Near infrared spectroscopy as a new method was proposed for rapid and non-destructive measurement of fatty acid composition in intramuscular fat of packed pork loin. Gas chromatography was used as a reference method for the spectral analysis of fatty acid composition. The fiber optic in interactance mode was adopted to measure the spectra of packed pork loins by low density polythene. The original spectra were pretreated by smoothing and 2nd derivative, and then PLS calibration model was builtby using software of Unscrambler 9.6. A total of eighty two samples were used in the experiment. The samples were divided into calibration set and validation set after removing the outliers. The calibration set was used to set up calibration model and then the model was adopted to predict the samples of validation set. The results show that the correlation coefficient for C14 : 0, C15 : 1, C16 : 0, C16 : 1, C18 : 0, C18 : 1, C18 : 2, C18 : 3, C20 : 1, C20 : 4, SFA, MUFA, PUFA is 0.57, 0.76, 0.71, 0.77, 0.62, 0.81, 0.86, 0.91, 0.85, 0.91, 0.67, 0.81 and 0.95, respectively. It means that evaluating fatty acid using near infrared spectroscopy in interactance mode has higher precision. Near infrared spectroscopy technique is a feasible and rapid method for nondestructive detection of fatty acid composition in intramuscular fat of packed pork loin.