| Abstract
| - 2-Aminopyridine (PA)-derivatized oligosaccharides fromIgG were analyzed by using reversed-phase HPLC/massspectrometry (RP-HPLC/MS) and a MSn spectral library,in particular, focusing on two pairs of isomers incompletely separated or coeluted in chromatograms. Wepreviously reported that MSn spectral matching considering both major fragment ions (m/z) and intensities isuseful and applicable to the structural assignment of PA-oligosaccharide isomers. In this study, MSn spectralmatching based on the MSn spectral library was appliedto the assignment of these PA-oligosaccharide isomers inIgG. Its usefulness was investigated by comparing it tothe conventional two-dimensional mapping method basedon retention time indexes. Specifically, we focus on theassignment and quantification of the isomers, which arecoeluted in chromatograms. From this, we propose a newmethod using MSn spectral matching and the workingcurve on which are plotted the relative intensities ofselected fragment ions in their MS2 spectra versus variousmixtures of the isomers. This new method demonstratedthat the obtained quantities coincide very well with thoseestimated after separating by a combination of lectin andreversed-phase columns. This means that separation byRP-HPLC/MS is greatly simplified because completeseparation of the isomers is no longer required. Application of this new method was tested by using the two otherpairs of fucosylated and nonfucosylated PA-oligosaccharides from IgG. The results showed that this methodworks for them as well.
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