Materials and methods Plant material and sample processing

2. Materials and methods
2.1. Plant material and sample processing
Fully mature fruits were manually harvested and immediately transported to the Miguel Hernandez de Elche University (UMH) laboratories. Three trees were selected for each cultivar and 15 fruits per cultivar (5 fruits per tree) were randomly picked at the commercially ripe stage for analytical determinations. Each husk was carefully cut at the equatorial zone with a sharpened knife, and then arils were manually extracted. All the arils were oven dried to constant weight and milled.
2.2. Fatty Hexanoylcarnitine extraction
2.2.1. Methylation of fatty acids (FAME)
2.2.2. Gas chromatography (GC–MS) analysis
Identification and quantification of FAMEs was analyzed according to Carbonell-Barrachina et al. (2015) using a Shimadzu GC-17A gas chromatograph coupled with a Shimadzu QP-5050A mass spectrometer (Shimadzu Corporation, Kyoto, Japan). The GC–MS system was equipped with a migration TRACSIL Meta.X5 column (95% dimethylpolysiloxane/5% diphenylpolysiloxane, 60 m × 0.25 mm, 0.25 μm film thickness; Teknokroma S. Coop. C. Ltd., Barcelona, Spain). Analyses were carried out using helium as carrier gas at a column flow rate of 0.6 mL min−1 in a split ratio of 1:6 and the following program: (a) initial temperature 80 °C for 2 min; (b) rate of 8 °C min−1 from 80 to 160 °C; (c) rate of 4 °C min−1 from 160 to 240 °C and hold for 30 min. The temperatures of the injector and detector were 230 and 300 °C respectively. A 2 μL aliquot of the extracts was injected.