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 lipase. In addition to its low cost, chitosan has several advantages for use as a support, including its lack of toxicity and chemical reactivity, allowing easy fixation of enzymes. In this article, we describe the immobilization of Candida rugosa lipase onto porous chitosan beads for the enzymatic hydrolysis of olive oil. The binding of the lipase onto the support was performed by physical adsorption using hexane as the dispersion medium. A comparative 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 for the soluble lipase (7.0). The optimum reaction temperature shifted from 37 degrees C for the free lipase to 50 degrees C 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 degrees C was equal to 0.71 h (Kd = 0.98 h(-1)), whereas for the immobilized lipase it was 1.10 h (Kd = 0.63 h(-1)). Kinetics was tested at 37 degrees C following the hydrolysis of olive oil and obeys the Michaelis-Menten type of rate equation. The Km was 0.15 mM and the Vmax was 51 micromol/(min x mg), which were lower than for free lipase, suggesting that the apparent affinity toward the substrate changes and that the activity of the immobilized lipase decreases during the course of immobilization.
Pereira
E B
EB
Department of Chemical Engineering, Maringi State University, PR, Brazil.
De Castro
H F
HF
De Moraes
F F
FF
Zanin
G M
GM
D002175:Candida / Q000201:enzymology*; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002686:Chitin / Q000031:analogs & derivatives; D048271:Chitosan; D004789:Enzyme Activation; D004795:Enzyme Stability; D004800:Enzymes, Immobilized; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008049:Lipase / Q000302:isolation & purification / Q000378:metabolism*; D013816:Thermodynamics
D004800:Enzymes, Immobilized; D002686:Chitin; D048271:Chitosan; D008049:Lipase