Vanillin is a main flavor ingredient of vanilla beans. It Is widely used as a food flavor, and is an important material in various industrial fields. The production of vanillin from vanillylamine was investigated using amine oxidase (AO) from Aspergillus niger and monoamine oxidase (MAO) from Escherichia coli, which have previously been isolated, characterized, and molecularly cloned in our laboratory. Oxidation of vanillylamine was observed with both AO and MAO. Kinetic analyses of the enzyme reactions revealed that AO was a more efficient producer of vanillin than MAO. With AO, 1 mM of vanillylamine was oxidized almost completely in about 30 min by 2.7 mu g/ml of the enzyme. The product of the enzyme reaction was confirmed to be vanillin by HPLC and GC-MS. Continuous production of vanillin with immobilized AO was also investigated. The results suggest that AO from A. niger will be useful for the industrial synthesis of vanillin.