Preparation and application of epoxy-chitosan/alginate support in the immobilization of microbial lipases by covalent attachment

摘要

The aim of this work was the preparation and application of highly hydrophobic epoxy-chitosan/alginate as a support to immobilize microbial lipases from Thermomyces lanuginosus commercially available as Lipolase~® (TLLl) and Lipex~® 100L (TLL2) and Pseudomonas fluorescens (PFL). The catalytic properties of the biocatalysts were assayed in olive oil hydrolysis and butyl butyrate synthesis. The results indicated that 12 h was enough for TLL1 to be immobilized on the support. Covalent attachment of TLL1 turned biocatalysts highly active and around 6-fold more stable than free lipase. Based on the results, a time of incubation of 24 h was selected for further studies about the maximum immobilized protein amount and butyl butyrate synthesis. Maximum protein loading immobilized was found to be 25.4 mgg~(-1) support for TLL1, followed by TLL2 (20.5 mg g~(-1)) and PFL (15.5 mg g~(-1)) offering 80 mg protein g~(-1) support. The immobilization of TLL1 and TLL2 resulted in highly active biocatalysts (around 1300 IU g~(-1) gel), almost fivefold higher than PFL (272.4 IU g~(-1) gel). In butyl butyrate synthesis, PFL showed similar activity to TLL1 and TLL2 derivatives, up to 60 mmol L~(-1). The biocatalysts displayed high activity after five successive cycles, retaining around 95% of the initial activity.