A comparison of the kinetic properties of free and immobilized Aspergillus oryzae β-galactosidase

Abstract The objective of this work was to compare the properties of free and immobilized β-galactosidase ( Aspergillus oryzae ), entrapped in alginate–gelatin beads and cross-linked with glutaraldehyde. The free and immobilized forms of the enzyme showed no decrease in enzyme activity when incubated in buffer solutions in pH ranges of 4.5–7.0. The kinetics of lactose hydrolysis by the free and immobilized enzymes were studied at maximum substrate concentrations of 90 g/L and 140 g/L, respectively, a temperature of 35 °C and a pH of 4.5. The Michaelis–Menten model with competitive inhibition by galactose fit the experimental results for both forms. The K m and V m values of the free enzyme were 52.13 ± 2.8 mM and 2.56 ± 0.3 g glucose /L min mg enzyme , respectively, and were 60.30 ± 3.3 mM and 1032.07 ± 51.6 g lactose /min m 3 catalyst , respectively, for the immobilized form. The maximum enzymatic activity of the soluble form of β-galactosidase was obtained at pH 4.5 and 55 °C. Alternatively, the immobilized form was most active at pH 5.0 at 60 °C. The free and immobilized enzymes presented activation energies of 6.90 ± 0.5 kcal/mol and 7.7 ± 0.7 kcal/mol, respectively, which suggested that the immobilized enzyme possessed a lower resistance to substrate transfer.