Rt-2560 Columns Ensure Accurate, Reliable AOAC 996.06 and AOCS Ce 1j-07 FAMEs Analysis
Restek’s Rt-2560 GC column was originally developed for detailed analysis of FAMEs in foods and food products. Its highly polar biscyanopropyl stationary phase and long column format make it an effective tool for the difficult task of separating and accurately quantifying complex mixtures of FAMEs. Recently, Restek optimized the manufacturing process and implemented a new, application-specific, QC-testing procedure for all new Rt-2560 columns (cat.# 13198). As shown here, Rt-2560 columns produced using the optimized process exhibit excellent performance for both AOAC 996.06 and AOCS Ce 1j-07 FAMEs analysis and meet all method requirements.
AOAC 996.06
AOAC official method 996.06 describes a standard procedure for the determination of total, saturated, and unsaturated fat in foods using capillary GC-FID. The procedure involves hydrolytic extraction, methylation, and capillary GC-FID analysis of the resulting fatty acid methyl esters (FAMEs). Triundecanoin, the C11:0 triglyceride, is used as an internal standard for quantitation, and response factors are included in method AOAC 996.06.
The method recommends a 100 m x 0.25 mm x 0.20 µm column with a highly polar biscyanopropyl stationary phase, such as the Rt-2560. The column must be able to separate the adjacent C18:3 and C20:1 FAME isomers as well as C22:1, C20:3, and C20:4 with a resolution of 1.0 or better. The Rt-2560 is specifically designed for FAMEs analysis and readily achieves these required critical separations as shown by peaks 22 to 24 and 29 to 31 in Figure 1.
Figure 1: Application-specific QC testing ensures that all Rt-2560 columns meet AOAC 996.06 method requirements.

Peaks | tR (min) | Conc. (mg/mL) | Structural Nomenclature | |
---|---|---|---|---|
1. | Methyl butyrate | 13.16 | 40 | C4:0 |
2. | Methyl caproate | 15.53 | 40 | C6:0 |
3. | Methyl octanoate | 19.43 | 40 | C8:0 |
4. | Methyl decanoate | 24.50 | 40 | C10:0 |
5. | Methyl undecanoate | 27.20 | 20 | C11:0 |
6. | Methyl dodecanoate | 29.87 | 40 | C12:0 |
7. | Methyl tridecanoate | 32.47 | 20 | C13:0 |
8. | Methyl myristate | 34.97 | 40 | C14:0 |
9. | Methyl myristoleate | 37.01 | 20 | C14:1 (c9) |
10. | Methyl pentadecanoate | 37.37 | 20 | C15:0 |
11. | Methyl pentadecenoate | 39.36 | 20 | C15:1 (c10) |
12. | Methyl palmitate | 39.68 | 60 | C16:0 |
13. | Methyl palmitoleate | 41.30 | 20 | C16:1 (c9) |
14. | Methyl heptadecanoate | 41.86 | 20 | C17:0 |
15. | Methyl heptadecenoate | 43.45 | 20 | C17:1 (c10) |
16. | Methyl stearate | 43.97 | 40 | C18:0 |
17. | Methyl octadecenoate | 44.92 | 20 | C18:1 (t9) |
18. | Methyl oleate | 45.34 | 40 | C18:1 (c9) |
Peaks | tR (min) | Conc. (mg/mL) | Structural Nomenclature | |
---|---|---|---|---|
19. | Methyl linolelaidate | 46.39 | 20 | C18:2 (t9,t12) |
20. | Methyl linoleate | 47.32 | 20 | C18:2 (c9,c12) |
21. | Methyl arachidate | 47.92 | 40 | C20:0 |
22. | Methyl linolenate | 48.77 | 20 | C18:3 (c6,c9,c12) |
23. | Methyl eicosenoate | 49.20 | 20 | C20:1 (c11) |
24. | Methyl linolenate | 49.54 | 20 | C18:3 (c9,c12,c15) |
25. | Methyl heneicosanoate | 49.78 | 20 | C21:0 |
26. | Methyl eicosadienoate | 51.09 | 20 | C20:2 (c11,c14) |
27. | Methyl behenate | 51.60 | 40 | C22:0 |
28. | Methyl eicosatrienoate | 52.52 | 20 | C20:3 (c8,c11,c14) |
29. | Methyl erucate | 52.88 | 20 | C22:1 (c13) |
30. | Methyl eicosatrienoate | 53.28 | 20 | C20:3 (c11,c14,c17) |
31. | Methyl arachidonate | 53.42 | 20 | C20:4 (c5,c8,c11,c14) |
32. | Methyl tricosanoate | 53.65 | 20 | C23:0 |
33. | Methyl docosadienoate | 54.85 | 20 | C22:2 (c13,c16) |
34. | Methyl lignocerate | 55.32 | 40 | C24:0 |
35. | Methyl eicosapentaenoate | 56.09 | 20 | C20:5 (c5,c8,c11,c14,c17) |
36. | Methyl nervonate | 56.74 | 20 | C24:1 (C15) |
37. | Methyl docosahexaenoate | 62.17 | 20 | C22:6 (c4,c7,c10,c13,c16,c19) |
Column | Rt-2560, 100 m, 0.25 mm ID, 0.20 µm (cat.# 13198) |
---|---|
Standard/Sample | Food industry FAME mix (cat.# 35077) |
Diluent: | Hexane/dichloromethane |
Conc.: | 1,000 µg/mL |
Injection | |
Inj. Vol.: | 1 µL split (split ratio 20:1) |
Liner: | Premium 4 mm Precision liner w/wool (cat.# 23305) |
Inj. Temp.: | 225 °C |
Oven | |
Oven Temp.: | 100 °C (hold 4 min) to 240 °C at 3 °C/min (hold 15 min) |
Carrier Gas | He, constant flow |
Flow Rate: | 1.0 mL/min |
Detector | FID @ 285 °C |
---|---|
Make-up Gas Flow Rate: | 45 mL/min |
Make-up Gas Type: | N2 |
Hydrogen flow: | 30 mL/min |
Air flow: | 300 mL/min |
Data Rate: | 20 Hz |
Instrument | Agilent 7890A GC |
Conclusion
The improved manufacturing process and application-specific quality control test both ensure that Rt-2560 columns provide a consistently high level of performance in the detailed analysis of FAMEs. The Rt-2560 column meets method requirements for standard methods of fat speciation in food, including AOAC 996.06 and AOCS Ce-1j-07. In addition, Rt-2560 columns can be used for other methods that call for a 100 m high-percentage biscyanopropyl capillary GC column.