BACKGROUND: Many researchers focus on exploring approaches to improve lipid productivity by photosynthetic organisms. Metabolomic analysis provides a new way to globally explore metabolic changes. To elevate lipid yields in the process of biodiesel production using photosynthetic organisms as feedstock, metabolomics combined with orthogonal partial least-squares discriminant analysis (OPLS-DA) is being applied widely to seek potential markers in regulation of lipid production. Herein, metabolic differences among three photosynthetic organisms Scenedesmus obliquus, Synechocystis sp. PCC6803 and Anabaena sp. PCC7120 were analyzed by gas chromatography coupled with time-of-flight mass spectrometry (GC-TOF-MS). RESULTS: In total, 74 metabolites were identified by GC-TOF-MS. The OPLS-DA model revealed good correlation between metabolites and lipid content (R2 = 0.9916). Nine compounds including ethanolamine were selected as potential markers affecting lipid accumulation. Herein, the highest level of C18:3 revealed its roles in regulation of the unsaturated degree of fatty acids. Further study revealed that exogenous ethanolamine (2 mmol L-1) significantly increased the lipid content (22%) in Scenedesmus obliquus. In particular, ethanolamine significantly elevated the levels of C16:2, C18:1, and C18:2. CONCLUSION: Metabolomic analysis is a useful tool to search for potential markers to improve lipid accumulation of photosynthetic organisms. Ethanolamine can be regarded as an alternative compound to improve lipid content and fatty acid levels for biodiesel production. Copyright (c) 2012 Society of Chemical Industry