Influence of Lipase Immobilization Mode on Ethyl Acetate Hydrolysis in a Continuous Solid-Gas Biocatalytic Membrane...

Solid-gas biocatalysis was performed in a specially designed continuous biocatalytic membrane reactor (BMR). In this work, lipase from Candida rugosa (LCR) and ethyl acetate in vapor phase were selected as model enzyme and substrate, respectively, to produce acetic acid and ethanol. LCR was immobilized on functionalized PVDF membranes by using two different kinds of chemical bond: electrostatic and covalent. Electrostatic immobilization of LCR was carried out using a membrane functionalized with amino groups, while covalent immobilization was carried out using membrane, with or without surface-immobilized polyacrylamide (PAAm) microgels, functionalized with aldehyde groups. These biocatalytic membranes were tested in a solid-gas BMR and compared in terms of enzyme specific activity, catalytic activity, and volumetric reaction rate. Results indicated that lipase covalently immobilized is more effective only when the immobilization is mediated by microgels, showing catalytic activity doubled with respect to the other system with covalently bound enzyme (4.4 vs 2.2 μmol h^-1). Enzyme immobilized by ionic bond, despite a lower catalytic activity (3.5 vs 4.4 μmol h^-1), showed the same specific activity (1.5 mmol·h^-1·g^-1_ENZ) of the system using microgels, due to a higher enzyme degree of freedom coupled with an analogously improved enzyme hydration. Using the optimized operating conditions regarding immobilized enzyme amount, ethyl acetate, and molar water flow rate, all three BMRs showed continuous catalytic activity for about 5 months. On the contrary, the free enzyme (in water/ethyl acetate emulsion) at 50 °C was completely inactive and at 30 °C (temperature optimum) has a specific activity 2 orders of magnitude lower (8.4 × 10^-2 mmol h^-1 g^-1) than the solid-gas biocatalytic membrane reactor. To the best of our knowledge, this is the first example of solid-gas biocatalysis, working in the gaseous phase in which a biocatalytic membrane reactor, with the enzyme/substrate system lipase/ethyl acetate, was used.