Kinetic studies of lipase from Candida rugosa - A comparative study between free and immobilized enzyme onto porous chitosan beads

The search for an inexpensive support has motivated our group to undertake this work dealing with the use of chitosan as matrix for immobilizing lipas e. in addition to its low cost, chitosan has several advantages for use as a support.. including its lack of toxicity and chemical reactivity, allowin g easy fixation of enzymes. In this article, we describe the immobilization of Candida rugosa lipase onto porous chitosan beads for the enzymatic hydr olysis of olive oil. The binding of the lipase onto the support was perform ed by physical adsorption using hexane as the dispersion medium. A comparat ive study between free and immobilized lipase was conducted in terms of pH, temperature, and thermal stability. A slightly lower value for optimum pH (6.0) was found for the immobilized form in comparison with that attained f or the soluble lipase (7.0). The optimum reaction temperature shifted from 37 degreesC for the free lipase to 50 degreesC for the chitosan lipase. The patterns of heat stability indicated that the immobilization process tends to stabilize the enzyme. The half-life of the soluble free lipase at 55 de greesC was equal to 0.71 h (K-d = 0.48 h(-1)), whereas for the immobilized lipase it was 1.10 h (K-d = 0.63 h(-1)). Kinetics was tested at 37 degreesC following the hydrolysis of olive oil and obeys the Michaelis-Menten type of rate equation. The K-m was 0.15 mM and the V-max was 51 mu mol/ (min . m g), which were lower than for free lipase, suggesting that the apparent aff inity toward the substrate changes and that the activity of Me immobilized lipase decreases during the course of immobilization.