Small molecule probes for peroxynitrite detection.

Peroxynitrite (ONOO/ONOOH) is a short-lived but highly reactive species that is formed in the diffusion-controlled reaction between nitric oxide and the superoxide radical anion. It can oxidize certain biomolecules and has been considered as a key cellular oxidant formed under various pathophysiological conditions. It is crucial to selectively detect and quantify ONOO to determine its role in biological processes. In this review, we discuss various approaches used to detect ONOO in cell-free and cellular systems with the major emphasis on small-molecule chemical probes. We review the chemical principles and mechanisms responsible for the formation of the detectable products, and plausible limitations of the probes. We recommend the use of boronate-based chemical probes for ONOO, as they react directly and rapidly with ONOO, they produce minor but ONOO‒specific products, and the reaction kinetics and mechanism have been rigorously characterized. Specific experimental approaches and protocols for the detection of ONOO in cell-free, cellular, and systems using boronate-based molecular probes are provided (as shown in Boxes 1-6).