Carya illinoinensis (Wangenh.) K. Koch, nuts are a renowned health food. However, there are many cultivars of this nut tree, and their mature kernel lipid composition has not been thoroughly studied. Therefore, we used liquid chromatography–mass spectrometry (LC–MS) to analyze the lipid composition of mature nuts of five C. illinoinensis cultivars. In the mature kernels of all cultivars, there were 58 lipid types which were mainly composed of glycerolipids (c. 65%) and phospholipids (>30%). Triacylglycerol (TG) accounted for the largest proportion of mature nuts of all cultivars, exceeding 50%; and diacylglycerol (DG), ceramide (Cer), phosphatidylcholine (PC), and phosphatidylethanolamine (PE) were also relatively high. Additionally, nuts contain fatty acids, mainly oleic, linoleic, and linolenic acids. Our research provides a new perspective for the processing and utilization of plant and edible oils, and for the use of C. illinoinensis kernels in the development of medicine and food science.