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Biodiesel offers an important alternative to fossil fuel. In this work, Eversa Transform 2.0 lipase was immobilized onto 3-aminopropyltriethoxysilane (APTES) modified silica clay (SC) by glutaraldehyde. The characteristics of the functionalized supports and the immobilized lipase were investigated by FTIR, TEM, BET, and XRD. The results show that the optimal conditions of lipase immobilization are as follows: 2% glutaraldehyde concentration, 15 mg/mL lipase concentration and incubating at 25 °C for 60 min. The immobilized lipase showed a high tolerance to temperature and pH variation in comparison to the free lipase. The immobilized lipase on SC was applied as a biocatalyst for the synthesis of biodiesel from methanol and canola oil. A biodiesel yield of 86% was obtained at a temperature of 45 °C via a three-step methanol addition. A conversion yield of 67% was maintained after reusing the immobilized lipase for five cycles. This work provides a strategy for the preparation of an efficient biocatalyst for the synthesis of biodiesel. [ABSTRACT FROM AUTHOR]