trans Fat: Resolving cis and trans FAME Isomers by GC

• Highly polar Rt-2560 column resolves individual cis and trans FAME isomers.
• Analytical reference mixes for quantifying FAMEs in foods and dietary supplements.
• Use column and reference mixes to meet new trans fat labeling regulations.

Concern over the detrimental effects of diets high in trans fats has prompted the US Food and Drug Administration (FDA) to require trans fat content to be reported separately on food labels after January 2006. The FDA estimates that by 2009 this rule will save $900 million to $1.8 billion per year in medical costs and lost productivity. The monetary savings will far more than offset the FDA-estimated $140-250 million in one-time costs of determining amounts of trans fats, revising Nutrition Facts panels, and voluntarily reducing amounts of trans fats(1) that the food industry will incur to comply with the rule.

The highly polar Rt-2560 biscyanopropyl stationary phase has the selectivity needed for resolving cis and trans FAME isomers to comply with the FDA guidelines. Individual cis and trans isomers are resolved on a 100-meter Rt-2560 GC column (cat.# 13199), making this the column of choice for analyzing partially hydrogenated fats. The trans isomers elute before the cis isomers (Figure 1), a reverse of the elution order on Carbowax®-based phases such as FAMEWAX™ or Rtx®-Wax. AOAC method 996.06 (2) specifies the determination of total fat content based on the fatty acid content after conversion of the fatty acids to the methyl esters, and is the accepted analytical method for determining total fat content for nutritional labeling. A 100-meter Rt-2560 column meets the requirements of this procedure, and also allows quantification of the total trans fat content.

To calibrate the GC system for these assays, we recommend a carefully formulated FAME mixture, such as our 37-component Food Industry FAME Mix (cat.# 35077, Figure 1) or our 28-component NLEA FAME Mix (cat.# 35078). Each of these mixes includes a gravimetric certificate of analysis to help ensure accurate quantification. To ensure correct identifications of individual C18:1 cis or trans isomers, use our cis/trans FAME Mix (cat.# 35079), as shown in Figure 2.

An Rt-2560 column is the column of choice when determining trans fat content and total fat content in food products. Whatever your fatty acid analysis requirements, Restek can provide the consistent-performance analytical columns and reference materials that will help you to accurately characterize your materials.

Figure 1  Food Industry FAME Mix (cat.# 35077) resolved on an Rt-2560 column.

Peaks % in Mix 1. C4:0 Methyl butyrate 4.0 2. C6:0 Methyl hexanoate 4.0 3. C8:0 Methyl octanoate 4.0 4. C10:0 Methyl decanoate 4.0 5. C11:0 Methyl undecanoate 2.0 6. C12:0 Methyl laurate 4.0 7. C13:0 Methyl tridecanoate 2.0 8. C14:0 Methyl myristate 4.0 9. C14:1 Methyl myristoleate (cis-9) 2.0 10. C15:0 Methyl pentadecanoate 2.0 11. C15:1 Methyl pentadecanoate (cis-10) 2.0 12. C16:0 Methyl palmitate 6.0 13. C16:1 Methyl palmitoleate (cis-9) 2.0 14. C17:0 Methyl heptadecanoate 2.0 15. C17:1 Methyl heptadecenoate (cis-10) 2.0 16. C18:0 Methyl stearate 4.0 17. C18:1 Methyl elaidate (trans-9) 2.0 18. C18:1 Methyl oleate (cis-9) 4.0 Peaks % in Mix 19. C18:2 Methyl linoleaidate (trans-9,12) 2.0 20. C18:2 Methyl linoleate (cis-9,12) 2.0 21. C20:0 Methyl arachidate 4.0 22. C18:3 Methyl γ-linolenate (cis-6,9,12) 2.0 23. C20:1 Methyl eicosenoate (cis-11) 2.0 24. C18:3 Methyl linolenate (cis-9,12,15) 2.0 25. C21:0 Methyl heneicosanoate 2.0 26. C20:2 Methyl eicosadienoate (cis-11,14) 2.0 27. C22:0 Methyl behenate 4.0 28. C20:3 Methyl eicosatrienoate (cis-8,11,14) 2.0 29. C22:1 Methyl erucate (cis-13) 2.0 30. C20:3 Methyl eicosatrienoate (cis-11,14,17) 2.0 31. C20:4 Methyl arachidonate (cis-5,8,11,14) 2.0 32. C23:0 Methyl tricosanoate 2.0 33. C22:2 Methyl docosadienoate (cis-13,16) 2.0 34. C24:0 Methyl lignocerate 4.0 35. C20:5 Methyl eicosapentaenoate (cis-5,8,11,14,17) 2.0 36. C24:1 Methyl nervonate (cis-15) 2.0 37. C22:6 Methyl docosahexaenoate (cis-4,7,10,13,16,19) 2.0 GC_FF00649 Column Rt®-2560, 100 m, 0.25 mm ID, 0.20 µm (cat.# 13199) Sample Food Industry FAME Mix (cat.# 35077) Diluent: methylene chloride Conc.: 30mg/mL total FAMEs Injection Inj. Vol.: 2.0 µL split (split ratio 200:1) Liner: Splitless (4mm ID) (cat.# 20814) Inj. Temp.: 225 °C Oven Oven Temp: 100 °C (hold 4 min.) to 240 °C at 3 °C/min. (hold 10 min.) Carrier Gas H2, constant flow Flow Rate: 1.2 mL/min. Detector FID @ 250 °C ,

Figure 2  An Rt-2560 column resolves cis C18 FAME isomers from trans C18 isomers.

Peaks Conc. (wt.%) 1. C18:0 methyl stearate 20.0 2. C18:1 methyl petroselaidate (trans-6) 8.0 3. C18:1 methyl elaidate (trans-9) 10.0 4. C18:1 methyl transvaccenate (trans-11) 12.0 Peaks Conc. (wt.%) 5. C18:1 methyl petroselinate (cis-6) 8.0 6. C18:1 methyl oleate (cis-9) 10.0 7. C18:1 methyl vaccenate (cis-11) 12.0 8. C18:2 methyl linoleate (cis-9,12) 20.0 GC_FF00652 Column Rt®-2560, 100 m, 0.25 mm ID, 0.20 µm (cat.# 13199) Sample cis/trans FAME Mix (cat.# 35079) Diluent: methylene chloride Conc.: 10 mg/ml total FAMEs Injection Inj. Vol.: 1 µL split (split ratio 20:1) Liner: Splitless (4mm ID) (cat.# 20814) Inj. Temp.: 225 °C Oven Oven Temp: 100 °C (hold 4 min.) to 240 °C at 3 °C/min. (hold 10 min.) Carrier Gas H2, constant flow Flow Rate: 1.2 mL/min. Detector FID @ 250 °C ,

References

1. www.cfsan.fda.gov/~dms/qatrans2.html
2. Official Methods of Analysis, 17th ed., AOAC International, 2000.

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