WHAT DO WE REALLY KNOW ABOUT HNO REACTIVITY? IMPLICATIONS FOR ITS FLUORESCENCE IMAGING IN VIVO

Abstract

Nitroxyl (HNO, IUPAC name azanone), being formally the protonated product of one-electron reduction of nitric oxide (NO), is an elusive reactive nitrogen species possessing interesting biological chemistry and high pharmacological importance. 1,2 Thiols, thiol proteins and metaloproteins are the most established biological targets for HNO. 1-3 Contrary to nitric oxide, HNO is a strong electrophile highly reactive towards thiols, phosphines and nitroso compounds. A number of fluorescent probes for HNO detection based on the reaction of HNO with arylphosphines have been developed recently. Here, we present the study on the HNO reactivity towards selected HNO scavengers: thiols, arylphosphines (including profluorescent probe PCM) and nitroso compounds. In this study we applied previously described competition kinetics method 4 based on two parallel, competing HNO reactions – with studied scavenger and with molecular oxygen. The latter, relatively fast reaction (k=(1.8 ± 0.3)10 4 M -1 s -1), results in the formation of peroxynitrite, 4 that can be easily detected fluorometrically with the use of fluorogenic boronate probes. Potential implications for fluorescent imaging of HNO in cells using phosphine-based fluorogenic probes are discussed.