Multi-coloured fluorescent sensing toolbox for selective detection of nitroxyl in vitro and ex vivo

Methods for the endogenous detection of nitroxyl (azanone; HNO), the reduced and protonated derivative of nitric oxide (NO), are required to define its cardiovascular function and its key role in chronic pain. This study reports the design, synthesis and biological evaluation of 3 super-bright, highly sensitive, specific and non-cytotoxic arylphosphine-based fluorescent nitroxyl sensors that enable the detection of endogenous nitroxyl in vitro and ex vivo. The presence of endogenous nitroxyl is observed in the murine microglial cell line (BV2), primary human coronary artery endothelial cells (HCAECs) and rat cardiomyocyte cell line (H9C2) by spectroscopy and/or confocal microscopy following the addition of relevant stimulants. Furthermore, the presence of endogenous nitroxyl is detected in rat blood samples. The novel sensor 3 (rhodol 4 on a 2-(diphenylphosphanyl)benzoate backbone) is the most sensitive in detecting endogenous nitroxyl following stimulation, in the presence of biological media such as HBSS and DMEM. This then provides a validated tool for detecting nitroxyl in biological systems.