Analysis of recombinant human follicle-stimulating hormone (FSH) by mass spectrometric approaches

Journal Article (2011)
Author(s)

J. Grass (BOKU-University of Natural Resources and Life Sciences)

Martin Pabst (BOKU-University of Natural Resources and Life Sciences)

Martina Chang (Polymun Scientific Immunbiologische Forschung GmbH)

Manfred Wozny (MassMap GmbH and Co. KG)

Friedrich Altmann (BOKU-University of Natural Resources and Life Sciences)

Affiliation
External organisation
DOI related publication
https://doi.org/10.1007/s00216-011-4923-5
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Publication Year
2011
Language
English
Affiliation
External organisation
Issue number
8
Volume number
400
Pages (from-to)
2427-2438

Abstract

Recombinant human follicle stimulating hormone is an important drug in reproductive medicine. Thorough analysis of the heterodimeric heavily glycosylated protein is a prerequisite for the evaluation of production batches as well as for the determination of "essential similarity" of new biosimilars. The concerted application of different liquid chromatography-mass spectrometry methods enabled the complete depiction of the primary structure of this pituitary hormone. Sequence coverage of 100% for the α- as well as the β-chain was achieved with tryptic peptides. Most of these peptides could be verified by tandem mass spectrometry. Site-specific analysis of all four glycosylation sites was, however, not possible with tryptic but with chymotryptic peptides. Quantification of the glycoforms of each glycopeptide was accomplished with the software MassMap®. Both protein subunits gave interpretable mass spectra upon S-alkylation and separation on a C5 reversed-phase column. Glycan isomer patterns were depicted by separation on porous graphitic carbon, using mass spectrometric detection for the evaluation of the glycopeptide liquid chromatography-electrospray ionization data. The currently marketed product Gonal-f™ and a potential biosimilar were compared with the help of these procedures.

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