A long-lived Donor-Acceptor fluorescent probe for sequential detection of Cu²⁺ and biothiols

A new long-lived Donor-Acceptor (D-A) fluorophore based on carbazolyl dicyanobenzene was developed as an ON-OFF-ON multifunctional fluorescent probe 1 for sequential detection of Cu²⁺ and biothiols (Cys, Hcy and GSH). The fluorescence of probe 1 can be significantly and selectively quenched by Cu²⁺. Meanwhile, the fluorescence lifetime decreased from 2.1 μs to 18.5 ns. The limit of detection was determined to be 33.6 nM. Upon addition of biothiols (Cys, Hcy and GSH), the generated ensemble 1-Cu²⁺ displayed a "turn-on" fluorescent response at 555 nm and an obvious recovery in fluorescence lifetime and UV-vis absorption within 1 min. The limit of detection for Cys, Hcy and GSH were calculated by fluorescence titration experiments to be 0.19, 0.21 and 0.29 μM, respectively. The ensemble 1-Cu²⁺ was further successfully applied in bioimaging.