Determination of fatty acids by high-performance liquid chromatography and fluorescence detection using precolumn derivatization with 9-fluorenylmethyl chloroformate

Abstract

ABSTRACTA new high-performance liquid chromatographic method is described for the determination of fatty acids in seed oils. The method was based on precolumn derivatization with 9-fluorenylmethyl chloroformate as a labeling agent and fluorescence detection. Fatty acids were extracted from the samples and subjected to derivatization with the reagent at 60°C for 10 min. The chromatographic separation of 14 fatty acids (C10–C22) was achieved on a combined loading compression octadecyl sulfate (CLC-ODS) column with a run time of 30 min. Three-step gradient elution of a mobile phase consisted of acetonitrile and water was used, and the signal was monitored at excitation and emission wavelengths of 265 and 315 nm, respectively. The method indicated favorable sensitivity and reproducibility for fatty acids’ derivatives. The detection limits, at a signal-to-noise ratio of 3, were 0.01–0.05 µg/ml and relative standard deviations (RSDs) were less than 0.27%. Excellent linear responses were observed with coefficients of 0.9995. This method was applied to quantify fatty acids in white, brown, and black sesame seeds’ oil.