Although it is clear that lipids are responsible for insulin resistance, it is poorly understood what types of lipids are involved. In this study, we verified the characteristic lipid species in skeletal muscle of a chronic exercise training model and a high-fat induced-obesity model. Three different lipidomics analyses revealed phospholipid qualitative changes. As a result, linoleic acid-containing phosphatidylcholine and sphingomyelin and docosahexanoic acid-containing phosphatidylcholine were characterized as chronic exercise training-induced lipids. On the contrary, arachidonic acid-containing phosphatidylcholines, phosphatidylethanolamines, and phosphatidylinositol were characterized as high-fat diet-induced lipids. In addition, minor sphingomyelin, which has long-chain fatty acids, was identified as a high-fat diet-specific lipid. This is the first report to reveal compositional changes in phospholipid molecular species in chronic exercise and high-fat-diet-induced insulin-resistant models. Due to their influence on cell permeability and receptor stability at the cell membrane, these molecules may contribute to the mechanisms underlying insulin sensitivity and several metabolic disorders.