Sustainable Immobilized Biocatalysts for Industrial Processes

The thesis is part of the EU-ITN INTERfaces project and the current project addressed the challenge of transforming immobilized enzymes into efficient and sustainable industrial biocatalysts. To achieve these objectives, a new bio-based crosslinking agent (2,5-Furandicarboxaldehyde, abbreviated as DFF) was developed with the aim to replace the fossil-based, volatile and toxic glutaraldehyde (GA). The efficiency of DFF was demonstrated in the hydrolysis of maltose catalyzed by glucoamylase from Aspergillus niger immobilized on an amino-functionalized poly(methyl methacrylate) carrier activated with DFF, which displayed high stability both in batch and in continuous flow settings, comparable to results observed for the GA-immobilized enzyme. The reactivity of DFF was also investigated, demonstrating that in water, unlike GA, it reacts with amines forming only stable imine bonds. The work on enzyme immobilization with glutaraldehyde led to a publication. In the second part of the project, rice husk was investigated as a renewable and inexpensive enzyme carrier. The lignocellulosic material was characterized using stereoscopical and SEM microscopy, porosimetry, contact angle measures. Its biodegradability in marine environment was also assessed. Rice husk was subject to delignification, in order to modify its hydrophilicity, and different pre-treatments were applied for introducing functional groups able to form covalent bonds with the enzymes. Glucoamylase and two industrially relevant lipases (TLL and CaLB) were immobilized using different immobilization techniques: (a) adsorption and crosslinking (with both DFF and GA); (b) covalent immobilization on rice husk functionalized by chemo- or enzymatic oxidation. The immobilization of lipases via adsorption and crosslinking displayed good hydrolytic activity. On the other hand, the immobilization of glucoamylase was unsuccessful with both tested techniques. This was attributed to the surface interactions between the highly glycosylated and hydrophilic enzyme and the rice husk, which presents the hydrophobic lignin on the surface. Moreover, the efficiency of samples of TLL physically immobilized on rice husk with the aid of binders was tested in the solvent-free interesterification of triglycerides, reaction used for the industrial production of cocoa-butter analogues. The tested formulations showed good stability and were able to successfully catalyze the interesterification reaction.