Enzyme immobilization technology as a tool to innovate in the production of biofuels: A special review of the Cross-Linked Enzyme Aggregates (CLEAs) strategy

This review emphasizes the crucial role of enzyme immobilization technology in advancing the production of two main biofuels, ethanol and biodiesel, with a specific focus on the Cross-linked Enzyme Aggregates (CLEAs) strategy. This method of immobilization has gained attention due to its simplicity and affordability, as it does not initially require a solid support. CLEAs synthesis protocol includes two steps: enzyme precipitation and cross -linking of aggregates using bifunctional agents. We conducted a thorough search for papers detailing the syn-thesis of CLEAs utilizing amylases, cellulases, and hemicellulases. These key enzymes are involved in breaking down starch or lignocellulosic materials to produce ethanol, both in first and second-generation processes. CLEAs of lipases were included as these enzymes play a crucial role in the enzymatic process of biodiesel production. However, when dealing with large or diverse substrates such as lignocellulosic materials for ethanol production and oils/fats for biodiesel production, the use of individual enzymes may not be the most efficient method. Instead, a system that utilizes a blend of enzymes may prove to be more effective. To innovate in the production of biofuels (ethanol and biodiesel), enzyme co-immobilization using different enzyme species to produce Combi-CLEAs is a promising trend.