The paper presents the validation and verification of an analytical method for the determination of total docosahexaenoic acid (DHA) in pig serum by liquid chromatography-electrospray ionization-tandem mass spectrometry. The characteristics studied during the validation included precision and accuracy, limit of quantitation (LOQ), selectivity, calibration range and linearity, parallelism, and stability. A separate verification study was also performed. The method was linear over the range. Precision and accuracy met acceptance criteria at all levels, and the LOQ was determined as 1 μg/mL. Parallelism experiments were conducted to show that there was no bias introduced in using a surrogate matrix to quantify DHA. Recoveries of free DHA were obtained for quality control samples, and stability studies were conducted over 1, 7, 31, and 180 days. The results of the verification study were in line with the validation study, and in conclusion, the method was deemed fit for purpose for measuring total DHA in pig serum.
Keywords: DHA; LC; MS; analytical method; enrichment; serum; validation and verification.