Optimal Immobilization ofβ-Galactosidase ontoκ-Carrageenan Gel Beads Using Response Surface Methodology and Its Applications

Abstract

β-Galactosidase (β-gal) was immobilized by covalent binding on novelκ-carrageenan gel beads activated by two-step method; the gel beads were soaked in polyethyleneimine followed by glutaraldehyde. 22full-factorial central composite experiment designs were employed to optimize the conditions for the maximum enzyme loading efficiency. 11.443 U of enzyme/g gel beads was achieved by soaking 40 units of enzyme with the gel beads for eight hours. Immobilization process increased the pH from 4.5 to 5.5 and operational temperature from 50 to 55°C compared to the free enzyme. The apparentKmafter immobilization was 61.6 mM compared to 22.9 mM for free enzyme. Maximum velocityVmaxwas 131.2 μmol·min−1while it was 177.1 μmol·min−1for free enzyme. The full conversion experiment showed that the immobilized enzyme form is active as that of the free enzyme as both of them reached their maximum 100% relative hydrolysis at 4 h. The reusability test proved the durability of theκ-carrageenan beads loaded withβ-galactosidase for 20 cycles with retention of 60% of the immobilized enzyme activity to be more convenient for industrial uses.