High-Affinity Ratiometric Fluorescence Probe Based on 6-Amino-2,2'-Bipyridine Scaffold for Endogenous Zn 2+ and Its Application to Living Cells.

Zinc is an essential trace element involved in many biological activities; however, its functions are not fully understood. To elucidate the role of endogenous labile Zn 2+ , we developed a novel ratiometric fluorescence probe, 5-(4-methoxyphenyl)-4-(methylsulfanyl)-[2,2'-bipyridin]-6-amine ( 6 ( rBpyZ )) based on the 6-amino-2,2'-bipyridine scaffold, which acts as both the chelating agent for Zn 2+ and the fluorescent moiety. The methoxy group acted as an electron donor, enabling the intramolecular charge transfer state of 6 ( rBpyZ ), and a ratiometric fluorescence response consisting of a decrease at the emission wavelength of 438 nm and a corresponding increase at the emission wavelength of 465 nm was observed. The ratiometric probe 6 ( rBpyZ ) exhibited a nanomolar-level dissociation constant ( K d = 0.77 nM), a large Stokes shift (139 nm), and an excellent detection limit (0.10 nM) under physiological conditions. Moreover, fluorescence imaging using A549 human lung adenocarcinoma cells revealed that 6 ( rBpyZ ) had good cell membrane permeability and could clearly visualize endogenous labile Zn 2+ . These results suggest that the ratiometric fluorescence probe 6 ( rBpyZ ) has considerable potential as a valuable tool for understanding the role of Zn 2+ in living systems.