A fluorescent probe for the dicriminatory detecion of Cys/Hcy, GSH and H2S in living cells and zebrafish

Biothiols such as Cys, Hcy, GSH, and H2S are a critical branch of reactive sulfur species (RSS) and play indispensable roles in human physiological processes. To better understand the intricate and symbiotic networks between these biothiols in physiological processes, single-molecule fluorescent probes to simultaneously distinguish them are highly desirable. Herein, we judiciously conjugated an azide-functionalized tetrahydroquinoxaline coumarin precursor with a NBD-masked 7-hydroxycoumarin through an ester bond to develop a robust fluorescent probe, TCAN, for the differentiation of Cys/Hcy, GSH, and H2S. TCAN displayed three distinct fluorescence signals in response to biothiols: blue-green for Cys/Hcy, blue for GSH, and blue-red for H2S. Significantly, the first discriminatory detection of Cys/Hcy, GSH and H2S in living organisms (zebrafish) was realized using probe TCAN.