GAS-CHROMATOGRAPHIC ANALYSIS OF FATTY-ACID METHYL-ESTERS

The full process of fatty acid methyl ester (FAME) analysis consists o f esterification of lipids, and of injection, separation, identificati on and quantitation of the FAMEs. In order for the required accuracy a nd precision to be attained, each of these steps has to be optimized. Esterification of lipids can be carried out with several reagents base d on acid-catalysed or base-catalysed reactions. The advantages and di sadvantages of these reagents are discussed. The most critical step in the gas chromatographic analysis of FAMEs is sample introduction. The classical split injection technique, which is the most widely used te chnique in the analysis of FAMEs, has the potential disadvantage of bo iling-point-dependent sample discrimination. Cold injection of the sam ple, either on-column or by programmed-temperature vaporization, does not present this problem and should therefore be preferred. Modern, co mmercially available fused-silica capillary columns offer excellent se paration of FAMEs from biological samples. Very polar stationary phase s give excellent separation of all FAMEs but have relatively low therm al stability, resulting in long retention times. Non-polar phases have a much greater thermal stability but inferior selectivity. For many a nalyses, phases of intermediate polarity, which combine the advantages of a relatively high resolution capability with relatively high therm al stability, are the most suitable. FAMEs can be identified by compar ison of their retention times with those of individual purified standa rds or secondary standards based on lipids that have been well charact erized in literature. Relative retention times and equivalent chain-le ngth values also provide useful information. FAMEs can be quantitated by peak areas via calibration factors, and absolute concentrations can be determined by adding an internal standard. Among numerous applicat ions in biomedical research, the analysis of fatty acids from body tis sues may contribute to the understanding of the link between the dieta ry intake of fatty acids and the diseases with which these acids are a ssociated.