Lipase-Ca2+ hybrid nanobiocatalysts through interfacial protein-inorganic self-assembly in deep-eutectic solvents (DES)/water two-phase system for biodiesel production

The production of biodiesel catalyzed by lipase has the advantages of simple process, low energy consumption, and low wastewater treatment requirements. However, low activity, poor stability, and difficulty in recycling still limit the application of lipase in production of biodiesel. Herein, a novel lipase hybrid biocatalyst with superior catalytic activity and stability (lipase from Aspergillus oryzae CJLU-3, AOCL@CaP in DES) was developed by dual activation of Ca2+ and interfacial effect of deep-eutectic solvents (DES)/water two-phase system. AOCL@CaP in DES exhibited significantly enhanced enzymatic activity with a Kcat/Km value of 692 s-1 mM(-1) and excellent thermostability, tolerance to chemical denaturants, substrate selectivity, and reusability compared with free lipase. Recovery activity of the prepared hybrid biocatalyst in pure water system (AOCL@CaP) and AOCL@CaP in DES was 195% and 250% using p-NPL as substrate, respectively. AOCL@CaP in DES exhibited about 253% of the original activity after 100 days of storage, while free AOCL, AOCL@CaP almost completely lost activity. The FAME content in product biodiesel catalyzed from soybean oil by AOCL@CaP was only 60%, However, the FAME content catalyzed by AOCL@CaP in DES reached 87%, and could be maintained at 50% even after 10 cycles.