Rational Design of a Fluorogenic Probe for Sphingosine Based on Precisely Controlled Intramolecular Spirocyclization...

We synthesized a series of coumarin-rhodamine (CR) derivatives with near-infrared fluorescence (NIRF) properties, and found their intramolecular spirocyclization equilibrium strongly depends on the hydrophilicity/hydrophobicity of their 5'-substituent: the hydrophobic substituent favors a closed spirolactone form (NIRF OFF), whereas the hydrophilic one prefers an open form (NIRF ON). On the basis of these findings, we developed a new design strategy to develop fluorogenic probes based on hydrophilicity-regulated ring-opening of CR scaffold. Based on this mechanism, a fluorogenic probe (CR-SE2) for sphingosine (Sph) was developed. CR-SE2 can react with Sph to afford the ligation product CR-Sph, which leads to the replacement of its salicylaldehyde ester recognition moiety by a less hydrophobic Sph, thereby shifting the equilibrium of CR platform toward the open form and thus resulting in a large turn-on fluorescence readout in the NIR region. CR-SE2 features fast response, high sensitivity, and high selectivity toward Sph, and has been applied to monitor Sph levels in senescent cells, mice, and their tissue slices, revealing a close correlation between aging and Sph levels in biological specimens. The present study provides a more generalizable design strategy for developing fluorogenic probes to study various pathological and biological processes.