Preparation of fatty acid methyl ester
Methylation of free fatty acids in the extracted lipids was achieved
by adding200μLof14%boron trifluoride in methanol(SigmaChemical
Co.) to the 600 μL extracts (500 μL hexane extract plus 100 μL
methanolic sodium hydroxide) and holding it at room temperature
(25 ◦C) for 30 min. Then, 200 μL of hexane and 100 μL of deionized
water were added, the samples were vortexed, and centrifuged in
a refrigerated (2 to 3 oC) microcentrifuge SFR13K-120 (Savant Instruments
Inc., Farmingdale, N.Y., U.S.A.) at 2000×g for 3 min. The
upper layer was recovered and stored in a sealed vial at –20 ◦C until
further analyses by gas chromatography.