2.3 Fatty acid analysis
The fatty acid composition was determined as fatty acid methyl esters (FAMEs) by capillary gas chromatography analysis after alkaline treatment (Christie, 1982). Methyl tridecanoate (C13:0, 2 mg/mL) was used as internal standard and FAMEs were measured on a GC 2010 Plus gas chromatograph (Shimadzu Corporation, Kyoto, Japan) equipped with a flame ionisation detector (FID) and an AOC-20s auto sampler (Shimadzu Corporation), at the same conditions reported in Marzocchi, Pasini, Baldinelli & Caboni (2018). Peak identification was accomplished by comparing peak retention time with GLC-463 standard mixture from Nu-Check (Elysian, MN, USA) and FAME 189-19 standard mixtures from Sigma-Aldrich Chemicals (St. Louis, MO, USA) and expressed as weight percentage of total FAMEs. FAMEs composition was measured in 2 replicates for each lipid extract (n = 4) and each analysis lasted 7 minutes.