Comparison of Methylation Methods for the Quantitation of Conjugated Linoleic Acid Isomers

Abstract

Abstract Four methylation methods were evaluated for use in the gas chromatographic (GC) quantitation of conjugated linoleic acid (CLA) isomers, which are potential anticarcinogen. The methods were (1) sodium methoxide in methanol (NaOMe-MeOH), (2) American Oil Chemists' Society (AOCS) procedure Ce 2-66, which involves methanolic sodium hydroxide followed by boron trifluoride in methanol, (3) tetramethylguanidine in methanol (TMG-MeOH), and (4) direct transesterification with methanolbenzene- acetyl chloride (DAC). Purified methyl esters of isomerized linoleic acid containing 86% CLA isomers were methylated and analyzed by GC. The AOCS and DAC methods resulted in 3 and 50% losses in cis-9,trans-11-octadecadienoic acid (9c, 111 CLA isomer) and trans-10,cis-12 octadecadienoic acid (10t, 12c CLA isomers), respectively. Compared with the control, the AOCS and DAC methods increased the yield of the trans,trans CLA isomers (trans-9,trans-11- and trans-10, trans-12-octadecadienoic acid) by 1.07-fold and a 10-fold, respectively. A non-CLA artifact that eluted close to CLA peaks was formed during methylation by the AOCS and DAC methods. Thus, the DAC and AOCS methods are not suitable for quantitation of CLA isomers. The NaOMe-MeOH and TMG-MeOH methods, however, are suitable for quantitation of CLA isomers in fats containing low concentrations of free fatty acids.