The reaction of hydrogen peroxide with protohemin has been examined in aqueous solution (pH 5-10) with potassium ferrocyanide as a substrate. Kinetic studies were carried out in the presence and absence of imidazole bases(B : imidazole, 1-methylimidazole, 2-methylimidazole, 4-methylimidazole and 1,2-dimethylimidazole). The apparent second-order rate constant (k(app)) for the formation of ferricyanide was used as the index of the peroxidase activity. in the absence of bases, hemin showed weak and constant activity at alkaline pH but has lost it at acidic pH. A plot of log k(app) vs pH has revealed that the association of one H+ equivalent with apparent pK(a) = 6.9 has inhibited the reaction. Along with the pH-dependency of the absorption spectra of ferric hemin, it was suggested that the hydroxide coordination to the ferric hemin is essential to the generation of the peroxidase activity. On the other hand, addition of bases resulted in an enhancement of the reaction rate at alkaline pH. This effect was well. accounted for by the mono-ligation of a base to the iron atom in the high-valent porphyrin intermediate(s). However, the reaction was found to be inhibited in the presence of large excess of unhindered bases, and the inhibition was strongly correlated to the formation of bis-coordinated ferric hemin. Therefore, bis-ligating bases have prevented the access of H2O2 to the ferric hemin and the subsequent formation of the intermediates. At acidic pH, the bis-coordination was promoted and the unhindered bases were less effective activators. Addition of sterically hindered 2-methylimidazole, by contrast, resulted in the higher activity at acidic pH than that in the base-free system. This may suggest that oxo-ligand in the intermediate was activated by the hydrogen bonding of the protonated 2-methylimidazole. A reaction scheme that satisfies the above criteria is proposed, and the correlation with the reactions of heme enzymes is discussed.