In the present work we applied the sol–gel method to obtain glass lentils entrapping β- d-galactosidase (β-Gal) (E β-Gal) within a silicate matrix. The effect of pH, temperature, polarity and salt concentration on the activity of E β-Gal was studied. Apparent kinetic parameters for ortho-nitro-phenyl-β- d-galactopyranoside hydrolysis catalyzed by E β-Gal ( V ′ max , K ′ M ) were lower compared to the soluble enzyme (S β-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 E β-Gal for the substrate as compared with S β-Gal. At pH < 4, E β-Gal stability was higher than that of S β-Gal. E β-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 S β-Gal while E β-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 E β-Gal was resistant to typical technological conditions applied in milk processing that would lead to the unfolding and inactivation of S β-Gal. The results are discussed from the biophysical viewpoint.