A PRACTICAL METHOD FOR PREPARING PEROXYNITRITE SOLUTIONS OF LOW IONIC-STRENGTH AND FREE OF HYDROGEN-PEROXIDE

The reaction of ozone (similar to 5% in oxygen) with sodium azide (0.0 2-0.2 M in water) at pH 12 and 0-4 degrees C is shown to yield concent rated, stable peroxynitrite solutions of up to 80 mM. The product of t his reaction is identified based on a broad absorption spectrum with a maximum around 302 nm and by its first-order rate of decomposition (k = 0.40 +/- 0.01 s(-1) at pH 7.05 and 25 degrees C). These peroxynitri te solutions can be obtained essentially free of hydrogen peroxide (de tection limit 1 mu M) and only traces of azide (detection limit 0.1 mM ). They are low in ionic strength and have a pH of about 12 but withou t buffering capacity; therefore, they can be adjusted to any pH by add ition of buffer. These preparations of peroxynitrite frozen at -20 deg rees C show negligible decomposition for about 3 weeks of storage and follow a first-order decomposition with a halflife of about 7 days at refrigerator temperatures (similar to 5 degrees C). These preparations give reactions that are characteristic of peroxynitrite. For example, at pH 7.0, they react with L-tyrosine to give a 7.3 mol % yield of ni trotyrosine(s), and with dimethyl sulfoxide to give a 8.2 mol % yield of formaldehyde, based on starting peroxynitrite concentration.