Activity modulation and reusability of β-d-galactosidase confined in sol–gel derived porous silicate glass

In the present work we applied the sol–gel method to obtain glass lentils entrapping β-d-galactosidase (β-Gal) (Eβ-Gal) within a silicate matrix. The effect of pH, temperature, polarity and salt concentration on the activity of Eβ-Gal was studied. Apparent kinetic parameters for ortho-nitro-phenyl-β-d-galactopyranoside hydrolysis catalyzed by Eβ-Gal ( V ′ max ,   K ′ M ) were lower compared to the soluble enzyme (Sβ-Gal), reflecting the solute diffusion restriction imposed by the matrix observed in the time curves, a partial protein inactivation upon encapsulation, and an improvement in the affinity of Eβ-Gal for the substrate as compared with Sβ-Gal. At pH < 4, Eβ-Gal stability was higher than that of Sβ-Gal. Eβ-Gal could be reused after storage at 4 °C for up to 90 days, and retained its activity profile within the range of pH = 2–10 and saline concentration 0–400 mM. Pre-incubation at 75 °C for 30 min fully inactivated Sβ-Gal while Eβ-Gal retained approximately 90% of its activity, even in the reused samples. Encapsulation did not introduce additional impairments to the reaction rate measured in heterogeneous dispersions, beyond those derived from their own particle-crowded environment. This reusable Eβ-Gal was resistant to typical technological conditions applied in milk processing that would lead to the unfolding and inactivation of Sβ-Gal. The results are discussed from the biophysical viewpoint.