Improved method of determination of biologically important C10:0-C22:6 fatty acids as their 2-nitrophenylhydrazides by reversed-phase high-performance liquid chromatography.

Fatty acids were separated by reversed-phase high-performance liquid chromatography after derivatization with 2-nitrophenylhydrazine hydrochloride in the presence of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride. The separation of a mixture of fourteen kinds of biologically important fatty acid hydrazides (C10:0-C22:6) was achieved within 15 min. Using margaric acid (C17:0) as internal standard, each fatty acid could be quantitated over the range of 2.5-5000 pmol per injection. Analytical recoveries ranged from 98.1 to 102.6%. The intra- and inter-assay coefficients of variation were less than 2.5 and 3.2%, respectively. For the determination of esterified fatty acids in fats and oils, the saponified mixture was directly derivatized without extraction. This method was compared gave similar fatty acid profiles to those obtained with the conventional liquid-liquid extraction method. It is simple, rapid and accurate for routine analyses of esterified fatty acids in biological materials.