, Volume 35, Issue 10, pp 1147–1153

Comparison of silver-ion high-performance liquid chromatographic quantification of free and methylated conjugated linoleic acids

  • Ewa Ostrowska
  • Frank R. Dunshea
  • Morley Muralitharan
  • Reginald F. Cross


Silver-ion high-performance liquid chromatography was used to fractionate a mixture of conjugated linoleic acid (CLA) isomers (as the free fatty acids, CLAFFA) in commercial CLA mixtures and biological samples. Due to the unchanged retention mechanism, it was assumed that the elution order of the isomers remained the same as that of methyl esters separated on the same column. The most abundant isomers, cis/trans 10,12-18:2 and cis/trans 9,11-18:2, were separated better as free acids on a single column than in the methyl ester form. Quantification of the CLA standard was used as the reference profile to evaluate different methylation methods commonly used to prepare CLA methyl esters for quantitation. Acid- and vuigi base-catalyzed derivatization methods resulted in CLA intraisomerization and losses in total conjugated dienes content. Acid (HCl and BF3) methylations significantly elevated the level of trans,trans isomers and significantly reduced the cis/trans isomers. Base methylation, tetramethylguanidine/methanol, resulted in loss of trans,trans isomers, and a substantial loss of total underivatized conjugated dienes. Other catalysts such as the trimethylsilyldiazomethane produced additional peaks of unidentified artifacts. The analysis of CLAFFA appears to provide more accurate quantification of CLA isomers in commercial and biological samples.





silver-ion high-performance liquid chromatography


allylic hydroxy oleate


conjugated linoleic acid


all the CLA isomers having a cis,trans or trans,cis configuration


conjugated linoleic acid as the free fatty acids


fatty acid methyl esters


free fatty acid


gas chromatography








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Copyright information

© AOCS Press 2000

Authors and Affiliations

  • Ewa Ostrowska
    • 3
    • 1
  • Frank R. Dunshea
    • 1
  • Morley Muralitharan
    • 2
  • Reginald F. Cross
    • 3
  1. 1.Agriculture VictoriaVictorian Institute of Animal ScienceWerribeeAustralia
  2. 2.Charles Sturt UniversityWagga WaggaAustralia
  3. 3.School of Engineering and ScienceSwinburne University of TechnologyHawthornAustralia

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