Liver Fatty Acid Quantification by GC-FID

Fatty acid composition was analysed in frozen liver samples by gas chromatography with a flame ionization detector as described previously.^{(33 (link))} Fatty acid methyl esters (FAMEs) were synthesized by incubation with 0.5 M sodium methoxide, and the fatty acid composition was calculated as a percentage of the total identified FAMEs. Analysis of FAMEs was conducted in a 7820A Agilent gas chromatograph (Agilent Technologies Spain) equipped with an Agilent HP-23 capillary column (60 m × 250 µm × 0.25 µm, Agilent Technologies Spain) and helium (1.0 ml/min) as the carrier gas. The temperature protocol was as follows: initial temperature 100°C, ramp 8°C/min to 145°C (20 min), ramp 5°C/min to 195°C (5 min), ramp 5°C to 215°C (15 min), ramp 5°C to 230°C (5 min). Chromatograms were recorded and analysed using Agilent EZChrom Elite software (Agilent Technologies Spain). The method was validated with original standards for every fatty acid quantified. Additionally, tridecanoic acid (C13:0) was used as an internal standard, and in every case, a response factor of 1 was used.