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This research investigated the usefulness of magnetic iron oxide nanoparticles (Fe 3 O 4 ) as a support to immobilize the lipase Eversa Transform 2.0 (ET 2.0) to obtain an active and stable biocatalyst, easily recoverable from the reaction medium for applications in the production of biodiesel. Biodiesel was an alternative fuel composed mainly of fatty acid esters with strong transesterification and esterification capabilities. The study focused on the esterification of oleic acid with ethanol to synthesize ethyl oleate. Magnetic nanoparticles were prepared by coprecipitation, then activated with glutaraldehyde and functionalized with γ-aminopropyltriethoxysilane (APTES). The optimal conditions for immobilizing ET 2.0 were pH 10, 25 mM sodium carbonate buffer, an enzymatic load of 200 U/g, and 1 h of contact time, obtaining 78% yield and enzymatic activity of 205.9 U/g. Postimmobilization evaluation showed that the immobilized enzyme performed better than its free form. Kinetic studies were conducted under these optimized conditions (2-96 h at 150 rpm and 37 °C). The biocatalyst was tested for the synthesis of ethyl oleate using oleic acid as the substrate and ethanol, achieving a conversion of 88.1%. Subsequent recirculation tests maintained approximately 80% conversion until the fourth cycle, confirming the sustainability of ester production. Molecular docking studies revealed that the binding affinity for the enzyme-docked oil composition was estimated at -5.8 kcal/mol, suggesting that the combination of the substrate and lipase was stable and suitable for esterification.

Magnetic nanoparticles were functionalized with polyethylenimine (PEI) and activated with epoxy. This support was used to immobilize Lipase (Eversa® Transform 2.0) (EVS), optimization using the Taguchi method. XRF, SEM, TEM, XRD, FTIR, TGA, and VSM performed the characterizations. The optimal conditions were immobilization yield (I.Y.) of 95.04 ± 0.79 %, time of 15 h, ionic load of 95 mM, protein load of 5 mg/g, and temperature of 25 °C. The maximum loading capacity was 25 mg/g, and its stability in 60 days of storage showed a negligible loss of only 9.53 % of its activity. The biocatalyst demonstrated better stability at varying temperatures than free EVS, maintaining 28 % of its activity at 70 °C. It was feasible to esterify free fatty acids (FFA) from babassu oil with the best reaction of 97.91 % and ten cycles having an efficiency above 50 %. The esterification of produced biolubricant was confirmed by NMR, and it displayed kinematic viscosity and density of 6.052 mm 2 /s and 0.832 g/cm 3 , respectively, at 40 °C. The in-silico study showed a binding affinity of -5.8 kcal/mol between EVS and oleic acid, suggesting a stable substrate-lipase combination suitable for esterification., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Magnetic biocatalysts combine magnetic properties with the catalytic activity of enzymes, achieving easy recovery and reuse in biotechnological processes. Lipases immobilized by magnetic nanoparticles dominate. This review covers an advanced bibliometric analysis and an overview of the area, elucidating research advances. Using WoS, 34,949 publications were analyzed and refined to 450. The prominent journals, countries, institutions, and authors that published the most were identified. The most cited articles showed research hotspots. The analysis of the themes and keywords identified five clusters and showed that the main field of research is associated with obtaining biofuels derived from different types of sustainable vegetable oils. The overview of magnetic biocatalysts showed that these materials are also employed in biosensors, photothermal therapy, environmental remediation, and medical applications. The industry shows a significant interest, with the number of patents increasing. Future studies should focus on immobilizing new lipases in unique materials with magnetic profiles, aiming to improve the efficiency for various biotechnological applications. [Display omitted] • Research progress, trends, and updates on magnetic biocatalysts were investigated. • The growing interest in research into magnetic biocatalysts is described. • The application of magnetic biocatalysts over the last 12 years was studied. • Comprehensive guidelines are given for magnetic biocatalyst research. • The most and least used raw materials for producing magnetic biocatalysts were investigated. [ABSTRACT FROM AUTHOR]