A near-infrared julolidine probe for visualization of mitochondrial peroxynitrite in living cells

Abstract

The overproduction of peroxynitrite (ONOO⁻) in mitochondria has been associated with various pathophysiological conditions and disorders. However, the use of fluorescent probes to visualize mitochondrial ONOO⁻ in biological systems is limited due to their low emission wavelengths and small Stokes shifts, which present significant challenges. In this study, we designed and synthesized julolidine-based near-infrared (NIR) fluorescent probes, named JQMe and JCN, specifically to monitor mitochondrial ONOO⁻. Comparative photophysical studies revealed that JQMe exhibits superior properties for sensing ONOO⁻ compared to JCN. Initially, JQMe emitted fluorescence emission at 706 nm via an intramolecular charge transfer (ICT) mechanism. Upon the addition of ONOO⁻, the NIR fluorescence emission of JQMe at 706 nm was suppressed, resulting in a rapid on–off fluorescence response within 5 minutes. JQMe exhibited high specific selectivity towards ONOO⁻ over other competing interferents, accompanied by a colorimetric change from deep blue to colorless. Additionally, JQMe exhibited a significant Stokes shift of 106 nm and a low detection limit of 6.5 nM. The proposed sensing mechanism was validated through ESI mass spectrometry and DFT studies. Furthermore, JQMe was successfully employed to monitor both endogenous and exogenous ONOO⁻ in living cells using inducer and inhibitor tests. Remarkably, time-dependent colocalization experiments revealed that JQMe effectively targets and reacts with mitochondrial ONOO⁻.