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Sequential GC/MS analysis of sialic acids, monosaccharides, and amino acids of glycoproteins on a single sample as heptafluorobutyrate derivatives

Pons, A; Richet, C; Robbe, C; Herrmann, Annkatrin LU ; Timmerman, P; Huet, G; Leroy, Y; Carlstedt, Ingemar LU ; Capon, C and Zanetta, JP (2003) In Biochemistry 42(27). p.8342-8353
Abstract
A GUMS procedure was developed for the analysis of all major constituents of glycoproteins. The rationale for this approach is that by using GC/MS analysis of the constituents as heptafluorobutyrate derivatives, it was possible to quantitatively determine the sialic acid, monosaccharide, fatty acids (when present), and the amino acid composition with the sample remaining in the same reaction vessel during the entire procedure. A mild acid hydrolysis was used to liberate sialic acids and was followed by formation of methyl-esters of heptafluorobutyrate (HFB) derivatives. After GC/MS analysis of sialic acids, the remaining material was submitted to acid-catalyzed methanolysis followed by the formation of HFB derivatives. After GUMS analysis... (More)
A GUMS procedure was developed for the analysis of all major constituents of glycoproteins. The rationale for this approach is that by using GC/MS analysis of the constituents as heptafluorobutyrate derivatives, it was possible to quantitatively determine the sialic acid, monosaccharide, fatty acids (when present), and the amino acid composition with the sample remaining in the same reaction vessel during the entire procedure. A mild acid hydrolysis was used to liberate sialic acids and was followed by formation of methyl-esters of heptafluorobutyrate (HFB) derivatives. After GC/MS analysis of sialic acids, the remaining material was submitted to acid-catalyzed methanolysis followed by the formation of HFB derivatives. After GUMS analysis of the monosaccharides, the sample was supplemented with norleucine (as internal standard) and hydrolyzed with 6 M HCl followed by the formation of isoamyl-esters of HFB derivatives and GC/MS analysis. His and Trp residues were modified during the step of acid-catalyzed methanolysis, but the resulting derivatives were stable during acid hydrolysis and quantitatively recovered by GUMS analysis. As a result, all constituents of glycoproteins (sialic acids, monosaccharides (or di- and trisaccharides) and amino acids) are identified in the electron impact mode of ionization and quantified using three GUMS analysis in the same chromatographic conditions and using a limited number of reagents, a considerable advantage over previous techniques. This method is very sensitive, all data (qualitative and quantitative) being obtained at the sub-nanomolar level of initial material. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Biochemistry
volume
42
issue
27
pages
8342 - 8353
publisher
The American Chemical Society
external identifiers
  • wos:000184100200025
  • pmid:12846583
  • scopus:0038503253
ISSN
0006-2960
DOI
10.1021/bi034250e
language
English
LU publication?
yes
id
f7a02a4a-dc11-4c5c-bf47-91f40dd25981 (old id 306878)
date added to LUP
2007-09-20 11:11:39
date last changed
2018-10-03 10:52:22
@article{f7a02a4a-dc11-4c5c-bf47-91f40dd25981,
  abstract     = {A GUMS procedure was developed for the analysis of all major constituents of glycoproteins. The rationale for this approach is that by using GC/MS analysis of the constituents as heptafluorobutyrate derivatives, it was possible to quantitatively determine the sialic acid, monosaccharide, fatty acids (when present), and the amino acid composition with the sample remaining in the same reaction vessel during the entire procedure. A mild acid hydrolysis was used to liberate sialic acids and was followed by formation of methyl-esters of heptafluorobutyrate (HFB) derivatives. After GC/MS analysis of sialic acids, the remaining material was submitted to acid-catalyzed methanolysis followed by the formation of HFB derivatives. After GUMS analysis of the monosaccharides, the sample was supplemented with norleucine (as internal standard) and hydrolyzed with 6 M HCl followed by the formation of isoamyl-esters of HFB derivatives and GC/MS analysis. His and Trp residues were modified during the step of acid-catalyzed methanolysis, but the resulting derivatives were stable during acid hydrolysis and quantitatively recovered by GUMS analysis. As a result, all constituents of glycoproteins (sialic acids, monosaccharides (or di- and trisaccharides) and amino acids) are identified in the electron impact mode of ionization and quantified using three GUMS analysis in the same chromatographic conditions and using a limited number of reagents, a considerable advantage over previous techniques. This method is very sensitive, all data (qualitative and quantitative) being obtained at the sub-nanomolar level of initial material.},
  author       = {Pons, A and Richet, C and Robbe, C and Herrmann, Annkatrin and Timmerman, P and Huet, G and Leroy, Y and Carlstedt, Ingemar and Capon, C and Zanetta, JP},
  issn         = {0006-2960},
  language     = {eng},
  number       = {27},
  pages        = {8342--8353},
  publisher    = {The American Chemical Society},
  series       = {Biochemistry},
  title        = {Sequential GC/MS analysis of sialic acids, monosaccharides, and amino acids of glycoproteins on a single sample as heptafluorobutyrate derivatives},
  url          = {http://dx.doi.org/10.1021/bi034250e},
  volume       = {42},
  year         = {2003},
}