Gas-phase pyridylamination of saccharides: development and applications.
Pyridylamination is a versatile method for fluorescence labeling of oligosaccharides. The technique affords sensitive detection of saccharides with reducing termini and high-resolution separation by high-performance liquid chromatography. The conventional method, based on a liquid-phase reaction, has been extensively used in various aspects of glycobiology and glycotechnology. Unfortunately, the necessity for removing excess 2-aminopyridine makes the technique both laborious and time-consuming. Furthermore, removal of excess reagent can result in a significant loss of short saccharide components. In the present paper, we report an alternative methodology based on a "gas-phase" reaction, in which dried saccharides are reacted with vaporized 2-aminopyridine. The resultant Schiff base was also reduced in the gas phase within the same reaction microtube using a purpose-built device. The newly developed procedure was applied to both monosaccharide (GlcNAc) and oligosaccharides (isomalto-oligosaccharides) at quantitative yields with no requirement to remove excess reagent. The acid-labile sialyl linkages of alpha2-6-disialobiantennary oligosaccharides proved to be fully stable during the procedure. The developed method was also successfully applied to profiling N-linked oligosaccharides liberated from glycoproteins by hydrazinolysis and, thus, should contribute to various fields of glycomics.
Nakakita
Shin-ichi
S
Department of Functional Glycomics, Life Science Research Center, Kagawa University, 1750-1 Ikenobe, Miki-cho, Kita-gun, Kagawa 761-0793, Japan.
Sumiyoshi
Wataru
W
Miyanishi
Nobumitsu
N
Natsuka
Shunji
S
Hase
Sumihiro
S
Hirabayashi
Jun
J
D000631:Aminopyridines / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D005740:Gases / Q000737:chemistry; D009844:Oligosaccharides / Q000737:chemistry* / Q000302:isolation & purification; D012545:Schiff Bases / Q000737:chemistry; D012680:Sensitivity and Specificity; D013997:Time Factors
D000631:Aminopyridines; D005740:Gases; D009844:Oligosaccharides; D012545:Schiff Bases; C032439:alpha-aminopyridine