2.2. Fatty Acid Component Analysis

TKO was filtrated with a 0.22 μm membrane filter after methyl esterification. The gas chromatography-mass spectrometer (GC-MS) analysis for TKO fatty acid components referenced the previous method with some modifications [11]. The gas chromatograph (Thermo Fisher Trace 1300, U.S.A.) was equipped with a DB-WAX column (30 m × 0.25 mm, 0.25 μm, U.S.A.). GC conditions are the following: high-purity nitrogen carrier gas as well as the sample injection volume was 1 μL with a split ratio of 1 : 20. The temperature rise procedure is the following: initial temperature of 140°C for 1 min, then increased to 250°C at a rate of 4°C/min for 15 min. According to the retention time of fatty acid methylated gas phase standard (Sigma), compare the peak area measured by the sample with the retention time. The peak area of each fatty acid can be used as the relative content of each fatty acid component in the total fatty acid.