between nitric oxide ( โข NO), a versatile biological media-A new method for the preparation of high concentrator (4-7), and superoxide (O โข0 2 ), a ubiquitous reactive tions of peroxynitrite (up to 1 M) is described. The synoxygen radical (8,9). Several potential loci exist in vivo thesis uses a two-phase system and involves a displacewhere peroxynitrite can be formed, including endothement reaction by the hydroperoxide anion (in the lial cells (10,11), macrophages (12), neuronal cells (13), aqueous phase) on isoamyl nitrite (in the organic phase). and neutrophils (14,15). While both โข NO and O โข0 2 are The product peroxynitrite remains in the aqueous weak oxidizing agents, peroxynitrite and its conjugate phase, whereas isoamyl alcohol forms a new organic acid (pK a ร
6.8) peroxynitrous acid (HOONO) are pophase along with the unreacted isoamyl nitrite. The tent one-and two-electron oxidants (16-18), and the aqueous phase contains some 0.15 M isoamyl alcohol and toxic effects of โข NO are thought to result in part from the unreacted hydrogen peroxide, but no isoamyl nitrite. the reactions of this inorganic peroxyacid.
Removal of isoamyl alcohol or traces of isoamyl nitrite
Eight synthetic methods have been reported for the is accomplished by washing the aqueous phase with dipreparation of peroxynitrite. These include (i) the reacchloromethane, chloroform, or hexane. A near total retion of ozone with azide ions in the presence of a low moval of hydrogen peroxide is then achieved by passing concentration of alkali (19-21), (ii) the autooxidation the solutions through a short column of manganese dioxof hydroxylamine in a moderately alkaline solution conide. The peroxynitrite in these postprocessed solutions taining 0.1-0.5 N NaOH (22, 23), (iii) the reaction of has a broad absorption spectrum with a maximum nitrite with acidified hydrogen peroxide followed by a around 302 nm, follows a characteristic first-order debase quench (24-26), (iv) the reaction of H 2 O 2 with composition at pH 7.2 and 25ะC (k ร
0.34 { 0.1 s 01 ), and 2-ethoxyethyl nitrite in a basic medium (27), (v) the reacts with organic compounds to give either nitrated reaction of โข NO with alkaline H 2 O 2 (28), (vi) the reacor one-electron transfer products. When stored frozen at 020ะC, these peroxynitrite solutions decompose at a tion of โข NO with solid K / O โข0 2 ( 23), (vii) the reaction of rate of about 1.7% per day and should be used within 2-โข NO with tetramethylammonium superoxide in liquid 4 weeks. For short-term storage of about 1 week or less, ammonia (29), and (viii) the UV photolysis of solid K / these solutions can be stored at refrigerator tempera-NO 0 3 (30, 31). Some of these methods produce peroxynitures (ร5ะC) where peroxynitrite has a half-life of about trite solutions that contain excess alkali as well as 7 days.