Abstract The oxidation of n-butanol to butyraldehyde by the bienzymatic system of alcohol oxidase (from Pichia pastoris)/catalase was studied in order to analyze the effect of organic media on the functioning of this system. Microaqueous, two-phase, and aqueous media were used, with three kinds of enzymatic preparations: whole cells, crude extract, and purified alcohol oxidase (with or without added catalase). Syntheses were carried out in flasks or in an oxygen probe reactor to investigate kinetics and yield of the reaction. The points of interest involved the role of water in enzyme activity, the effect of internal diffusion limitations in the various systems, and the possibilities offered by organic media concerning the transfer of butanol, oxygen, and butyraldehyde. The cellular system was found to be active in low-water media. Particular attention was directed to the case of H 2O 2, a hydrophilic compound, and to the crucial role of catalase, which was found to function in microaqueous media.