Development of an optimized sample preparation method for quantification of free fatty acids in food using liquid chromatography-mass spectrometry

Accurate and precise determination of free fatty acid (FFA) contents is essential for quality control and assurance in food production. Herein, a mass spectrometric study was performed to develop a sample preparation protocol that can minimize exogenous FFA contamination during the quantification of FFA in food. The quantities of exogenous FFAs were measured using various combinations of seven pretreatment methods for a sample tube, three extraction methods, and four types of sample tubes. Methanol washing could effectively reduce exogenous palmitic acid (PA) and stearic acid (SA) by 73 and 64%, respectively, in contrast to furnace baking resulting in a decrease in the amount of PA and SA contaminants by 50 and 37%, respectively. A lower amount of FFA contaminants was extracted from glass tubes during comparative analysis of the four different sample tubes studied. A combination of a methanol-washed glass tube and chloroform extraction solvent was determined to be the optimal method for minimizing the error in FFA quantification. The optimized sample preparation protocol used for FFA quantification can lower the amount of foreign palmitic acid and stearic acid to the sub-nanomolar level in the analysis of FFAs in skimmed milk.