Rationally modifying the dicyanoisophorone fluorophore for sensing cysteine in living cells and mice

Abstract

Cysteine (Cys), as one of the biothiols in organisms, is regarded as a significant participant in the physiological and pathological process. However, monitoring endogenous Cys in living organisms is still hindered due to the lack of effective fluorescent probe with excellent photophysical properties. Therefore, in this work, we attempted to develop excellent fluorescent probes for Cys detection with the aim of improving the photophysical properties of fluorophores. Firstly, through the rational structural modification of the dicyanoisophorone fluorophore by introducing different electron-withdrawing or electron-donating substituent groups, we found that three fluorophores with electron-withdrawing substituent group (-F, -Cl, -Br) exhibited remarkably decreased pKa value and complete deprotonation behavior in physiological pH, endowing the fluorophores with strong fluorescence emission in physiological condition and the wide pH adaptability. Subsequently, we developed three fluorescent probes (F-Cys, Cl-Cys, Br-Cys) for the Cys detection by introducing the acrylate group as the recognition unit into these three fluorophores, respectively. By comprehensively evaluating the sensing performance of three probes toward Cys, we screened out the best fluorescent probe Br-Cys. In solution, Br-Cys exhibited high sensitivity (detection limit: 86.9 nM), fast response (10 min), large fluorescence enhancement (214-fold), good selectivity, and wide pH adaptability. Furtherly, Br-Cys was successfully employed for the specific monitoring of Cys in cells and mice, providing a potential tool for understanding the physiological and pathological functions of Cys in living systems.