Quantitation of total fatty acids in plasma and serum by GC-NCI-MS

Abstract

Polyunsaturated fatty acids (PUFAs), including essential omega-3 and omega-6 fatty acids, play important roles in diverse physiological and pathological processes. Diligent monitoring of PUFAs is recommended for individuals with increased risk of developing essential fatty acid deficiency (EFAD), including premature and very low birth weight infants, patients on prolonged parenteral nutrition, and those with dietary restrictions, for example due to inborn errors of metabolism. Here, we present a gas chromatography-negative chemical ionization-mass spectrometry (GC-NCI‐MS) method for the quantitation of total levels of twenty-two fatty acids (C12‐C22) in serum/plasma, including omega-3 and omage-6 PUFAs. Hydrolysis was used to release esterified fatty acids, which were analyzed by GC-NCI-MS as pentafluorobenzyl esters in selected-ion monitoring (SIM) mode. The calibration curves for all analytes had consistent slopes with R2 of ⩾0.990. Intra- and inter-assay precision CVs were ⩽9.0% and ⩽13.2%, respectively. Samples were found to be stable for 24h at room temperature, at least 7days at 4°C, at least 75days at −20°C, and for three freeze/thaw cycles. No matrix effects or interferences were observed. This method offers improvements over published studies including smaller sample volume, inclusion of additional internal standards, analysis in a single injection, and use of methane reagent gas. This method could be used in a clinical laboratory setting for the diagnosis of EFAD, evaluation of nutritional status, and diet monitoring.