A highly selective ratiometric fluorescent probe based on naphthalimide for detection and imaging of CYP1A1 in living cells and zebrafish.

Real-time monitoring of the cytochrome P450 1A1 (CYP1A1) activity in complex biological systems via a practical tool is highly sought after because of its significant role in the metabolism and bioactivation of various xenobiotics. Herein, according to slight differences in the 3D structure and substrate preference between CYP1A1 and its homologous CYP1A2, a series of novel ratiometric fluorescent probes were designed and synthesized using 1,8-naphthalimide because of its trait of naked-eye visualization and ratiometric fluorescence to achieve the detection of CYP1A1 in biological samples. Among these probes, NEiPN showed good water solubility, highly isoform selectivity and great sensitivity (LOD = 0.04874 nM) for CYP1A1 under simulated physiological conditions, which makes it favorable for monitoring CYP1A1 in vivo. Remarkably, NEiPN exhibited excellent reproducibility when it was used to detect the CYP1A1 content in human liver microsomes, which indicated that it has a great potential for quantifying the CYP1A1 content in real biological samples. Furthermore, NEiPN showed relatively low cytotoxicity and has been successfully applied in biological imaging in living cells and zebrafish. These findings indicate that NEiPN is capable of real-time monitoring of the activity of endogenous CYP1A1, which could provide support for CYP1A1-associated pathological processes.

A novel probe for colorimetric and near-infrared fluorescence detection of cysteine in aqueous solution, cells and zebrafish.

Cysteine(Cys) is tightly related to physiological and pathological of human, and the imbalance of concentration of cysteine in the intracellular are associated with many diseases. Here, a novel NIR fluorescent probe TCF-Cys was designed and synthesized, and both the optimal excitation and emission wavelength of them were between 650 and 900 nm, that within the "optical window" of biological tissues. In aqueous solution, TCF-Cys, which with an acrylate extremity as a recognizing unit, exhibited excellent "turn-on" fluorescence response for Cys superior to other amino acids and thiols with a limit of detection of 0.1323 µM. Moreover, as an excellent naked-eye colorimetric indicator, TCF-Cys could effectively distinguishing the Cys, Hcy and GSH in aqueous solution through color change. Then, the response mechanism of TCF-Cys for Cys was revealed by TLC, 1H NMR, HPLC, HRMS and DFT calculation. Finally, TCF-Cys was successfully employed to fluorescence specifically map of exogenous and endogenous Cys in living cells and zebrafish with low toxicity.

A novel isophorone-based red-emitting/NIR probe for thiophenol and its application in real water sample and vivo.

Fast, highly selective and sensitive thiophenol probes are highly desirable in the field of bioimaging and environmental monitoring. For that, based on the mechanism that thiophenol can effectively cleave the sulfonamide bond selectively, we herein report a dicyanoisophorone-based Red-emitting/NIR probe for thiophenol detection. This probe had some desirable properties such as rapid response, high selectivity and sensitivity, remarkable large Stokes shift (181 nm), Red-emitting/NIR fluorescence region and low LOD value (80 nM, according to 3σ/s). Moreover, this novel Red-emitting/NIR probe can potentially be applied to the detection of thiophenols in real water samples quantitatively and fluorescent imaging in living cells and zebrafishes.

Synthesis of a selective ratiometric fluorescent probe based on Naphthalimide and its application in human cytochrome P450 1A.

Cytochrome P450s have brought considerable attention to researchers for their significant correlations with metabolic behaviors of procarcinogenic chemicals. To better understand the roles of CYP1A in biological and physiological systems, we developed a novel ratiometric fluorescence probe N-((2-hydroxyl ethoxy) ethyl)- 4-methoxy-1, 8-naphthalimide (NEMN) allowing for selectively and sensitively monitoring the target enzymes under physiological conditions and living cells. The probe was designed based on substrate predilection of CYP1A and its outstanding O-dealkylation capacity, and 1, 8-naphthalimide was chosen as fluorophore on account of its desirable photophysical properties. Absorption and emission spectra of the probe solution and reacted metabolism showed obvious red-shift with remarkable colour changes, which indicated that NEMN could be a promising ratiometric detector of CYP1A. Additionally, the selectivity assays displayed that NEMN only sensitive to CYP1A1 and CYP1A2 enzymes with scarce interference of other CYPs. Furthermore, the excellent linear relationships between the ratio of fluorescent intensities and incubation time and enzymes concentration signified time- and concentration- dependence of the probe, which were of desire benefit to quantify and monitor the CYP1A-involved biological behaviors in physiological conditions. The assay in real living samples (Human liver microsomes) further proved the analytical utility of the probe. Finally, the cytotoxicity assay and confocal fluorescence imaging demonstrated that this probe was of great promise for detecting the activity of endogenous CYP1A in human living cells.