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Abstract
Conventional sample preparation for fatty acid analysis is a complicated, multiple-step process, and gas chromatography (GC) analysis alone can require >1 h per sample to resolve fatty acid methyl esters (FAMEs). Fast GC analysis was adapted to human plasma FAME analysis using a modified polyethylene glycol column with smaller internal diameters, thinner stationary phase films, increased carrier gas linear velocity, and faster temperature ramping. Our results indicated that fast GC analyses were comparable to conventional GC in peak resolution. A conventional transesterification method based on Lepage and Roy was simplified to a one-step method with the elimination of the neutralization and centrifugation steps. A robotics-amenable method was also developed, with lower methylation temperatures and in an open-tube format using multiple reagent additions. The simplified methods produced results that were quantitatively similar and with similar coefficients of variation as compared with the original Lepage and Roy method. The present streamlined methodology is suitable for the direct fatty acid analysis of human plasma, is appropriate for research studies, and will facilitate large clinical trials and make possible population studies.
Highlights
Conventional sample preparation for fatty acid analysis is a complicated, multiple-step process, and gas chromatography (GC) analysis alone can require Ͼ1 h per sample to resolve fatty acid methyl esters (FAMEs)
In as much as dietary intakes of n-3 polyunsaturated fatty acids (PUFAs) are well indicated in the blood compartment [19, 20], it has been proposed that n-3 highly unsaturated fatty acid (HUFA) status in blood may be an important, modifiable, clinical biomarker for the prevention of various chronic diseases [21, 22]
The organic phases were combined and evaporated under nitrogen to a volume of 60 l. This solution was transferred to a GC vial, and the vial was crimped under nitrogen for FAME analysis by GC
Summary
Conventional sample preparation for fatty acid analysis is a complicated, multiple-step process, and gas chromatography (GC) analysis alone can require Ͼ1 h per sample to resolve fatty acid methyl esters (FAMEs). Analysis of the fatty acid composition of human blood and plasma samples involves tedious and time-consuming extraction and transesterification procedures [3,4,5,6,7,8]. The effect of extending heating times from 60 min to 120 min for each transesterification on concentrations of fatty acids as determined by fast GC FAME analysis was examined.
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