Fluorescent probe for simultaneous discrimination of Cys/Hcy and GSH in pure aqueous media with a fast response under a single-wavelength excitation

Cysteine (Cys), homocysteine (Hcy), and glutathione (GSH) are of great importance in many physiological and pathological processes. Due to their similar structure and reactivity of these three biothiols, it is a great challenge to simultaneously distinguish one from another. By virtue of two different fluorescence signals, nitrobenzofurazan ( NBD ) has been proved to be an effective sensing functional group as well as a green fluorophore precursor to construct fluorescent probes for simultaneous discrimination of Cys/Hcy and GSH. However, most of the reported NBD-based fluorescent probes suffered from a poor water-solubility, a slow response, double-wavelength excitation or two interfering fluorescence signals (narrow wavelength separation in λ max Em and/or big difference in Φ Fl ). In this study, we developed a fluorescent probe, MCQ-O-NBD , for simultaneous discrimination of Cys/Hcy and GSH using two matched fluorescent signals (well separated red and green fluorescence and comparable Φ Fl ), which could function in a pure aqueous media with a rapid response under a single-wavelength excitation. This probe exhibited high sensitivity and good selectivity as well as low detection limit, and had been successfully applied to visualize and differentiate Cys/Hcy and GSH in living cells.