Abstract
The main goal of this study was to investigate the immobilization of commercial ß-galactosidase from Kluyveromyces lactis (Lactozym®) on Eupergit® C. A Plackett-Burman design was proposed. The ionic strength and pH were the variables that presented significant effect (p<0.1) on immobilization. The increase in the ionic strength from 0.1 to 1.5 M and the increase in pH from 6.6 to 7.4 represented an increase of 28.56% and a reduction of 18.19% in the immobilization yield, respectively. At 25°C, pH 6.6, ionic strength of 1.5 M, immobilization for 8 h, 1 mM of divalent magnesium ion and 0.4 mL of enzyme added, reached 85% immobilization yield. The free and immobilized enzymes were characterized. pH and temperature profiles showed maximum activity at pH 6.6 and 45°C, for both free and immobilized enzymes. There was a gain in thermal stability with enzyme immobilization and there was an increase of about four times in the half-life of the immobilized derivative at 45°C (from 0.43 h to 1.78 h). This greater thermal stability was also made clear through the calculation of thermodynamic parameters (ΔH, ΔG and ΔS). Km values, 30.33 mM and 104.00 mM for free and immobilized enzymes, respectively, represented a reduction in substrate affinity after immobilization, possibly owing to stereo-conformational factors. In a batch reactor for lactose hydrolysis from cheese whey, an increase in lactose conversion with immobilization was observed at 40°C and 45°C (90.43% and 65.36%, respectively) in relation to the free enzyme (84.17% and 39.58%, respectively).