Lipase from Candida rugosa has been immobilized in different formulations of calcium alginate beads, prepared by ionotropic gelation, which differ from each other in CaCl2 concentration and hardening time, to investigate the effects of immobilization conditions on enzyme properties. Morphological studies on all hydrated beads, performed by SEM equipped with a Peltier plate, revealed a different internal compactness. Despite this, all types of beads had an immobilization yield of 100% measured with the Bradford method and about 94% evaluated from the residual activity of the preparation solutions; moreover, all entrapped biocatalysts catalyzed the complete hydrolysis of p-nitrophenyl acetate, even after one month of storage in distilled water at 4 °C. When the internal microstructure of the beads was more compact, the rate of hydrolysis of the most hydrophobic p-nitrophenyl dodecanoate was halved, probably due to mass transfer limiting effects. The immobilized lipase had better resistance to temperature inactivation than the free form: enzyme residual activity at 50 °C after a week were approximately 70% and 20% for the immobilized and free forms respectively. An excellent recyclability in water at 25 °C of entrapped enzyme was also found, having residual activity greater than 80% at the tenth reaction cycle. The best bead formulation was then used for the resolution of (R)-1-phenylethanol in aqueous solution starting from racemic (R,S)-1-phenylethyl acetate. The enantioselectivity found (E=10) was slightly higher but did not differ significantly from that of free lipase towards the same substrate (E=4).

Characterization of lipase from Candida rugosa entrapped in alginate beads to enhance its thermal stability and recyclability

Vetrano, Alice;Gabriele, Francesco;Spreti, Nicoletta
2022

Abstract

Lipase from Candida rugosa has been immobilized in different formulations of calcium alginate beads, prepared by ionotropic gelation, which differ from each other in CaCl2 concentration and hardening time, to investigate the effects of immobilization conditions on enzyme properties. Morphological studies on all hydrated beads, performed by SEM equipped with a Peltier plate, revealed a different internal compactness. Despite this, all types of beads had an immobilization yield of 100% measured with the Bradford method and about 94% evaluated from the residual activity of the preparation solutions; moreover, all entrapped biocatalysts catalyzed the complete hydrolysis of p-nitrophenyl acetate, even after one month of storage in distilled water at 4 °C. When the internal microstructure of the beads was more compact, the rate of hydrolysis of the most hydrophobic p-nitrophenyl dodecanoate was halved, probably due to mass transfer limiting effects. The immobilized lipase had better resistance to temperature inactivation than the free form: enzyme residual activity at 50 °C after a week were approximately 70% and 20% for the immobilized and free forms respectively. An excellent recyclability in water at 25 °C of entrapped enzyme was also found, having residual activity greater than 80% at the tenth reaction cycle. The best bead formulation was then used for the resolution of (R)-1-phenylethanol in aqueous solution starting from racemic (R,S)-1-phenylethyl acetate. The enantioselectivity found (E=10) was slightly higher but did not differ significantly from that of free lipase towards the same substrate (E=4).
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11697/186972
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