Application of ethyl esters and d3-methyl esters as internal standards for the gas chromatographic quantification of transesterified fatty acid methyl esters in food

Ethyl esters (FAEE) and trideuterium-labeled methyl esters (d3-FAME) of fatty acids were prepared and investigated regarding their suitability as internal standards (IS) for the determination of fatty acids as methyl esters (FAME). On CP-Sil 88, ethyl esters of odd-numbered fatty acids eluted approximately 0.5 min after the respective FAME, and only coelutions with minor FAME were observed. Depending on the problem, one or even many FAEE can be added as IS for the quantification of FAME by both GC-FID and GC-MS. By contrast, d3-FAME coeluted with FAME on the polar GC column, and the use of the former as IS requires application of GC-MS. In the SIM mode, m/z 77 and 90 are suggested for d3-methyl esters of saturated fatty acids, whereas m/z 88 and 101 are recommended for ethyl esters of saturated fatty acids. These m/z values give either no or very low response for FAME and can thus be used for the analysis of FAME in food by GC-MS in the SIM mode. Fatty acids in sunflower oil and mozzarella cheese were quantified using five saturated FAEE as IS. Gravimetric studies showed that the transesterification procedure could be carried out without of loss of fatty acids. GC-EI/MS full scan analysis was suitable for the quantitative determination of all unsaturated fatty acids in both food samples, whereas GC-EI/MS in the SIM mode was particularly valuable for quantifying minor fatty acids. The novel GC-EI/MS/SIM method using fatty acid ethyl esters as internal standards can be used to quantify individual fatty acids only, that is, without determination of all fatty acids (the common 100% method), although this is present. This was demonstrated by the exclusive quantification of selected fatty acids including methyl-branched fatty acids, erucic acid (18:1n-9trans), and polyunsaturated fatty acids in cod liver oil and goat's milk fat.