(P26-026-23) Integrated and Quantitative Multi-Omic Method for Analyzing the Macronutrients of Food Using Rapid-Throughput LC-MS

Objectives: Proteins, lipids, and carbohydrates are the primary energy sources of the human diet and play integral roles in overall health that arise from their diverse structures. However, analyzing these critical compounds has proven difficult since they often require different sample extraction procedures and analytical instrumentation. Herein, we introduce a novel, rapid-throughput, mass spectrometry-based method for characterizing food carbohydrates, lipids, and proteins in a single extraction.

Methods: Foods were lyophilized and ground into fine a powder. A four-in-one extraction of food samples was performed using a modified Folch: MeOH/H₂O/CHCl₃ (1:1:2 v/v). Polysaccharides, proteins, and lipids were subjected to hard acid hydrolysis with 4 M trifluoracetic acid to get total monosaccharide, amino acid, and fatty acid composition. Monosaccharides and fatty acids were chemically derivatized with 1-phenyl-3-methyl-5-pyrazolone (PMP) and 2-picolylamine, respectively. Glycosidic linkage analysis included one additional permethylation step before hydrolysis. All sample preparations were conducted in 96-well plates, enabling rapid-throughput, quantitative analysis with UHPLC-dMRM MS.

Results: The same UHPLC dMRM-QqQ platform was used for all five analyses. Limits of detection ranged from 2 to 60 ng/ml for derivatized monosaccharides and fatty acids and 5 to 50 ug/mL for native amino acids and free saccharides. The average coefficient of determination for each calibration curve was about 0.99. Method validations were performed to show the impact of the food matrix on the accuracy, precision, and reproducibility of the quantitation. Relative standard deviation (RSDs) for all analyses were lower than 20%, with a recovery range from 70% to 130%. Representative animal-based meats and plant-based meat substitutes were subjected to all five analyses for comprehensive macronutrient quantitation. The preliminary results showed that lean meats had a lower amount of monounsaturated fatty acids (MUFAs), polyunsaturated fatty acids (PUFAs), amino acids, and monosaccharides than seafood and plant-based products.

Conclusions: A novel UHPLC-QqQ MS platform for comprehensive macronutrient analysis of food using a single extraction protocol has been developed.

Funding Sources: The U.S. Department of Agriculture