Analysis of Fatty Acids in Food Lipids

Abstract

Analysis of Fatty Acids in Food Lipids Fatty acid compositional analysis of lipids is usually carried out by gas-liquid chromatography (GC). Lipids obtained from food samples have complex structures and often fall into the following classes: triacylglycerols, phospholipids, sphingolipids, and sterols. Sterols exist in both animal-and plant-originated foods in either a free form (e.g., free cholesterol) or esterified form (e.g., cholesterol esters). Generally, most of the fatty acids found in food lipids are covalently bound to either an alcohol (glycerol) via ester bonds (triacylglycerols, phospholipids, and sterol esters), or to a long-chain base (sphingosines) via amide bonds (sphingolipids). To analyze the fatty acid composition of food lipids, the complex lipids must be pretreated so that the individual fatty acids are available for chromatographic analysis. For this purpose, fatty acids in the complex lipids are converted to their corresponding methyl esters by various derivatization methods to make them volatile for GC analysis. Since the focus of this unit is fatty acid analysis, and fatty acids from sterol esters contribute only a small portion of the total fatty acid pool, issues typical to the analysis of sterol-bound fatty acids will not be discussed; however, it should be noted that the protocols described in this unit do not work well for fatty-acid esterified sterols. There are basically two mechanisms to convert the fatty acids in a complex lipid to fatty acid methyl esters (FAMEs): methylation following hydrolysis of the fatty acids from the complex lipids, or direct transesterification. The first mechanism involves saponification (alkaline hydrolysis) in which the ester bond is cleaved between the fatty acid and the glycerol moiety (e.g., triacylglycerols and phospholipids) under heat and in the presence of an alkali (usually sodium hydroxide), followed by methylation performed in the presence of an acidic catalyst in methanol. Direct transesterification is usually a one-step reaction involving alkaline or acidic catalysts. This unit describes two basic protocols and one alternate protocol for FAME derivatization, and one support protocol for lipid extraction. The derivatization procedures described in this unit are suitable for a small amount of sample, ranging from 2 to 50 mg, which is particularly compatible with GC analysis using today's popular capillary or megabore fused-silica columns. In the first protocol, FAMEs are prepared from lipid samples by sodium hydroxide saponification followed by boron trifluoride (BF 3)-catalyzed esterification (see Basic Protocol 1). This procedure is adopted from the official methods (AOCS, 1989; Firestone, 1995) with modification. Basic Protocol 1 is suitable for most lipid analyses. Since BF 3 and other acidic catalysts will change the conformation of conjugated dienoic fatty acids (e.g., conjugated octadecadienoic acids; conjugated linoleic acids; CLA), this method is not recommended for lipid samples having these special structures, such as dairy product...

Implications of Method Chosen for Analysis of Fatty Acids in Meat: A Review

“…In accordance with Li and Watkins (2001), the procedure adopted by AOCS, 1989 and(Stark andFirestone, 1996), in the preparation of FAME, occurs if the small sample from lipids (2-50 mg), by saponification with sodium hydroxide, is followed by esterification catalyzed by boron trifluoride (BF 3 ). This protocol fits most analysis of lipids.…”

“…As research on Conjugated Linoleic Acid (CLA) and other fatty acid conjugates, have become more popular, it is essential not to leave missing information on the analysis of its composition, due to improper application protocol methylation (Li and Watkins, 2001). The CLA isomers, occur naturally in food derived from ruminants (Sun et al, 2009) being produced as intermediates of the biohydrogenation of polyunsaturated fatty acids, specifically linolenic (C18: 2 cis-9, cis-12) and α-linolenic acid (C18: 3 cis-9, cis-12, cis-15) by ruminal bacteria (Luna et al, 2008;Fuentes et al, 2006).…”

“…Thus, with increasing interest in the cis and trans isomers of conjugated fatty acids, where an accurate determination of the isomers is desired, the basic catalysis, performed at room temperature is considered as the most reliable identification of conjugated fatty acid isomers, such as CLA, isomers, or do not produce artifacts and reduce the loss of short-chain fatty acids (Fuentes et al, 2006;Palmiquist and Jenkins, 2003;Li and Watkins, 2001). …”

“…In accordance with Li and Watkins (2001), there is no single method that works optimally in all situations. The researcher must know the nature of the sample and select the appropriate method.…”

“…Therefore, the fatty acids from lipid complexes must be converted into their corresponding methyl esters, turning them volatile and thus facilitating the GC analysis. Various methods of derivatization may be employed for the esterification of fatty acids (Eder, 1995, Li and Watkins 2001, Millinsk et al, 20082011).…”