A comparison of methods for the high-performance liquid chromatographic and capillary gas-liquid chromatographic analysis of fatty acid esters.

A reversed-phase high-performance liquid chromatographic (RP-HPLC) method for the analysis of free fatty acids in plasma was compared with a method using capillary gas-liquid chromatography (GLC). The same extraction procedure was used for both assays. In the RP-HPLC method, the acids were separated as their anthrylmethyl esters on a C18 reversed-phase column, and detected by fluorescence. The coupling agent 2-bromo-1-methylpyridinium iodide was used with 9-(hydroxymethyl)anthracene. A mobile phase of acetonitrile-water (98:2) was used with flow programming. The derivatives of the C14:0, C16:1 and C18:2 acids could not be fully resolved. For capillary GLC, the acids were separated as their methyl esters following on-column injection into a 25-m OV-101 glass capillary column and detected using flame ionization detection. The esterifying agent used was diazomethane. The C18:2 and C18:3 esters were not fully resolved. The precision and sensitivity of both methods were similar. In an application of the methods, the free fatty acid concentrations in the plasma of a group of diabetic patients and their age-matched controls were estimated. Fatty acid concentrations tended to be higher in the diabetic group but, in the small number of patients studied, wide inter-individual variations prevented a significant difference from being detected. Estimates of individual fatty acids were higher by the RP-HPLC method. The identity of the acids in the extract was confirmed by gas chromatography-mass spectrometry of their methyl esters.