Unraveling the Chemosensing Mechanism by the 7-(Diethylamino)coumarin-hemicyanine Hybrid: A Ratiometric Fluorescent Probe for Hydrogen Peroxide.

Abstract

The hemicyanine hybrid containing the 7-(diethylamino)coumarin (ACou) donor attached to the cationic indolenium (Ind) acceptor through a vinyl linkage (ACou-Ind) represents a classic ratiometric fluorescent probe for detecting nucleophilic analytes, such as cyanide and reactive sulfur species (RSS), through addition reactions that disrupt dye conjugation to turn off red internal charge transfer (ICT) fluorescence and turn on blue coumarin emission. The chemosensing mechanism for RSS detection by ACou-Ind suggested in the literature has now been revised. Our studies demonstrate that thiolates react with ACou-Ind through conjugate addition to afford C4-SR adducts that lack coumarin fluorescence due to photoinduced electron transfer quenching by the electron-rich enamine intermediate. Thus, ACou-Ind serves as a turn-off probe through loss of red ICT fluorescence upon RSS addition. The literature also suggests that blue coumarin emission of thiolate adducts is enhanced in the presence of reactive oxygen species (ROS) due to ROS-mediated cellular changes. Our studies predict that such a scenario is unlikely and that thiolate adducts undergo oxidative deconjugation in the presence of H2O2, the pervasive ROS. Under basic conditions, H2O2 also reacts directly with ACou-Ind to generate intense coumarin fluorescence through an epoxidation process. The relevance of our chemosensing mechanism for ACou-Ind was assessed within live zebrafish, and implications for the utility of ACou-Ind for unraveling the interplay between RSS and ROS are discussed.