Effect of column and software on gas chromatographic determination of fatty acids

Four capillary columns (A: CP-WAX 52 CB 25 m x 0.25 mm; B: CP WAX 52 CB 30 m x 0.25 mm; C: CP-WAX 58 CB 25 m x 0.25 mm, Chrompack; D: OMEGAWAX 320 30 m x 0.32 mm, Supelco) and two integration software (Mosaic v.5.10, Chrompack and CSW v.1.7, Data Apex5) were compared for analysis of fatty acids. Column A was mounted stepwise in two different instruments. Fatty acids of blood plasma phosphatidylcholine and standard mixture of saturated fatty acids were analysed as methyl esters under identical chromatographic conditions. Both integrating software did not differ significantly in most results; differences were observed only for minor components: 16:1n9 (0.10+/-0.020 vs. 0.17 +/- 0.005 M%, P < 0.0001, column Al; 0.09 +/- 0.011 vs. 0.16 +/- 0.007 M%, P< 0.0001, column A2; 0.09 +/- 0.010 vs. 0.17 +/- 0.003 M%, P < 0.0001, column C; 0.09 +/ -0.008 vs. 0.19 +/- 0.003 M%, P < 0.0001, column D), 20:0 (0.10+0.001 vs. 0.06 +/- 0.005 M%, P < 0.05, column C) and 20:2n6 (0.43 +/- 0.030 vs. 0.91 +/- 0.016 M%, P < 0.0001, column A2). Increased values for 16:1n9 and 20:2n6 integrated by MOSAIC are caused by cointegration of two poorly resolved peaks: fatty acid and impurity from sample matrix. Lower values for 20:0 are caused by incomplete integration of minor peak. Differences between columns were observed mostly for minor fatty acids. The results indicate that CSW is more suitable software for integration of complicated chromatograms. Linear calibration dependences measured with standard mixture of saturated fatty acids (carbon number 10-24) were observed in wide range of concentrations (three orders). Slope close to unity and minimal value of intercept confirmed theoretical relations when analyses are run under optimal conditions. Use of one column is advisable in small intervention or experimental metabolic studies.