New colorimetic methods for the estimation of cytochrome c oxidase and of cytochrome c-cyto-chrome oxidase system

New colorimetric methods for the estimation of the activities of cytochrome c oxidase and of cytochrome c-cytochrome oxidase system in tissue homogenates are described, using neotetrazolium chloride in the presence of p-phenylenediamine with or without additional cytochrome c. Optimal time of incubation, optimal concentration of the incubation medium and amounts of tissue, and simple method for the extraction of the reduced neotetrazolium chloride were determined. The reduction of neotetrazolium chloride was proportional to the amount of enzyme. Using this method, colorimetric estimations of cytochrome c-cytochrome oxidase system activity in the kidney, heart, liver, brains, and skeletal muscle were made. The procedures of these methods are very simple, and they are considered to be feasible in the combination with histochemical demonstrations of these enzyme activities.