Food Lipids: Chemistry, Nutrition, and Biotechnology

in the transition periods in spring and late autumn. The average contents were (c9t11-
18:2) 0.76%, (trans-C18:1) 3.67%, (trans-C18:2) 1.12%, and (total trans fatty acids)
4.92%. High correlation coefficients (r) were reported between the content of the
CLA isomer c9t11-18:2 and the contents of trans-C18:1, trans-C18:2, total trans
fatty acids, and C18:3 of 0.97, 0.91, 0.97, and 0.89, respectively. A probable metabolic
pathway in the biohydrogenation of linolenic acid was also reported: c9c12c15
→ c9t11c15 → t11c15 → t11 [65].
3. Equivalent Chain Length
It is difficult to identify peaks in gas chromatograms, mainly because of the complexity
of the fatty acid composition of hydrogenated vegetable and fish oils. Some
of the problems of using capillary GC for qualitative and quantitative analysis of
hydrogenated oils have been reviewed [56]. The major identification problem is a
result of the large number of fatty acid isomers present. One difficulty in identifying
peaks is due to the fact that some polyunsaturated fatty acids can have longer retention
times than the next longer chain fatty acid. This happens most frequently on
highly polar columns used to separate fatty acid isomers.
One way to predict where a certain fatty acid will elute on a GC column is
based on the so-called equivalent chain length (ECL). The use of ECL to identify
fatty acid isomers on GC columns has been reviewed [56,66]. It was reported that
ECLs are constants for a specific carrier gas and column and are independent of
experimental conditions [67]. A specific fatty acid can be characterized by obtaining
its ECL on both polar and nonpolar columns. The ECL consists of one or two integers
indicating the positional chain length, and two numbers after the decimal that indicate
the fractional chain length (FCL). The ECL values for fatty acids on a specific
column are determined by first using semilog paper to plot the log of the retention
times (y axis) against those of the saturated fatty acids (e.g., 16:0 = 16.00, 18:0 =
18.00, 20:0 = 20.00) on the x axis observed under isothermal conditions. Using the
same column, the retention time (y-axis value) of an unsaturated fatty acid is then
plotted to find its ELC on the x axis. For example, the ECL of linoleic acid (18:2n-
6) was found to be 18.65. The FCL value was 0.65 (18.65 � 18.00). Using these
data, the ECL for 20:2n-6 was predicted to be 20.65 (20.00 � 0.65). The actual ECL
of 20:2n-6 was experimentally determined to be 20.64 [36]. Equivalent chain length
data from a GC analysis of rapeseed oil showed that the cis-11 20:1 fatty acid
coelutes with trans triene isomers at oven temperatures above 160�C [68].
4. Relative Response Factors
Area normalization is often used to report the relative concentration of fatty acid
isomers in an oil mixture. This method is accurate only if all isomers exhibit the
same response in the GC detector. However, all the fatty acid isomers do not give
the same relative response in the GC flame ionization detector. Relative response
factors, first proposed in 1964 [69], were found to depend on the weight percent of
all carbons except the carbonyl carbon in the fatty acid chain. A later study confirmed
these findings and concluded that the required corrections were not insignificant,
even though they were often not used [70]. Failure to use a correction factor could
result in an error of about 6% (relative to 18:0) in the case of 22:6. Table 2 summarizes
some of the correction factors recommended.
Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.