The kinetic study of the hydrolysis of lactose and o-nitrophenol-beta -D-galactoside (ONPG) with a beta -galactosidase from Escherichia coli, both in solution and covalently immobilized on a silica-alumina, is presented. The enzyme employed in this work had been modified previously by genetic engineering and purified to homogeneity by affinity chromatography. Firstly, the influence of pH and temperature on the activity and the stability of the enzyme, both free and immobilized, have been studied. Secondly, hydrolysis runs of lactose and ONPG with both forms of the enzyme were carried out in a wide experimental range of temperature and concentrations of substrates, products and enzyme. Data obtained were fitted to several kinetic models based on the Michaelis-Menten mechanism by non-linear regression. Finally, the models and their parameters were compared to determine the influence of the immobilization process and the substrate on the activity of the enzyme. In the hydrolysis of lactose and with both forms of the enzyme, acompetitive inhibition due to glucose was observed while the most common inhibition by galactose (which is usually a competitive inhibitor of beta -galactosidases) was not observed. Curiously, when the immobilized enzyme was the catalyst employed, lactose was an acompetitive inhibitor of the hydrolysis. When the substrate hydrolysed was the o-nitrophenol-beta -D-galactoside (ONPG), the galactose acted as a competitive inhibitor and the o-nitrophenol (ONP) was an acompetitive inhibitor for the free enzyme, being the immobilization process able to avoid the interaction between the ONP and the enzyme.