A new xanthene-based platform for developing NIR fluorogenic probes for in vivo bioimaging

Fluorescence imaging has attracted much attention due to its unique advantages. In particular, near-infrared (NIR, 650–900 nm) probes are beneficial since they can greatly reduce the interference of autofluorescence and cause negligible photodamage to organisms. Rhodamine B, a kind of xanthene dyes, is widely used for constructing fluorogenic probes because of its outstanding photophysical properties. However, its short absorption and emission wavelengths (<590 nm) impede the bioapplication in living mice. Herein, by extending the conjugated structure of xanthene, we present a novel fluorophore F3 with high quantum yield (Φ = 16.1%), NIR fluorescence (maxima emission at 770 nm), as well as the tunable fluorescence off–on switch via the incorporation of analyte-specific moieties. As a proof of concept, we synthesize two NIR fluorescence probes (F3ONOO and F3NTR) for peroxynitrite (ONOO–) and nitroreductase (NTR) based on fluorophore F3. Using the two probes, we have realized fluorescence imaging of not only ONOO– fluctuations in hepatotoxicity, but also NTR changes in tumor mice, demonstrating the feasibility of the platform for developing activatable NIR fluorogenic probes for other biospecies in the future.