Fatty acid methyl ester analysis

Oil extraction from the ground soybean seed powder (100 mg) was extracted twice with hexane (5 mL) in an ultrasonic bath (power 600 watts) for a period of 15 min. The extracts were centrifuged at 5000 rpm for 10 min and the supernatant was collected in a separate vial. The residue was re-extracted with 5 mL fresh hexane. The pool supernatant was evaporated to dryness under slow stream of nitrogen gas. The concentrated soybean oil was re-suspended with 2 mL hexane. A partial aliquot of one mL was evaporated to dryness and transesterified to FAMEs using 5 mL of acidified methanol (10 mL of acetyl chloride to 90 mL of cold methanol). The mixture was stirred at ambient temperature overnight under inert nitrogen atmosphere. To the above mixture, 3 mL of water was added and the fatty acid methyl esters were extracted with 2 mL of hexane. The hexane layer was separated and analyzed with GC [28]. Fatty Acid Methyl Ester were characterized by comparison of detention time with an authentic FAMEs standard. All analyses are conducted in triplicates and standard errors are calculated.