An improved direct transesterification method for fatty acid determination of Phaeodactylum tricornutum

Development of microalgae biotechnology demands a rapid and accurate method of fatty acid determination. In this study, an improved two-step direct transesterification (TSDT) using a sequential combination of alkaline catalyst (NaOH) and acid catalyst (AcCl) was found to enhance the recovery yield of fatty acids in Phaeodactylum tricornutum compared with the methods of traditional solvent extraction (the Folch, the Bligh and Dyer, or the soxhlet methods) followed by transesterification. There was no significant difference (p > 0.05) in the transesterification efficiency between TSDT and AOAC 991.39. TSDT converted intracellular lipid to saponifiable lipids, which were then in situ transesterified to fatty acid methyl esters. All the reactions occurred in one tube, so the extraction step was eliminated and the small samples were enough to assay. Higher levels of fatty acid content with recovery yield above 96 % were obtained with TSDT compared with the traditional extraction-transesterification methods. In the presence of water, TSDT was more effective than each catalyst used alone. A water content up to 10 % of total reaction volume was allowed in the TSDT determination with no effect on results. TSDT is a simple and more accurate method suitable for wet and small samples assay in P. tricornutum cultures.