Identification of Sialylated Oligosaccharides Derived from Ovine and Caprine Caseinomacropeptide by Graphitized Carbon Liquid Chromatography-Electrospray Ionization Ion Trap Tandem Mass Spectrometry

Isolation and characterization of oligosaccharides from caseinomacropeptide (CMP) are important in understanding the biological and functional properties of CMP. However, it is difficult to achieve this goal, due to the high degree of isomerism present in these types of compounds. In this study, the sialylated oligosaccharides derived from ovine and caprine CMP were released as oligosaccharide alditols by reductive β-elimination and subsequently separated and characterized using graphite carbon column liquid chromatography-negative electrospray ionization ion trap tandem mass spectrometry (LC/ESI(-)-MSn). Although, the chromatographic resolution of isomeric oligosaccharides was not achieved perfectly, the characteristic tandem mass spectra of these compounds allowed differentiating and confirming unequivocally the structure of each one of the oligosaccharides. In CMP of both species, four trisaccharides and four tetrasaccharides were identified as O-glycans. Their chemical structures were identified to be Galβ1-3(NeuAcα2-6)GalNAcol, NeuAcα2-3Galβ1-3GalNAcol, Galβ1-3(NeuGcα2-6)GalNAcol, NeuGcα2-3Galβ1-3GalNAcol, NeuAcα2-3Galβ1-3(NeuGcα2-6)GalNAcol, NeuGcα2-3Galβ1-3(NeuAcα2-6)GalNAcol, NeuAcα2-3Galβ1-3(NeuAcα2-6)GalNAcol, and NeuGcα2-3Galβ1-3(NeuGcα2-6)GalNAcol. The LC/MSn methodology using an ion trap-type mass analyzer shown in this study is of general applicability for determination of short O-glycan oligosaccharides.Peer Reviewe