Lipase from Rhizomucor miehei onto home-made hydrophobic polymers: Stable and efficient biocatalysts

Abstract

Immobilized lipases are biotechnological spotlights as tools to improve and enhance enzyme applicability. Here, lipase from Rhizomucor miehei (RML) was immobilized onto core/shell supports, named poly(methyl methacrylate) (PMMA/PMMA); poly(methyl methacrylate) copolymerized with divinylbenzene (PMMA-co-DVB/PMMA-co-DVB); and polystyrene copolymerized with divinylbenzene (PS-co-DVB/PS-co-DVB), and on a commercial non-core/shell porous support (polypropylene - Accurel MP 1000). Different enzyme loadings were evaluated (400 U.g−1, 500 U.g−1, 600 U.g−1, 4000 U.g−1, and 6000 U.g−1), and the immobilized biocatalysts were used in hydrolysis (using p-nitrophenyl laurate (p-NPL) and p-nitrophenyl butyrate (p-NPB) as substrates) and in esterification reactions (using oleic acid and ethanol as substrates). The influence of pH, temperature, and buffer concentration in the hydrolysis of p-NPL was also investigated. The immobilized biocatalysts were compared to free RML and/or to commercially immobilized RML (RM-IM) and/or to Novozym 435. RML interacted differently with each support, being RML-PMMA-co-DVB/PMMA-co-DVB and RML-PMMA/PMMA the biocatalysts with the highest enzymatic activities (hydrolytic activity: >400 U.g−1, using p-NPL; esterification activity: 25,532 U.g−1). Immobilization improved the thermal and pH tolerance and buffer concentration tolerance of RML. Both free RML and the immobilized biocatalysts exhibited their maximum activity at pH 7, with the exception of RML-Accurel MP 1000, which displayed optimal performance at pH 9.0. In addition, RML-PS-co-DVB/PS-co-DVB demonstrated increased activity at 70 °C. Notably, RML-PMMA/PMMA and RML-PMMA-co-DVB/PMMA-co-DVB showcased a maximum activity of 120 U/g, and this was achieved at different temperatures, 30°C and 50°C, respectively. This diverse library of immobilized biocatalysts shows promising potential for application in various industrial processes.