A study of galacto-oligosaccharides (GOS) synthesis from lactose with β-galactosidase from Kluyveromyces lactis (Maxilact® L2000) was carried out. The synthesis was performed using various initial lactose concentrations ranging from 220 to 400 mg/mL and enzyme concentrations ranging from 3 to 9 U/mL, and was investigated at 40°C and pH 7, in a stirred-tank reactor. In the experimental range examined, the results showed the amount of GOS formed depended on lactose concentration but not on enzyme concentration. Galactose was a competitive inhibitor, while glucose was a non-competitive inhibitor. In a further study, a laboratory-scale reactor system, fitted with a 10-kDa NMWCO composite regenerated cellulose membrane, was used in a continuous process. The reactor was operated in cross-flow mode. The effect of operating pressures on flux and productivity was investigated by applying different transmembrane pressures to the system. The continuous process showed better production performance compared to the batch synthesis with the same lactose and enzyme concentrations at 40°C, pH 7. Comparison of product structures from batch and continuous processes, analyzed by HPAE-PAD and methylation analysis, showed similarities but differed from the structures found in a commercial GOS product (Vivinal®GOS). © 2004 Wiley Periodicals, Inc.