Nitric oxide reacts with superoxide to form peroxynitrite, a strong oxidizing species. Peroxynitrite can either directly oxidize molecules such as thiols or protonate to peroxynitrous acid, which can yield an oxidant with a reactivity similar to that of hydroxyl radical in a transition metal-independent mechanism. This oxidative chemistry of peroxynitrite, however, is inhibited by the metal chelator desferrioxamine. Indeed, desferrioxamine, was a potent inhibitor of dimethylsulfoxide, hydrogen peroxide, 5,5-dimethyl-1-pyrroline-N-oxide, and luminol oxidation, whereas the metal chelator diethylenetriaminepentaacetic acid, and ferrioxamine, the iron complex of desferioxamine, were not. Two other hydroxamates, acetohydroxamate and salicylhydroxamate, were also effective inhibitors. Stopped-flow experiments showed that there is no direct reaction between peroxynitrite anion or cis-peroxynitrous acid with desferrioxamine. Electron paramagnetic resonance (EPR) studies showed the formation of the desferrioxamine nitroxide radical in incubations containing desferrioxamine, but not ferrioxamine, indicating that the hydroxamic group acts as a one-electron donor to peroxynitrite-derived oxidants. Taken together, our results led us to propose that desferrioxamine can inhibit the oxidative chemistry of peroxynitrite by reaction of the hydroxamic acid moieties with trans-peroxynitrous acid.