Reactive oxygen species (ROS) such as superoxide and reactive nitrogen species (RNS) such as nitric oxide are considered to be involved in the pathogenesis of various diseases. Although nitric oxide and superoxide are both endogenous compounds and important for physiological reactions including vasodilatation, signal transduction, and protection from infection, they have toxic effects on various biological components when overproduced. Peroxynitrite is produced after the diffusion rate-limiting reaction between nitric oxide and superoxide, and is a strong oxidant for various biological components as well as a nitrating reagent for free tyrosine and protein tyrosine residues. Since peroxynitrite is formed from superoxide and nitric oxide under physiological conditions in vitro, it is also considered to be produced in vivo when sufficient amounts of superoxide and nitric oxide are produced to react directly with one another. Because of its high reactivity, peroxynitrite can cause oxidative damage to important biological components, such as low density lipoprotein (LDL) oxidation, lipid peroxidation, and DNA strand breakage, [1][2][3] as well as the nitration of tyrosine residues to produce nitrotyrosine.4) The presence of nitrotyrosine in tissues and cell cultures is often used as a marker for peroxynitrite production although nitrotyrosine is not the sole product of peroxynitrite.With reports that nitrotyrosine is upregulated in tissue in several kinds of diseases, peroxynitrite is assumed to be produced not only in inflammation, 2,3) but also in neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and amyotropic lateral sclerosis.1,5-10) Additionally, the tyrosine nitration in the substrate proteins of tyrosine kinases is assumed to prevent the tyrosine phosphorylation by the kinases. Tyrosine phosphorylation is very important to the signal transduction in cells.11,12) Furthermore, the oxidation and nitration activities of peroxynitrite are considered to play important roles in the functional damage to enzymes and receptors. It has been reported that the activity of the respiratory chain in mitochondria is inhibited by peroxynitrite treatment due to the oxidative destruction of iron-sulfur containing proteins, 13) the inhibition of the redox cycling of cytochrome c by the oxidation of ferrocytochrome c, 14) and the inhibition of cytochrome c oxidase.15) It has also been shown that cytochrome c is released from mitochondria and involved in the activation of caspases during the apoptotic process. [16][17][18] Although cytochrome c is an important component of the electron transfer system in mitochondria, few studies have been conducted on the effect of peroxynitrite on the redox activity of cytochrome c. Very recently, Radi and colleagues, reported nitration of cytochrome c by peroxynitrite in vitro, 19,20) and demonstrated that a low concentration of peroxynitrite caused mono-nitration of the tyrosine residue at position 67. Since cytochrome c is a water-soluble protein in the intermembran...