G.C. analysis of plasma free fatty acids


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Added on July 29, 2015 at 2:25 PM by Harris, Carla
Modified on October 19, 2017 at 3:29 PM by Harris, Carla
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HAASC: Lipid Core Master

Plasma Free Fatty Acids by G.C.

Free fatty acids are extracted from the plasma using heptane/isopropanol (30:70) (1) and separated from other lipids by thin layer chromatography on silica gel plates developed in petroleum ether/ethyl ether/acetic acid (80:20:1 v/v/v).  The free fatty acid band is visualized using rhodamine 6G (0.1% in ethanol), scraped from the TLC plate, and methylated with BF3/methanol(2) without elution from the silica gel. 

The fatty acid methyl esters are analyzed by gas chromatography using an Agilent 7890 gas chromatograph equipped with flame ionization detector and a capillary column (SP2380, 0.25 mm x 30 m, 0.20 µm film, Supelco, Bellefonte, PA).  Helium is used as a carrier gas.  The oven temperature is programmed from 160 °C to 230 °C at 4 °C/min.  Fatty acid methyl esters are identified by comparing the retention times to those of known standards.  Inclusion of a pentadecanoic acid (15:0) internal standard permits quantitation of the amount of FFA in the sample.

References

  1. Ko, H., and Royer, M. E.  1974. A gas-liquid chromatographic assay for plasma free fatty acids.  J. Chrom.  88:  253-263.
  2. Morrison, W. R., and Smith, L. M.  1964.  Preparation of fatty acid methyl esters and dimethylacetal from lipids with boron trifluoride-methanol.  J. Lipid Res.  5:  600-608.

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