This paper aims to study phospholipid (PL) profiling of muscle from Ctenopharyngodon idellus during room-temperature storage for 72 h by direct-infusion electrospray ionization tandem mass spectrometry (ESI-MS/MS). Five classes of PLs, including phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylinositol (PI), phosphatidylserine (PS), and sphingomyelin (SM), were analyzed. At least 110 molecular species of PLs were identified, including 32 species of PC, 34 species of PE, 24 species of PS, 18 species of PI, and 2 species of SM. The result showed that oxidation and hydrolysis are the two main causes for the deterioration of PLs in fish muscle during storage. Most content of PL molecular species increased and then decreased gradually. However, some special PE molecular species with former low abundance, such as PE 32:1, PE 34:2, and PE 34:1, emerged during the storage in quantity. It indicated that those PE molecular species may come from the microbe bred in the muscle. This phenomenon was found and discussed for the first time. The possible relevance between the emergence of these special PE molecular species and the freshness of the fish muscle during storage will be investigated in further studies.