Entrapment of lipase into K-carrageenan beads and its use in hydrolysis of olive oil in biphasic system

Abstract

The porcine pancrease lipase was immobilized by entrapment in the beads of K-carrageenan and cured by treatment with polyethyleneimine (PEI) in the phosphate buffer. The retention of hydrolytic activity of lipase and compressive strength of the beads were examined. The activity of free and immobilized lipase was assessed by using olive oil as the substrate. The immobilized enzyme exhibited a little shift towards acidic pH for its optimal activity and retained 50% of its activity after 5 cycles. When the enzyme concentration was kept constant and substrate concentration was varied the K m and V max were observed to be 0.18 × 10 −2 and 0.10, and 0.10 × 10 −2 and 0.09 respectively, for free and for entrapped enzymes. When the substrate concentration was kept constant and enzyme concentration was varied, the values of K m and V max were observed to be 0.19 × 10 −7 and 0.41, and 0.18 × 10 −7 and 0.41 for free and entrapped enzymes. Though this indicates that there is no conformational change during immobilization, it also shows that the reaction velocity depends on the concentration. Immobilized enzyme showed improved thermal and storage stability. Hydrolysis of olive oil in organic–aqueous two-phase system using fixed bed reactor was carried out and conditions were optimized. The enzyme in reactor retained 30% of its initial activity after 480 min (12 cycles).