Journal
ANALYTICAL BIOCHEMISTRY
Volume 388, Issue 2, Pages 242-253Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ab.2009.03.004
Keywords
Milk oligosaccharide; Multibranched polylactosamine-type; oligosaccharide; MALDI-TOF MS; Exoglycosidase digestion; HPLC
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A complex mixture of diverse oligosaccharides related to the carbohydrates in glycocojugates involved in various biological events is found in animal milk/colostrum and has been challenging targets for separation and Structural studies. In the current study, we isolated oligosaccharides having high molecular masses (MW similar to 3800) from the milk samples of bearded and hooded seals and analyzed their structures by off line normal-phase-high-performance liquid chromatography-matrix-assisted laser desorption/ionization-time-of flight (NP-HPLC-MALDI-TOF) mass spectrometry (MS) by combination with sequential exoglycosidase digestion. Initially, a mixture of oligosaccharides from the seal milk Was reductively aminated with 2-aminobenzoic acid and analyzed by a combination of HPLC and MALDI-TOF MS. From MS data, these oligosaccharides contained different numbers of lactosamine units attached to the nonreducing lactose (Gal beta 1-4Glc) and fucose residue. The isolated oligosaccharides were sequentially digested with exoglycosidases and characterized by MALDI-TOF MS. The data revealed that oligosaccharides from both seal species were composed from lacto-N-neohexaose (LNnH, Ga beta 1-4GlcNAc beta 1-6[Gal beta 1-4Glc-NAc beta 1-3]Gal beta 1-4Glc) as the common core structure, and most of them contained Fuc alpha 1-2 residues at the nonreducing ends. Furthermore, the oligosaccharides from both samples contained multibranched oligosaccharides having two Gal beta 1-4GlcNAc (N-acetyllactosamine, LacNAc) residues on the Gal beta 1-4Glc-NAc beta 1-3 branch or both branches of LNnH. Elongation of the chains was observed at 3-OH positions of Gal residues, but most of the internal Gal residues were also Substituted with all N-acetyllactosamine at the 6-OH position. (c) 2009 Elsevier Inc. All rights reserved.
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