Cp sil 88 for fame

Avocado oil (301 mg) was transesterified by ISO method 5509:1978 (21) to obtain the methyl esters of fatty acids. In a tube, the sample and 5 mL of n-heptane were added. The tube was vortexed for 1 min, following which 1 mL of a 0.5 mol L⁻¹ NaOH methanol solution was added and then vortexed again for 1 min. The upper organic phase was collected, anhydrous sodium sulfate was added and the solution was filtered.
The fatty acid methyl esters were determined on a Shimadzu gas chromatograph (GC 2010-Plus, Shimadzu, Kyoto, Japan) with a split/splitless injector, AOC-i-20 auto-injector, and flame ionization detection. A fused silica capillary column was used (CP-SIL 88 for FAME; 100 m × 0.25 mm × 0.2 μm, Agilent Technologies, Palo Alto, CA, USA). The chromatographic conditions were as follows: Injection volume of 1.0 μL in split mode 1:100, injector temperature at 250 °C, detector temperature at 260 °C, initial oven temperature set to 100 °C and held for 5 min, increased from 4 °C/min to 200 °C, and waited 30 min after reaching the final temperature. Hydrogen was used as the entrainment gas with a flow rate of 1.0 mL/min and a pressure of 140.3 kPa. Quantification of the fatty acid methyl esters was done by area normalization and the concentrations were expressed in g per 100 g sample (Table 2) [20 (link)].

Gas chromatography–mass spectrometry (GC/MS), using an Agilent 7890B/7000D (Agilent Technologies, Santa Clara, CA, USA), was performed for measurement of the concentration of palmitic acid in murine sera, liver, feces, and plantaris muscle. Fecal pellets were collected by resection of the distal colon during sacrifice. A total of 25 milliliters of sera and 15 µg of the liver, feces, and plantaris muscle were methylated using a fatty acid methylation kit (Nacalai Tesque, Kyoto, Japan). The final product was loaded onto a Varian capillary column (DB-FATWAX UI; Agilent Technologies). For fatty acid separation, the capillary column used was CP-Sil 88 for FAME (length, 100 mm; inner diameter, 0.25 mm; membrane thickness, 0.20 μm; Agilent Technologies). The column temperature was held at 100 °C for 4 min, gradually increased to 240 °C at 3 °C/min, and maintained for 7 min. Samples were injected in split mode at a ratio of 5:1. Palmitic acid methyl ester was detected in the selective ion monitoring mode. All results were normalized to the peak height of the C17:0 internal standard [36 (link)].

The composition of fatty acids in murine epididymal adipose tissue and serum was measured by gas chromatography–mass spectrometry (GC/MS), Agilent 7890B/5977B (Agilent Technologies, Santa Clara, CA, USA). A total of 15 µg of epididymal adipose tissue and 25 µl of serum were methylated with a fatty acid methylation kit (nacalai tesque, Kyoto, Japan). The final product was loaded onto a Varian capillary column (DB-FATWAX UI; Agilent Technologies). The capillary column used for fatty acid separation was CP-Sil 88 for FAME (100 m × an inner diameter of 0.25 mm × membrane thickness of 0.20 μm, Agilent Technologies). The column temperature was maintained at 100°C for 4 min and then increased gradually by 3°C/min to 240°C and held there for 7 min. The sample was injected in split mode with split ratio 5:1. Each fatty acid methyl ester was detected in selected ion monitoring mode. All results were normalized to the peak height of the C17:0 internal standard (24 (link)).