Walnut oil is an excellent source of essential fatty acids. Systematic evaluation of walnut lipids has significance for the development of the nutritional and functional value of walnut. Ultra-performance liquid chromatography/Orbitrap high-resolution mass spectrometry (UHPLC-Orbitrap HRMS) was used to characterize the lipids of walnut. A total of 525 lipids were detected and triacylglycerols (TG) (18:2/18:2/18:3) and diacylglycerols (DG) (18:2/18:2) were the main glycerolipids present. Essential fatty acids, such as linoleic acid and linolenic acid, were the main DG and TG fatty acid chains. Many types of phospholipids were observed with phosphatidic acid being present in the highest concentration (5.58%). Using a combination of metabolome and transcriptome analysis, the present study mapped the main lipid metabolism pathway in walnut. These results may provide a theoretical basis for further study and specific gene targets to enable the development of walnut with increased oil content and modified fatty acid composition.