Determination of N-acetylneuraminic acid in poultry eggs by ultra performance liquid chromatography–tandem mass spectrometry

Abstract

N-acetylneuraminic acid is an important member of sialic acids which is a family of nine-carbon carboxylated sugars most frequently found in terminal position in glycoconjugates. It was widely distributed in different portions of organisms. In this study, a method of ultra performance liquid chromatography–tandem mass spectrometry was developed for the determination of N-acetylneuraminic acid in poultry eggs. The N-acetylneuraminic acid was freed by heating the materials in 0.1 M H₂SO₄ at 80°C for 2 h. Then, chloroform reagent was used to eliminate phospholipids, cholesterol and other organic impurities, meanwhile, it was able to precipitate protein. In the cleanup step, C18 solid phase extraction columns were used to purify the matrix. Determination of N-acetylneuraminic acid was performed with electrospray ionization in negative ion mode. Chromatographic separation was performed on a Waters Acquity UPLC BEH C18 (100 × 2.1 mm, 1.7 μm) analytical column. The gradient elution reagent was acetonitrile and water (containing 0.1% formic acid). The tandem spectrometer was operated in the Multiple Reaction Monitoring mode. The linearity over of sialic acids on-column had a correlation coefficient greater than 0.999. The detection method of N-acetylneuraminic acid in the poultry eggs had good precision and acceptable recovery.