Monophenolase Activity of Polyphenol Oxidase from Artichoke Heads (Cynara scolymusL.)

Abstract

Artichoke polyphenol oxidase (PPO) has been isolated avoiding the use of severe methods which use acetone powder. The enzyme extract showed both monophenolase and diphenolase activities. Monophenolase activity of artichoke PPO was characterized using 4-hydroxyanisole as substrate with 3-methyl-2-benzothiazolinone hydrazone as a coupled nucleophile. This assay method showed high sensitivity and repeatability and allowed us to obtain a calibration curve which may be useful for determining microquantities of PPO. Monophenolase activity of artichoke PPO showed a lag period (τ) prior to the attainment of the steady state rate (Vss). Both kinetic parameters, τ andVss, depended on pH and enzyme, monophenol ando-diphenol concentrations. Optimum rate and longest lag occurred between pH 4.3 and 5. The effect of pH was related with two pKavalues of the free enzyme but not of enzyme-substrate complexes when 4-hydroxyanisol was assayed. This paper is an extension of recent studies which give uniformity to the kinetic reaction mechanism of PPOs from fruits and vegetables. The kinetic reaction mechanism of the monophenolase activity previously proposed for other PPOs was tested and confirmed for artichoke PPO. © 1997 Academic Press Limited.