High-Level Production of Recombinant Lipase by Fed-Batch Fermentation in Escherichia coli and Its Application in Biodiesel Synthesis from Waste Cooking Oils.

The enzymatic production of biodiesel from waste cooking oils (WCOs) offers a green and sustainable solution for the liquid fuel manufacture as well as waste resource recovery. In present study, liquid lipase was used to simplify the catalysis process, thereby reducing biodiesel production costs. An engineered Escherichia coli expressing Geobacillus thermocatenulatus lipase 2 (GTL2) was screened at an enzyme activity of 6.96 U/mg, after evaluating the propagating stability of the recombinant plasmids exceeding 86.11%. Through the beneficial feeding strategy and effective pH control, high-level production of GTL2 by fed-batch fermentation was achieved with an enzyme activity of 434.32 U/mg, which was almost 62 times that of shake flask fermentation. In addition, liquid GTL2 was used to prepare fatty acid methyl esters (FAMEs) using WCOs. The effects of the reaction time, catalyst loading, temperature, and methanol-to-oil molar ratio on FAMEs production using WCOs were explored, and a maximum FAMEs yield of 96.62% was achieved under optimized conditions. These results indicate that liquid GTL2 is a promising biocatalyst for efficient utilization of WCOs in the synthesis of biodiesel and provide a novel enzymatic process for biodiesel reducing the cost of production.