Db ffap fused silica capillary column

Lipids were extracted from ileum via the Folch Method [32 (link)] and phospholipid fractions were separated by thin layer chromatography (TLC) as previously described [29 (link)]. Briefly, samples were spotted on H-plates (EMD Chemicals, Gibbstown, NJ, USA) to separate phospholipid species. Bands corresponding to lyso-phosphatidylcholine (lyso-PC), sphingomyelin (SM), phosphatidylcholine (PC), phosphatidylserine (PS), phosphatidylinositol (PI), and phosphatidylethanolamine (PE) were collected and methylated with 14% boron trifluoride-methanol (Fisher Scientific, Mississauga, ON, Canada). Fatty acid methyl esters were separated on a DB-FFAP fused-silica capillary column (15 m, 0.1 m film thickness, 0.1 mm i.d.; Agilent, Mississauga, ON, Canada) and quantified on an Agilent 6890 gas chromatograph. Peaks were identified by retention times of fatty acid methyl ester standards (Nu-Chek-Prep, Elysian, MN, USA) using EZchrom Elite version 3.2.1 software. Fatty acid results were calculated as percent composition or ug/0.1 g tissue with 10 µg of heptadecanoic acid (17:0) added to each fraction for the internal standard.

All solvents and reagents were obtained from Fisher Scientific (Toronto, ON, Canada). Prefrontal cortex (PFC), hippocampal (HPC) and hypothalamic (HYP) samples (0.1 g) were maintained on ice and homogenized in a 0.1 M KCl solution. Total fatty acid methyl esters were extracted from samples with a chloroform:methanol (2:1, v/v) solution, according to Folch et al. [23 (link)]. Complete details of the fatty acid analysis have been previously published [24 (link)]. Fatty acid methyl esters were detected using an Agilent 6890A gas chromatograph with flame ionization detector (Agilent Technologies, Palo Alto, CA, USA) and separated on DBFFAP fused silica capillary column (15 m, 0.1 µm film thickness, 0.1 mm i.d.; Agilent Technologies, USA). Fatty acid peaks were identified by comparison to retention times of known fatty acid standard peaks (Nu-Chek-Prep, Elysian, MN, USA) using EZChrom Elite software (Version 3.3.2) to determine the relative abundance of individual fatty acids. Fatty acids are reported as a percentage of total fatty acids detected. The n-3 PUFA:LC-PUFA ratio was calculated as the sum of EPA+DPAn-3+DHA/sum of all PUFA ≥ 20 carbons.