OXYMETRIC AND SPECTROPHOTOMETRIC STUDY OF THE ASCORBATE OXIDASE ACTIVITY SHOWN BY FROG EPIDERMIS TYROSINASE

Many studies concerning the effect of ascorbic acid on the action of t yrosinase on several substrates have been carried out with contradicto ry results. The results shown in this work comprise a hypothetical rea ction mechanism, which explains the ascorbate oxidase activity of frog epidermis tyrosinase. The reaction between frog epidermis tyrosinase and L-ascorbic acid was studied by oxymetric and spectrophotometric as says. The activity was linearly related to enzyme concentration, with a Michaelis constant for L-ascorbic acid of 0.160 +/- 0.009 mM and V-m ax of 90 +/- 4 nM/s. Maximum activity was obtained at pH 7.5. The stoi chiometry of the reaction was calculated by measuring the substrate (O -2 and L-ascorbic acid) consumption as web as the initial rates of the consumption of oxygen and the disappearance of L-ascorbic acid. The s toichiometry was found to be 1:2 (O-2:L-ascorbic acid). The action of the tyrosinase inhibitor tropolone was also studied. All the results p resent evidence concerning the ascorbate oxidase activity of frog epid ermis tyrosinase and a possible reaction mechanism based on the differ ent enzymatic forms of tyrosinase to explain such activity. Copyright (C) 1996 Elsevier Science Ltd