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Chromatographic separation of asparagine-linked oligosaccharides labeled with an ultravioletabsorbing compound,p-aminobenzoic acid ethyl ester

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Abstract

We have expanded on the suitability ofp-aminobenzoic acid ethyl ester as an ultraviolet-absorbing reagent [Wanget al., (1984) Anal Biochem 141:366–81] for the analysis of asparagine-linked oligosaccharides derived from glycoproteins. The oligosaccharides released from glycoproteins by hydrazinolysis/N-reacetylation were derivatized withp-aminobenzoic acid ethyl ester and the derivatives were purified and separated into neutral and acidic oligosaccharides on a PRE-SEP C18 cartridge. The acidic oligosaccharides could be further separated into a few species by high-voltage paper electrophoresis.

p-Aminobenzoic acid ethyl ester derivatives of neutral oligosaccharides were analyzed by gel permeation chromatography on Bio-Gel P-4 and HPLC on a silica-based amide column. The elution profile and the proportion of the oligosaccharides were in agreement with literature values. The overall yield of oligosaccharides from glycoproteins was approximately 70%. Fifty pmol of oligosaccharide were detectable on Bio-Gel P-4 and 4–5 pmol on HPLC.

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Abbreviations

HPLC:

high performance liquid chromatography

NABEE:

p-aminobenzoic acid ethyl ester

FAB-MS:

fast-atom bombardment mass spectrometry

(GlcNAc)2, (GlcNAc)3, (GlcNAc)4, (GlcNAc)5 and (GlcNAc)6 :

chito-oligosaccharides containing 2,3,4,5 and 6 residues ofN-acetylglucosamine

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Authors and Affiliations

  1. Department of Biotechnology, Faculty of Engineering, Fukuyama University, 985 Sanzo, 729-02, Fukuyama, Hiroshima, Japan

    Fumito Matsuura & Akiko Imaoka

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  1. Fumito Matsuura
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  2. Akiko Imaoka
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Matsuura, F., Imaoka, A. Chromatographic separation of asparagine-linked oligosaccharides labeled with an ultravioletabsorbing compound,p-aminobenzoic acid ethyl ester. Glycoconjugate J 5, 13–26 (1988). https://doi.org/10.1007/BF01048328

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  • DOI: https://doi.org/10.1007/BF01048328

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