The cytochrome oxidase content of rat tissues was investigated, utilizing the accumulation of the free radical formed by the emzymatic, univalent oxidation of a stable, non-toxic substrate, N-phenyl- p-phenylenediamine ( p-aminodiphenylamine). This procedure eliminated the non-enzymatic coupling of the radical with α-naphthol, found in the classic Nadi reaction. Using homogenates, the following decreasing order of enzyme activity was found: heart, kidney, diaphragm, liver, rectus abdominis. A more sensitive assay was also developed, involving the reduction of cytochrome c by ascorbic acid, and using 2-( p-iodophenyl)-3-( p-nitrophenyl)-5-phenyl tetrazolium chloride to analyze unoxidized ascorbic acid. The same order of tissue activity was demonstrated. The cytochrome oxidase content of a tissue may be a reflection of its physiological role.
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Pearl et al. (1963) studied this question.