[Determination of docosahexenoic acid in human serum by capillary gas chromatography].

ABSTRACT

A reliable CGC method was developed for the determination of docosahexenoic acid (DHA) in human serum. The serum sample was acidified by adding two drops of 4.5 mol.L-1 H2SO4, and DHA was extracted from serum using ethyl acetate. The extract was evaporated to dryness under nitrogen stream. To each residue, 1 ml 1.3 mol.L-1 methanolic hydrogen chloride solution was added and the mixture was allowed to stand for 30 min in 60 degrees C water bath. After derivatization, the mixture was extracted with 1 ml of n-hexane. The solvent was evaporated under a stream of nitrogen to dryness and the residue was dissolved in 30 microliters n-hexane and subjected to capillary GC, which was equipped with a fused silica capillary column (26.3 m x 0.25 mm ID) coated with FFAP (free fatty acid phase, 0.1 micron film thickness). Tricosanoic acid was used as an internal standard. The retention time of DHA-M and internal standard was 23.41 min and 20.79 min, respectively. The minimum detection concentration of DHA in serum was 40 ng.ml-1 with a serum volume of 200 microliters and S/N value of 2. A good linear relationship between the peak area ratios and concentrations was found at the DHA concentrations ranging from 25 to 200 micrograms.ml-1. The within-day and between-day precision was 5% and 9%, respectively.