Part:BBa_K5193003

This is a type of lipase used to esterify alcohol and acid into ester. In order to enhance the scent of our essential oil, we aimed to increase the amount of ester by using lipase (lip4) to catalyze acid and alcohol into ester.[1] See graph 1 for the mechanism of lipase-catalyzed esterification.

Figure 1. Lipase-catalyzed synthesis of ester through direct esterification, alcoholysis or acidolysis. Source: Kuo, C.-H. et. al. 2020. [2]

We incubate our essential oil extract with enzyme extract of lip4 for 2.5 hours in a 37C 200 rpm shaker, allowing the reaction to take place. Then, we centrifuge the mixture at 4956 x g for 10 minutes before taking the upper oil layer (desired product) to a new tube. The final esterified oil product went through Gas Chromatography–Mass Spectrometry (GC-MS) to validate the change of chemical composition in essential oil with and without post-treatment. The result is shown below.

We used GC-MS (Agilent 8890-7000D) for component analysis. We selected 10 top fold change component

References: 1. Tang SJ, Sun KH, Sun GH, Chang TY, Lee GC. Recombinant expression of the Candida rugosa lip4 lipase in Escherichia coli. Protein Expr Purif. 2000 Nov;20(2):308-13. doi: 10.1006/prep.2000.1304. PMID: 11049754. 2. Kuo, C.-H.; Huang, C.-Y.; Lee, C.-L.; Kuo, W.-C.; Hsieh, S.-L.; Shieh, C.-J. Synthesis of DHA/EPA Ethyl Esters via Lipase-Catalyzed Acidolysis Using Novozym® 435: A Kinetic Study. Catalysts 2020, 10, 565. https://doi.org/10.3390/catal10050565