Enantioselective determination of anteiso fatty acids in food samples

Abstract

Anteiso fatty acids (aFAs)-long-chain carboxylic acids with a methyl branch on the (n - 2)-carbon-are among the most simple fatty acids that are chiral. The most frequently occurring aFAs in food are 12-methyltetradecanoic acid (a15:0) and 14-methylhexadecanoic acid (a17:0), structures where the asymmetric carbon is more than 10 carbons separated from the polar head group. Previously, only enantioseparation of 4-methyl-substituted carboxylic fatty acids has been reported by gas chromatography. Here we present the first direct partial enantioresolution of synthesized racemic a15:0-a17:0 on a capillary column coated with 50% heptakis(6-O-tert-butyldimethylsilyl-2,3-di-O-methyl)-beta-cyclodextrin diluted in OV1701. Synthesized (S)-(+)-enantiomers were used to demonstrate that the elution order was (R)- prior to (S)-enantiomers. Using this system, food samples (butter, goat's milk fat, suet, human milk, seal oil, cod liver oil) known to contain aFAs were analyzed. Prior to the enantioselective gas chromatography, unsaturated fatty acids were preseparated by urea complexation, silver ion high performance liquid chromatography (Ag+-HPLC), or both from food samples. The fractions of the food samples enriched with methyl-branched fatty acids were then analyzed by GC/MS in the SIM mode. The measurements confirmed that the (S)-enantiomer of a15:0 (ee >96%), a16:0, and a17:0 (ee >90%, respectively) dominated in all samples. While the (R)-enantiomers could not be identified in samples from ruminants and human milk, their presence could be established in cod liver and seal oil (ee <86%).