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Journal of Lipid Research, Vol. 46, 2299-2305, October 2005
Copyright © 2005 by American Society for Biochemistry and Molecular Biology

Methods

A simplified and efficient method for the analysis of fatty acid methyl esters suitable for large clinical studies

Athar Masood^*, Ken D. Stark and Norman Salem, Jr.^1,*

^* Laboratory of Membrane Biochemistry and Biophysics, National Institutes on Alcohol Abuse and Alcoholism, National Institutes of Health, Rockville, MD
Laboratory of Nutritional and Nutraceutical Research, Department of Kinesiology, University of Waterloo, Waterloo, Ontario, Canada

Published, JLR Papers in Press, August 1, 2005. DOI 10.1194/jlr.D500022-JLR200

¹ To whom correspondence should be addressed. e-mail: nsalem{at}niaaa.nih.gov

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.

Abbreviations: BHT, butylated hydroxytoluene; DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid; FAME, fatty acid methyl ester; FID, flame ionization detector; GC, gas chromatography; HUFA, highly unsaturated fatty acid; RRF, relative response factor

Supplementary key words fatty acid analysis • plasma • transesterification method • robotic chemistry • fast gas chromatographic analysis

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