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

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 H-1-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 Le x epitope, are also novel structures.