Comparative analysis of standardised and common modifications of methods for lipid extraction for the determination of fatty acids

Abstract

► Lower solvent volumes can be used to quantify fatty acids with AOAC method. ► Mono- and poly-unsaturated fatty acids were underestimated with ISO method. ► Lipid peroxidation was lowest with ISO method, which uses high temperatures. Various standard techniques are commonly used for lipid extraction, but a comparative analysis amongst the different methods has not been assessed. To compare three commonly used procedures for the extraction of fatty acids from flaxseed, with adjustments for lower solvent volumes. Lipids were extracted from ground flaxseed using the International Organization for Standardization (ISO) method, the Association of Official Analytical Chemists (AOAC) method, the Folch method, and a modified AOAC procedure using lower solvent volume. Fatty acid concentrations and percent composition were determined by gas chromatography (GC) and the identities of the fatty acids were confirmed by gas chromatography–mass spectrometry (GC–MS). The extent of lipid peroxidation was also assessed using the thiobarbituric acid reactive species (TBARS) assay. GC analysis revealed that total fatty acid recoveries were not statistically different across the various methods ( P > 0.05). However, the ISO extraction procedure resulted in higher concentrations of saturated fatty acids and lower concentrations of unsaturated fatty acids, as compared with the Folch and conventional AOAC methods. Fatty acid concentrations were similar with the conventional high-volume and the modified low-solvent AOAC procedures. GCMS analysis confirmed the identity of the measured fatty acids. All assays produced TBARS, which were highest in the conventional AOAC method, followed by the Folch method ( P < 0.05) and the ISO method, respectively ( P < 0.05). Total fatty acid concentrations were equal after the different extraction methods. It appears that mono- and poly-unsaturated fatty acids in flaxseed are underestimated after extraction with the ISO method and that the solvent volumes used in the AOAC method can be decreased without compromising fatty acid concentrations. The lower yield in unsaturated fatty acids with the ISO method is not likely due to an increase in fatty acid peroxidation.