Abstract: Lipids are a class of compounds with unique physiological functions and activities. They participate in the regulation of a variety of plant responses to abiotic stress, which are essential for maintaining physiological homeostasis in plants. Since its proposal in 2003, lipidomics has rapidly developed as an emerging discipline that provides novel technical support for traditional metabolomics. Lipidomics can aid in clarifying the mechanisms of lipid metabolic regulation in plants. Moreover, the advantages of being label-free, non-specific, highly sensitive an d allowing for simultaneous elemental and molecular analysis mass spectrometry imaging (MSI) has been widely used to decipher the spatial distribution of various lipid molecules in plant tissues. Here, we introduce the current research status of lipidomics and MSI technologies, and comprehensively review recent progress in applying lipid analysis techniques to study lipid metabolic regulation in plant seeds, especially the application of MSI techniques for mapping lipid species in plants. Lipidomics and MSI are important complements to commonly used multi-omics technologies, and they will provide new opportunities for deeper exploration of plant metabolic pathways and regulatory mechanisms. The continuous development of high-spatial-resolution MSI technologies is expected to further promote the deep analysis and prospecting applications of lipid metabolism network regulation at the spatially resolved level in plants.