Reduction of the metallochromic indicators arsenazo III and antipyrylazo III to their free radical metabolites by cytoplasmic enzymes.

At a concentration much lower than that usually employed for measuring cytosolic ionized Ca2+ concentrations, arsenazo III underwent a one-electron reduction by rat liver cytosolic fraction or a hypoxanthine-xanthine oxidase system to produce an azo anion radical metabolite. NADH, NADPH, N1-methylnicotinamide, hypoxanthine, and xanthine, in that order, could serve as a source of reducing equivalents for the production of this free radical by the cytosolic fraction. The steady-state concentration of the azo anion radical and the arsenazo III-stimulated O2 consumption were enhanced by calcium and magnesium. Antipyrylazo III was ineffective in increasing O2 consumption by rat liver cytosolic fraction and gave a much weaker ESR signal of an azo anion radical with both the liver cytosolic fraction, in the presence of NADH, and the hypoxanthine-xanthine oxidase system.