Essential fatty acids are nutrients that must be obtained from the diet because humans lack the anabolic processes for their synthesis. Two closely related groups of essential fatty acids are the omega 3 and omega 6 fatty acids. These unsaturated fatty acids contain the initial double bond located directly after the third (omega 3) or the sixth (omega 6) carbon atom, as measured from the methyl end of the compound.
Omega 3 and omega 6 fatty acids are typically analyzed using gas chromatography (GC) after their conversion to fatty acid methyl esters (FAMEs). Columns described in promulgated methods1,2 contain a stationary phase based on polyethylene glycol (PEG), and often have ‘wax’ in the name.
In this work, a new ionic liquid GC column, SLB®-IL60, is evaluated against an Omegawax® column for its suitability for this application. The SLB-IL60 column has selectivity similar to ‘wax’ columns, but is different enough to provide a unique elution pattern. It is not based on a PEG phase. Instead, it has various functional groups that allow for an increased number of interaction mechanisms compared to a PEG phase. Specifications for both columns are:
The GC conditions used are from the AOAC® 991.39 and AOCS® Ce 1i-07 methods. This was possible as both methods share the same set of run conditions. Two standard mixes were initially analyzed to gauge elution patterns. An individual FAME was also analyzed to confirm identification. Overall, the following standards were used:
A custom 38-component mix was prepared by the addition of C22:5n3 FAME to the stock 37-component FAME mix. The resulting mix was then analyzed on both columns under identical conditions. Resulting chromatograms are shown in Figure 1. Observations are that the SLB-IL60 column provides:
Figure 1.C4-C24 FAME Standard
column 1: Omegawax, 30 m x 0.25 mm I.D., 0.25 µm (Product No. 24136); column 2: SLB-IL60, 30 m x 0.25 mm I.D., 0.20 µm (Product No. 29505-U); oven: 170 °C, 1 °C/min to 225 °C; inj. temp.: 250 °C; carrier gas: helium, 1.2 mL/min; det.: FID, 260 °C; injection: 1 µL, 100:1 split; liner: 4 mm I.D., split/splitless type, single taper wool packed FocusLiner™ design; sample: Supelco 37-Component FAME Mix + C22:5n3, in methylene chloride (Product No. 17269)
| 1. C4:0
A qualitative polyunsaturated fatty acid (PUFA) methyl ester standard (made from menhaden oil) was also analyzed on both columns under identical conditions. Figure 2 shows both chromatograms. Use of the SLB-IL60 column resulted in:
Figure 2.PUFA No. 3 Standard
column 1: Omegawax, 30 m x 0.25 mm I.D., 0.25 µm (Product No. 24136); column 2: SLB-IL60, 30 m x 0.25 mm I.D., 0.20 µm (Product No. 29505-U); oven: 170 °C, 1 °C/min to 225 °C; inj. temp.: 250 °C; carrier gas: helium, 1.2 mL/min; det.: FID, 260 °C; injection: 1 µL, 100:1 split; liner: 4 mm I.D., split/splitless type, single taper wool packed FocusLiner™ design; sample: PUFA No. 3 Mix (Product No. 47085-U), diluted in 1 mL of hexane
| 1. C14:0
Compared to existing ‘wax’ columns, the SLB-IL60 column offers unique selectivity, some cis/trans resolution, and the faster elution of analytes. The elution of trans isomers before cis isomers of the same carbon chain length, plus degree and position of unsaturation, demonstrates the analyte-stationary phase mechanism difference of the SLB-IL60. These differences make the SLB-IL60 a good column choice for the analysis of omega 3 and omega 6 fatty acid methyl esters.