Evaluation of two methods for plasma fatty acid analysis by GC

The present study was designed to establish the best method for the quantitative analysis of fatty acids in blood plasma. In the “two steps” method, internal standards of C15:0, C19:0, C19:1, or C21:0 fatty acids were added to plasma and lipids were extracted in chloroform/methanol (2:1). Lipid extracts resuspended in methanol/toluene (4:1) were subjected to methanolysis at 80°C for 2.5 h with acetyl chloride for GC analysis. In the “direct” method, plasma containing internal standards were suspended in methanol/toluene and after methanolysis at 100°C for 1 h with acetyl chloride were processed as above. When C15:0 was used to calculate recoveries, values were above 100%, these being higher when using the “direct” method. When methylated C17:0 was the reference standard, the peak area ratios between internal standards and methylated reference were lower for C15:0 than for any of the others. When using C19:0 as the internal standard, quantitative values of individual fatty acids were higher when the “two steps” method was used as compared to the “direct” method. Thus, the “two steps” method is more appropriate and reliable for the quantitative analysis of fatty acid profiles in plasma samples, and C19:0 but not C15:0 should be used as the internal standard.