Isoamyl acetate production was investigated using immobilized Rhizomucor miehei and Candida antarctica lipases by esterification of acetic acid with isoamyl alcohol in a solvent-free system. The acid-alcohol molar ratio, enzyme type and amount, reaction time, temperature and shaking rate were chosen as efficient reaction parameters. The initial reaction rate, isoamyl acetate concentration and conversion values obtained using Novozym 435 were higher (8.3-fold) than those of the values of Lipozyme RM IM. Isoamyl acetate production was maximal at an acid-alcohol molar ratio of 1/2, 5% enzyme (g/g substrates), 30 degreesC and 150 rpm for Novozym 435. A 6 h reaction time is sufficient to reach a high ester concentration (380 g/l) and conversion (80%) under these conditions. Acid inhibition of enzyme activity occurred above a critical acid-alcohol molar ratio of 1/2 (3.6 M acid). The isoamyl acetate concentration achieved in a solvent-free system is higher (2.5-fold) than those reported by other researchers, although the same conversion was obtained. Isoamyl acetate production is possible at a high acetic acid concentration (3.6 M) using Novozym 435 in a solvent-free system. (C) 2002 Elsevier Science Ltd. All rights reserved.