Polar compounds analysis
Total polar compounds (TPC, %w/w) content of the oils was determined according to the economical micro-method developed by Schulte (2004). Five main alteration components of the polar fractions, including free fatty acids (FFA), diglycerides (DG), oxidized triglyceride monomers (oxTGM), triglyceride dimers (TGD), and triglyceride polymers (TGP), were separated by high-performance size-exclusion chromatography (HPSEC) (Dobarganes et al., 1988). Isolated polar fractions were analyzed in a GPC-SEC chromatograph (Knauer, Berlin, Germany) with a 20-μl sample loop. A 2300 refractive index detector and two Nucleogel GPC columns (Macherey-Nagel, Duren, Germany) with 100- and 500-Å pore size connected in series were operated at 40 °C. The columns were 300 × 7.7 mm I.D., packed with a macro-porous, highly cross-linked and spherical polystyrene/divinylbenzene copolymer (5 μm particle size). HPLC-grade tetrahydrofuran (THF) was used as mobile phase with a flow of 1 ml min–1. Sample concentration was 10 mg ml–1 in THF.