- Agrocybe aegerita peroxidase (AaP) is a versatile heme-thiolate protein that can act as a peroxygenase and catalyzes, among other reactions, the hydroxylation of aromatic rings. This paper reports a rapid and selective spectrophotometric method for directly detecting aromatic hydroxylation by AaP. The weakly activated aromatic compound naphthalene served as the substrate that was regioselectively converted into 1-naphthol in the presence of the co-substrate hydrogen peroxide. Formation of 1-naphthol was followed at 303 nm (ɛ 303 = 2,010 M−1 cm−1), and the apparent Michaelis–Menten (K m) and catalytic (k cat) constants for the reaction were estimated to be 320 μM and 166 s−1, respectively. This method will be useful in screening of fungi and other microorganisms for extracellular peroxygenase activities and in comparing and assessing different catalytic activities of haloperoxidase–peroxygenases.