Abstract Lipidomics, the comprehensive illumination of lipid-based information in biology systems, involves in identifying lipids and profiling lipids and lipid-derived mediators. The development of lipidomics enables the characterization of lipid species and detailed lipid profiling in body fluid, tissue or cell, and allows for a wider understanding of the biological roles of lipid networks. Lipidomic research has been greatly facilitated by recent advances in ultra-performance liquid chromatography–mass spectrometry (UPLC–MS) and involved in lipid extraction, lipid identification and data analysis supporting applications from qualitative and quantitative assessment of multiple lipid species. UPLC technique, different mass spectrometry technique, lipid extraction and data analysis in lipidomics are reviewed. Afterwards, examples are provided on the use of UPLC–MS for finding lipid biomarkers in disease, drug, food, nutrition and plant fields. We also discuss the UPLC–MS-based lipidomics for the future perspectives and their potential problems.