Magnetic nano-crosslinked lipase aggregates: Preparation and catalytic synthesis of OPO

1,3-Dioleoyl-2-palmitoylglycerol (OPO) is crucial for infant nutrition; however, conventional immobilized lipase requires high-purity enzymes, which increases costs and limits industrial scalability. Herein, Rhizomucor miehei lipase (RML) was immobilized on surface-modified magnetic nanoparticles using cross-linked enzyme aggregates (CLEAs) technology to produce Fe₃O₄@SiO₂@TPOAC@RML CLEAs. This approach combines the separation and immobilization of enzymes, allowing for the use of lower-purity lipase, which enhances its suitability for industrial-scale processes. The optimized Fe₃O₄@SiO₂@TPOAC@RML CLEAs exhibited excellent thermal, pH, and storage stability. The performance of Fe₃O₄@SiO₂@TPOAC@RML CLEAs was evaluated by catalyzing the synthesis of OPO through the enzymatic hydrolysis of glycerol tripalmitate (PPP) and oleic acid (OA). Under optimal conditions, the OPO content in products reached 63.33 ± 0.30%, highlighting the high efficiency of the enzymatic process. After eight reuse cycles, the Fe₃O₄@SiO₂@TPOAC@RML CLEAs retained around 75% of their relative activity. Therefore, this strategy of lipase immobilization using CLEA technology, coupled with magnetic nanoparticles, offers a promising approach for OPO synthesis.