A chromatographic and mass spectrometric strategy for the analysis of oligosaccharides: determination of the glycan structures in porcine thyroglobulin.
Oligosaccharides released from porcine thyroglobulin were first derivatised with 2-aminoacridone (2-AMAC) and analysed by capillary electrophoresis to determine the complexity of this glycan pool. The same glycan mixture was then subjected to either a sialidase digest or a sialidase and fucosidase digest prior to derivatisation with 2-AMAC and analysis by normal phase high pressure liquid chromatography (HPLC). Comparison of the three chromatographic profiles with known standards allowed an initial identification of the glycan structures. The 2-AMAC derivatised glycans were then collected from HPLC for matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) analysis and the molecular weights of predicted structures were confirmed. This study demonstrates that a two enzyme array and subsequent MALDI-TOF analysis can be used successfully to assign the major glycans present in a complex mixture.
Charlwood
J
J
SmithKline Beecham Pharmaceuticals, Harlow, Essex, UK.
Birrell
H
H
Organ
A
A
Camilleri
P
P
D000609:Aminoacridines; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D019075:Electrophoresis, Capillary; D005456:Fluorescent Dyes; D006868:Hydrolysis; D009439:Neuraminidase; D009844:Oligosaccharides / Q000032:analysis*; D011134:Polysaccharides / Q000032:analysis*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013552:Swine; D013954:Thyroglobulin / Q000032:analysis*; D005644:alpha-L-Fucosidase
D000609:Aminoacridines; D005456:Fluorescent Dyes; D009844:Oligosaccharides; D011134:Polysaccharides; C072515:2-aminoacridone; D013954:Thyroglobulin; D009439:Neuraminidase; D005644:alpha-L-Fucosidase