[Study on evaluating methods for the quality control of glycoprotein products. (III)--Erythropoietin products. Part...

We reported previously that peptide mapping using high-performance liquid chromatography with electrospray ionization mass spectrometry (LC/MS) and liquid chromatography with tandem mass spectrometry (LC/MS/MS) are useful for determination of the glycosylation sites, carbohydrate structure, and site-specific carbohydrate heterogeneity of glycoproteins. Here, with intention to enhance the sensitivity and shorten the time-span of analysis to characterize glycoproteins, especially biotechnological products with carbohydrate moieties, we studied the introduction of HPLC with a microbore column to LC/MS with recombinant erythropoietin (rh-EPO). In addition, we evaluated the ability of LC/MS/MS precursor-ion scanning to make identification of glycopeptides and facilitate the analysis of carbohydrate moieties. We found that the peptide mapping with microbore HPLC is highly sensitive and more rapid than the previous method, and the precursor-ion scanning is helpful for identifying glycopeptides. Our results indicate that these methods are very useful for characterization and quality control of the carbohydrate moieties of biotechnological products.