A hemicyanine-based dual-modal probe for fluorescence and mass spectrometry imaging of peroxynitrite in biosamples

Peroxynitrite (ONOO^-) is a highly reactive oxidant formed by the reaction of nitric oxide with superoxide. Excessive ONOO^- can be produced by the body in response to multiple diseases, resulting in cell death through oxidation and nitration processes. However, technical challenges arise in detection due to its high reactivity and very short lifespan. In this work, we rationally designed and synthesized a novel tetrazine-hemicyanine-based probe (TZN-HCY). This probe demonstrates the ability to capture ONOO^- with high selectivity and sensitivity, enabling the detection and imaging of ONOO^- in biological samples with both fluorescence and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS). The 3,6-disubstituted tetrazine moiety could react with ONOO^-, and the hemicyanine skeleton with a permanent positive charge could enhance MALDI MS detection. The responsive performances of the TZN-HCY probe were verified on cell models and livers of hepatic ischemia-reperfusion injury (HIRI) model mice. Due to the applicability to dual-mode imaging, the formation of ONOO^- and its content change in the liver of living mice were visualized by fluorescence imaging, while the fine-scale spatial distribution of ONOO^- in liver tissues was revealed by MALDI MS imaging. This dual-modal probe could serve as a powerful tool for elucidating the diverse and complex roles of biogenic ONOO^- in ischemia-reperfusion injury.