A highly selective and sensitive carbazole-based fluorescent probe for peroxynitrite detection and cellular imaging

• Probe EHMC showed high selectivity and sensitivity for peroxynitrite (ONOO − ). • The probe EHMC have been applied in intra-cellular fluorescence imaging. • The probe EHMC has good biocompatibility and low toxicity. Peroxynitrite, a colorless and odorless reactive nitrogen species, plays a critical role in various physiological processes within biological systems. Therefore, developing a reliable method for quantitative detection of peroxynitrite in living systems is essential. In this study, we synthesized a simple, two-step, reaction-based fluorescent probe, designated as EHMC , specifically for peroxynitrite detection. The synthesis involved the use of 4-(9-phenyl-9H-carbazol-3-yl)benzaldehyde as the fluorophore and 2-hydrazineylpyridine as the recognition group, both of which exhibit inherently weak fluorescence. Upon oxidation of the C=N bond by peroxynitrite, EHMC undergoes conversion into an aldehyde, resulting in a significant fluorescence response. EHMC demonstrated high selectivity and sensitivity, achieving a detection limit of 5.43 µM under physiological conditions. Additionally, EHMC was successfully applied to HeLa cellular models, where it exhibited excellent cellular imaging capability for peroxynitrite detection in living systems.