Biomedical subjects

Satsuki Itoh

Publications and source records attributed to Satsuki Itoh.

22 records · Page 2Linked to original sources

PubMed2002

Simultaneous microanalysis of N-linked oligosaccharides in a glycoprotein using microbore graphitized carbon column liquid chromatography-mass spectrometry.

We previously reported that graphitized carbon column liquid chromatography-mass spectrometry (GCC-LC-MS) is very useful for the structural analysis of carbohydrates in a glycoprotein. In this study, GCC-LC-MS was adapted for the simultaneous microanalysis of oligosaccharides. A variety of oligosaccharide alditols prepared from fetuin, ribonuclease B, and recombinant human erythropoietin were used as model oligosaccharides. The use of microbore GCC-LC-MS was found to be successful for rapid, sensitive, and simultaneous analysis of high-mannose-type, desialylated fucosyl complex-type, sialylated complex-type, and sialylated fucosyl complex-type oligosaccharide alditols. Furthermore, we demonstrate that this method is applicable to the analysis of carbohydrate heterogeneity in a glycoprotein that possesses diverse oligosaccharides. Microbore GCC-LC-MS was able to characterize high-mannose-type, hybrid-type, and complex-type oligosaccharides in tissue plasminogen activator produced from human melanoma cells in a single analysis.

Carbohydrate Sequence↗

PubMed2002

Usefulness of sugar mapping by liquid chromatography/mass spectrometry in comparability assessments of glycoprotein products.

We have previously reported that sugar-mapping by liquid chromatography/mass spectrometry (LC/MS) equipped with a graphitized carbon column (GCC) can be useful for structural analysis of carbohydrates in a glycoprotein. In this paper, we evaluated sugar-mapping with regard to its use in comparability assessment of glycoprotein products. Erythropoietins (EPO) produced from three different sources were chosen as models of the closely related glycoprotein products. The two-dimensional displays of sugar maps drawn by LC/MS with GCC clearly showed the differences in carbohydrate heterogeneity with regard to sialylation, acetylation, and sulphation patterns among three EPOs. Exoglycosidase digestion followed by sugar-mapping provided information regarding the structure of characteristic carbohydrates in each EPO. These results demonstrate that LC/MS with GCC can reveal the details of carbohydrate heterogeneity in order to distinguish between closely related glycoprotein products. Our method can thus be useful in comparability assessments of therapeutic glycoproteins.

Animals↗

PubMed2002

Usefulness of glycopeptide mapping by liquid chromatography/mass spectrometry in comparability assessment of glycoprotein products.

We previously reported on glycopeptide mapping of erythropoietin (EPO) by liquid chromatography/mass spectrometry (LC/MS). Using this method, glycopeptides in proteolytic digestion can be eluted before peptides, and are further separated on the basis of the carbohydrate structure. The detailed glycosylation at each glycosylation site can be elucidated based on mass chromatography and mass spectroscopy. In this study, we evaluated glycopeptide mapping with regard to its use in comparability assessment of glycoprotein products possessing multiple glycosylation sites. Models of closely related glycoprotein products used in this study are EPOs produced from three different sources. We previously reported that there are differences in the carbohydrate heterogeneity of these EPOs with regard to sialylation, acetylation, and sulphation patterns, using sugar mapping by LC/MS. In this paper, we demonstrated that glycopeptide mapping can distinguish site-specific glycosylation among these three EPOs and reveal the differences in acetylation, sialylation, and sulphation at each glycosylation site in one analysis. Our method can thus be useful in comparability assessment of therapeutic glycoproteins in terms of glycosylation.

Animals↗

PubMed2002

[Study on evaluating methods for the quality control of glycoprotein products. (IV)--erythropoietin products. Part 4].

We have previously reported on peptide mapping of recombinant human erythropoietin (rhEPO) by liquid chromatography/mass spectrometry (LC/MS). Using this method, both peptides and glycosylation at each glycosylation site can be elucidated based on the mass chromatogram and mass spectrum. In this study, we evaluated the mass spectrometric peptide mapping with regard to its use in comparability assessment of both protein parts and carbohydrates parts in glycoprotein products. Models of closely related glycoprotein products used in this study are rhEPOs produced from three different sources. We demonstrated that the mass spectrometric peptide mapping can elucidate the identity of protein part, and differences in site-specific carbohydrates heterogeneity due to acetylation and sulfation among the three rhEPOs. Our method can thus be useful in comparability assessment of therapeutic glycoproteins.

Binding Sites↗