Hemicyanine-based turn-off fluorescent probe for monitoring of nitric oxide in living cells

In situ imaging of nitric oxide (NO) is of great importance to study the critical roles of NO in various physiological and pathological processes. In most currently available fluorescent probes for NO, the o-phenylenediamine (OPD) based derivatives are the most greeted and studied, in which the OPD serves as the reactive site and fluorescence quencher to modulate a turn-on process via photoinduced electron transfer (PET). In this work, a unique turn-off probe containing OPD moiety, N-1, was designed and synthesized, which was radically different from the typical PET-based turn-on fluorescent probes. By theoretical calculation of molecular orbital, it was verified that not the widely used PET mechanism, but the intramolecular charge transfer (ICT) mechanism should be responsible for the turn-off process. Probe N-1 exhibited a remarkable performance with 7-folds fluorescence intensity decrease (at 490 nm), a low detection limit (20 nM), a fast response to NO within 12 min and a high selectivity for NO over other relevant analytes. Furthermore, probe N-1 was also applied to imaging NO in living cells. All results demonstrated that the new probe N-1 could be a practical and ideal NO indicator with good biological significance.