Enzymatic Hydroesterification of Waste Oils: Combilipase-Assisted Ethyl Ester Synthesis via a Dual-Step Biocatalytic Process

Abstract

Hydroesterification is a synthetic route involving hydrolysis followed by esterification. This study aimed to investigate the production of ethyl esters via enzymatic hydroesterification through theoretical and experimental approaches. Complete hydrolysis of residual frying oil was achieved using a 1:1 mass solution at 40 °C for 4 h with 0.4% Eversa Transform 2.0 lipase relative to the oil mass. A combination of Eversa Transform 2.0 and lipase B from Candida antarctica (CAL B) was used in the esterification step. A Taguchi statistical design evaluated the effects of enzyme combination (1:1, 1:2, 1:3), FFA/alcohol molar ratio (1:1, 1:8, 1:15), biocatalyst percentage (5%, 10%, 15%), and reaction time (2, 4, 6 h). Optimal conditions were identified as a 1:8 molar ratio (FFA/ethanol), 10% biocatalyst, and a 1:3 enzyme combination for 6 h, theoretically yielding 80.1 ± 0.02% conversion. Experimentally, a 71.4 ± 0.1% conversion was achieved, slightly lower due to biocatalyst susceptibility to interferences affecting catalytic activity. Viscosity and density analyses confirmed the potential of the produced ethyl esters for future applications. This combined theoretical and experimental study highlights the feasibility of enzymatic hydroesterification using waste frying oil and combilipases as a sustainable approach for biodiesel production.