Abstract: The peroxyoxalate chemiluminescence (CL) detection of fatty acids in human serum combined with high-performance liquid chromatography (HPLC) is described. Some fatty acids in serum were extracted with a 1:1 (v/v) mixture of chloroform-n-heptane. 2-(4-Hydrazinocarbonyl- phenyl)-4,5-diphenylimidazole (HCPI) was used as a fluorescent labelling reagent of the fatty acids. The labelling reaction was carried out at 30 ºC for 1 h at pH 6.5 and the resulting reaction mixture was subjected to HPLC. The labelled fatty acid C17 (P-C17) was used as the internal standard. The labelled fatty acids C16 and C18 were separated within 18 min on an ODS-80TM column (150 mm×6 mm ID, 5 μm, Tosoh Japan). The calibration curves of fatty acids from the spiked control serum were Y1 = -0.0037 + 0.0028X1, r = 0.994 for FA C16 and Y2 = 0.0012 + 0.00098X2, r = 0.999 for FA C18, respectively. The average recoveries of facids from the spiked control serum were 107.2% (n = 8, RSD = 4.3%)for FA C16 and 97.35%(n=8, RSD=4.0%) for FA C18, respectively. The lower detection limits of fatty acids after reaction were 12μmol per 20 μl injection for FA C16 and 18 μmol per 20 μl injection for FA C18, respectively (signal to noise ratio, S/N = 2). The HPLC/CL method was applied to the determination of FA C16 and FA C18 in normal human serum and the results showed that the concentrations of fatty acids in normal human serum were 0.134±0.009 μ mol/ml serum (n = 5) for FA C16 and 0.052±0.028 μmol/ml serum (n = 5) for FA C18, respectively.

Key words: Fatty acids, High-performance Liquid Chromatography, Peroxyoxalate chemiluminescence detection, Human serum