Department of Biochemistry and Biophysics, Oregon State University, Environmental Health Science Center, The Linus Pauling Institute Corvallis, OR 97331, USA.
We report a simple method for the synthesis of peroxynitrite from nitrite and hydrogen peroxide that can generate hundreds of milliliters of 180 mM peroxynitrite within 1 h from start to finish. It can be scaled down to make small quantities of isotope-labeled peroxynitrite. The method requires only a syringe pump and tubing connectors and is feasible for any biochemical laboratory. Unreacted hydrogen peroxide is eliminated with manganese dioxide, using an improved preparation compared to commercially available manganese dioxide. A number of contaminants were detected by mass spectrometry in peroxynitrite solutions cleaned with commercially purchased manganese dioxide. Nitrite contamination of the peroxynitrite solution is less than 2% as determined using the Griess method. The residual contaminants are principally 0.28 M sodium chloride and 0.1 M sodium hydroxide, which pose few problems when peroxynitrite is diluted for use in biological experiments.