Determination by electrospray mass spectrometry and 1H-NMR spectroscopy of primary structures of variously fucosylated neutral oligosaccharides based on the iso-lacto-N-octaose core

Abstract

We have isolated a nonfucosylated and three variously fucosylated neutral oligosaccharides from human milk that are based on the iso-lacto-N-octaose core. Their structures were characterized by the combined use of electrospray mass spectrometry (ES-MS) and NMR spectroscopy. The branching pattern and blood group-related Lewis determinants, together with partial sequences and linkages of these oligosaccharides, were initially elucidated by high-sensitivity ES-MS/MS analysis, and then their full structure assignment was completed by methylation analysis and 1H-NMR. Three new structures were identified. The nonfucosylated iso-lacto-N-octaose, Galbeta1-3GlcNAcbeta1-3Galbeta1-4GlcNAcbeta1-6[Galbeta1-3GlcNAcbeta1-3]Galbeta1-4Glc, has not previously been reported as an individual oligosaccharide. The monofucosylated and trifucosylated iso-lacto-N-octaose, Galbeta1-3GlcNAcbeta1-3Galbeta1-4(Fucalpha1-3) GlcNAcbeta1-6[Galbeta1-3GlcNAcbeta1-3]Galbeta1-4Glc and Galbeta1-3(Fucalpha1-4)GlcNAcbeta1-3Galbeta1-4(Fucalpha1-3)GlcNAcbeta1-6[Galbeta1-3(Fucalpha1-4)GlcNAcbeta1-3]Galbeta1-4Glc, both containing an internal Lex epitope, are also novel structures.