Dynamic proton coupled electron transfer quenching as a sensing modality in fluorescent probes

Fluorescent off-on probes based on a modular design where an analyte sensitive PET moiety is attached to a fluorophore are extremely successful. Here we report a new modular fluorescence probe design switched by dynamic quenching due to proton coupled electron transfer (PCET) mediated by collision with weak bases in solution. The fluorescence lifetime of this probe directly reports on the rate of deprotonation by the weak bases in the solution. We investigate the probe design, mechanism of response, and sensitivity to various abundant weak bases/metabolites including acetate, glutamate, phosphate, valine, and amines. We find that this modular PCET based probe design, in contrast to traditional PET probes, can work efficiently with a fluorescence lifetime readout providing a calibration free probe for weak bases. Upon further development we envision such dynamic PCET probes as sensitive tools for studies of cellular buffer systems and metabolite pools.