe enables the imaging of CE activity in situ with a high-resolution three-dimensional view for the first time. This study may promote advances in optical imaging approaches for precise imaging-guided diagnosis of HCC in situ and its evaluation of treatment.

Highly Efficient Aggregation-Induced Red-Emissive Organic Thermally Activated Delayed Fluorescence Materials with Prolonged Fluorescence Lifetime for Time-Resolved Luminescence Bioimaging.

Organic thermally activated delayed fluorescence (TADF) materials are emerging as potential candidates for time-resolved fluorescence imaging in biological systems. However, the development of purely organic TADF materials with bright aggregated-state emissions in the red/near-infrared (NIR) region remains challenging. Here, we report three donor-acceptor-type TADF molecules as promising candidates for time-resolved fluorescence imaging, which are engineered by direct connection of electron-donating moieties (phenoxazine or phenothiazine) and an electron-acceptor 1,8-naphthalimide (NI). Theoretically and experimentally, we elucidate that three TADF materials possessed remarkably small ΔEST to promote the occurrence of reverse intersystem crossing (RISC). Moreover, they all exhibit aggregation-induced red emissions and long delayed fluorescence lifetimes without the influence of molecular oxygen. More importantly, these long-lived and biocompatible TADF materials, especially the phenoxazine-substituted NI fluorophores, show great potential for high-contrast fluorescence lifetime imaging in living cells. This study provides further a molecular design strategy for purely organic TADF materials and expands the versatile biological application of long-lived fluorescence research in time-resolved luminescence imaging.

Design and synthesis of efficient heavy-atom-free photosensitizers for photodynamic therapy of cancer.

Novel thiocarbonyl derivatives (NIS and CRNS) with excellent ROS generation abilities are synthesized and studied as potential photosensitizers for one- and two-photon excited photodynamic therapy. In particular, NIS-Me and CRNS display outstanding phototoxicity toward HeLa cells under two-photon excitation (800 nm) with negligible dark toxicity.

Fine-tuning the electronic structure of heavy-atom-free BODIPY photosensitizers for fluorescence imaging and mitochondria-targeted photodynamic therapy.

Theranostics that combines both diagnosis and therapy into a single platform has recently emerged as a promising biomedical approach for cancer treatment; however, the development of efficient theranostic agents with excellent optical properties remains a challenge. Here, we report novel mitochondria-targeting BODIPY photosensitizers (R-BODs) that possess considerable singlet oxygen generation capabilities and good fluorescence properties for imaging-guided photodynamic therapy (PDT). The incorporation of sulfur atoms into the π-conjugated skeleton of BODIPY along with the introduction of different functional groups at the meso-position of the BODIPY core is essential for tuning the photophysical and photosensitizing properties. Notably, the MeOPh-substituted thiophene-fused BODIPY (MeO-BOD, R = p-methoxyphenyl) displayed the highest singlet oxygen generation capability (Φ Δ ≈ 0.85 in air-saturated acetonitrile) and a moderate fluorescence quantum yield (Φ f = 17.11). Furthermore, MeO-BOD showed good biocompatibility, low dark toxicity and superior fluorescence imaging properties in living cells. More importantly, the PDT efficacy of mitochondria-specific anchoring of MeO-BOD was remarkably amplified with an extremely low half-maximal inhibitory concentration (IC50) value of 95 nM. We believe that the incorporation of an electron-donating group at the meso-position of the thiophene-fused BODIPY platform may be an effective approach for developing theranostic agents for precision cancer therapy.

A molecular approach to rationally constructing specific fluorogenic substrates for the detection of acetylcholinesterase activity in live cells, mice brains and tissues.

Acetylcholinesterase (AChE) is an extremely critical hydrolase tightly associated with neurological diseases. Currently, developing specific substrates for imaging AChE activity still remains a great challenge due to the interference from butyrylcholinesterase (BChE) and carboxylesterase (CE). Herein, we propose an approach to designing specific substrates for AChE detection by combining dimethylcarbamate choline with a self-immolative scaffold. The representative P10 can effectively eliminate the interference from CE and BChE. The high specificity of P10 has been proved via imaging AChE activity in cells. Moreover, P10 can also be used to successfully map AChE activity in different regions of a normal mouse brain, which may provide important data for AChE evaluation in clinical studies. Such a rational and effective approach can also provide a solid basis for designing probes with different properties to study AChE in biosystems and another way to design specific substrates for other enzymes.

An Emerging Molecular Design Approach to Heavy-Atom-Free Photosensitizers for Enhanced Photodynamic Therapy under Hypoxia.

A novel strategy for designing highly efficient and activatable photosensitizers that can effectively generate reactive oxygen species (ROS) under both normoxia and hypoxia is proposed. Replacing both oxygen atoms in conventional naphthalimides (RNI-O) with sulfur atoms led to dramatic changes in the photophysical properties. The remarkable fluorescence quenching (ΦPL ≈ 0) of the resulting thionaphthalimides (RNI-S) suggested that the intersystem crossing from the singlet excited state to the reactive triplet state was enhanced by the sulfur substitution. Surprisingly, the singlet oxygen quantum yield of RNI-S gradually increased with increasing electron-donating ability of the 4-R substituents (MANI-S, ΦΔ ≈ 1.00, in air-saturated acetonitrile). Theoretical studies revealed that small singlet-triplet energy gaps and large spin-orbit coupling could be responsible for the efficient population of the triplet state of RNI-S. In particular, the ROS generation ability of MANI-S was suppressed under physiological conditions due to their self-assembly and was significantly recovered in cancer cells. More importantly, cellular experiments showed that MANI-S still produced a considerable amount of ROS even under severely hypoxic conditions (1% O2) through a type-I mechanism.

Coumarin/fluorescein-fused fluorescent dyes for rapidly monitoring mitochondrial pH changes in living cells.

On base of the good optical properties of coumarin and fluorescein, we designed and synthesized two coumarin/fluorescein-fused fluorescent dyes (CF dyes), which enlarged the emission wavelength and increased the Stokes shift of fluorescein moiety. The corresponding optical properties of CF dyes were investigated in detail. CF dyes could easily introduce other groups to design different functional molecules. CF dyes also exhibited rapid and sensitive responses to pH values in the range of 4.0-7.4 through the characterization of absorption and fluorescence spectra in buffer solution. More importantly, CF ethyl ester dye (CFE dye) not only showed good cell membrane permeability and low cytotoxicity, but also had the ability to rapidly monitor mitochondrial pH changes in living cells.

Colorimetric and fluorescent detection of hydrazine with high sensitivity and excellent selectivity.

It is critical to develop probes for rapid, selective, and sensitive detection of the highly toxic hydrazine in both environmental and biological science. In this work, under mild condition, a novel colorimetric and off-on fluorescent probe was synthesized for rapid recognition of hydrazine with excellent selectivity over other various species including some biological species, metal ions and anions. The limit of quantification (LOQ) value was 1.5×10-4M-3.2×10-3M (colorimetric method) and 1.5×10-4M-3.2×10-3M (fluorescent method) with as low as detection limit of 46.2µM.