ABSTRACT
BACKGROUND: Hypochlorous acid (HClO) is an important biomarker for inflammation, cardiovascular disease, and even cancer. It is of great significance to accurately monitor and quantitatively analyze the fluctuations of HClO to better understand their physiological functions. Traditional HClO detection methods such as high-performance liquid chromatography (HPLC), and mass spectrometry are preferred, but are costly and unsuitable in vivo. Near-infrared (NIR) fluorescence imaging has the advantages of high sensitivity, high temporal and spatial resolutions, minimal autofluorescence, and deep tissue penetration, which facilitates its application in biological systems. Therefore, the development of sensitivity and simple NIR fluorescence monitoring HClO methods in vivo and in vitro is essential and desirable. RESULTS: Herein, we present a NIR probe NOF3 by integrating the rhodamine scaffold and HClO-triggered moiety for the real-time detection of HClO in vitro and in vivo. NOF3 reacts with the HClO and releases the NOF-OH fluorophore of emitted signals at 730 nm, which is in the NIR region. The designed probe detected concentrations of HClO ranging from 0 to 17 µM with a low detection limit of 0.146 µM, presenting excellent sensitivity and selectivity toward HClO over other species. NOF3 manifests significantly turn-on NIR fluorescent signals in response to HClO concentration, which makes it favorable for monitoring dynamic HClO distribution in vivo. We exemplify NOF3 for the tracking of endogenously overexpressed HClO distribution in RAW 264.7 cells, and further realize real-time in vivo bioimaging of HClO activity in inflammation mice. SIGNIFICANCE: The facile NIR NOF3 probe was successfully applied to visualize endogenous and exogenous HClO in living cells and mice. This study provides not only an effective tool for spatial and temporal resolution HClO bioimaging in vivo but also possesses great potential for use in future research on HClO-related biology and pathology.
Subject(s)
Hypochlorous Acid , Xanthenes , Mice , Animals , Hypochlorous Acid/analysis , Rhodamines/chemistry , Fluorescent Dyes/chemistry , Inflammation/diagnostic imagingABSTRACT
Thiol-containing amino acids, cysteine (Cys) and homocysteine (Hcy), play crucial roles in the biosystem; their abnormal contents in the cells are linked to many diseases. Herein, we designed and synthesized a novel near-infrared (NIR) phosphorescent iridium(iii) complex-based probe (FNO1) that can detect Cys and Hcy in real-time in the biosystem. Due to the advantages of the iridium complex, the FNO1 probe had excellent chemical stability and photostability, high luminescence efficiency, and long luminescence lifetime. In addition, the probe showed a fast response, high sensitivity, and low cytotoxicity. As verified by high resolution mass spectra (HR-MS) and density functional theory (DFT) calculations, the detection was achieved through the addition of the α,ß-unsaturated ketone group in FNO1 by the nucleophilic thiol group in Cys and Hcy. Through time-resolved emission spectroscopy (TRES) and in the presence of a strongly fluorescent dye rhodamine B, the FNO1 probe could detect Cys and Hcy due to its long luminescence lifetime (260/197 ns). Finally, owing to its NIR-emitting properties, the FNO1 probe was successfully applied in the imaging of Cys and Hcy in living cells, zebrafish, and mice.
Subject(s)
Coordination Complexes/chemistry , Cysteine/analysis , Homocysteine/analysis , Luminescent Agents/chemistry , Animals , Coordination Complexes/chemical synthesis , Coordination Complexes/toxicity , Density Functional Theory , Humans , Iridium/chemistry , Iridium/toxicity , Luminescent Agents/chemical synthesis , Luminescent Agents/toxicity , Luminescent Measurements , MCF-7 Cells , Mice , Microscopy, Confocal , Models, Chemical , Rhodamines/chemistry , ZebrafishABSTRACT
Peroxynitrite (ONOO-), one of the reactive oxygen/nitrogen species (ROS/RNS) found in vivo, plays crucial roles in many physiological and pathological processes. The ability to selectively and sensitively determine ONOO-in vivo is important for the understanding of its biological roles. Thus, by utilizing the excellent chemical stability and photostability, high luminescence efficiency, and long luminescence lifetime of iridium complexes, we developed a novel near-infrared (NIR) phosphorescent iridium(iii) complex (FNO2) probe to detect ONOO- within seconds. The probe FNO2 showed better selectivity towards ONOO- over other interfering biomolecules, including O2- and ClO-. Moreover, it possessed a long luminescence lifetime, which enabled successful elimination of the interference from background fluorescence in vitro (simulated by Rhodamine B) in time-resolved emission spectra. Finally, in addition to its low cytotoxicity, the probe FNO2 showed emission wavelength in the NIR region and was able to specifically sense ONOO- induced in living cells and inflamed mouse models.
