Abstract: To establish a highly efficient and accurate determination of Sn-2 palmitic acid in triglycerides of infant formulas.Lipids of milk powder were extracted by the method of ISO14156. The Sn-1,3 fatty acids in the triglycerides were specially hydrolyzed by pancreatic lipase.The 2-monoglyceride was separated and prepared by solid phase extraction( SPE),and analyzed by gas chromatography-mass spectrometry( GC-MS). Results showed that the content of palmitic acid esterified at the Sn-2 position was 40.5%,40.3%,40.8%. The average spike recoveries for palmitic acid were 91.7% ~ 98.5%,and the relative standard deviation was 1.56%. The developed method was proved to be simple,precise,accurate and reliable,thereby being suitable for the determination of Sn-2 palmitic acid in infant formulas.