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

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® (TLL1) 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 mg g−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.