Abstract: In order to study the difference of fatty acid and phospholipid composition in different families and species of Perciformes freshwater fish, we analyzed the fatty acids and phospholipids in muscles of Scortum barcoo, Perca fluviatilis, Lucioperca lucioperca, Micropterus salmoides and Oreochromis mossambicus by gas chromatography-mass spectrometry (GC-MS) and high performance liquid chromatography (HPLC-ELSD). Twenty-one kinds of fatty acids had been detected in those fish. Eight kinds of common fatty acids were detected (C_14:0, C_16:0, C_18:0, C_16:1, C_18:1, C_18:2, C_20:5 and C_22:6). Polyunsaturated fatty acids (PUFA) were rich and the content was higher than 35% except for S. barcoo (26.81%). Six kinds of phosphatidylcholine were detected in those fish, including phosphatidylethanolamine (PE), sphingomyelin (SM), phosphatidylinositol (PI), lysophosphatidylcholine (LPC), phosphatidylcholine (PS) and phosphatidylcholine (PC). The PC contents in those fish muscles phospholipids were the highest, accounting for 45.84%−62.55% of total phospholipid, followed by PE and PI contents. The PS and SM contents were relatively low, accounting for only 2.19%−5.27% and 2.06%−3.65% of total phospholipids.