title: Dry heat forced degradation of buserelin peptide: kinetics and degradant profiling.
abstract: Buserelin is a GnRH agonist peptide drug, comprising a nine amino acid sequence (pGlu-His-Trp-Ser-Tyr-D-Ser(tBu)-Leu-Arg-Pro-NH-Et) and most commonly known for its application in hormone dependent cancer therapy, e.g. prostate cancer. In order to evaluate its hot-melt extrusion (HME) capabilities, buserelin powder in its solid state was exposed to elevated temperatures for prolonged time periods. A stability indicating UPLC-PDA method was used for quantification of buserelin and the formed degradants. Different solid state kinetic models were statistically evaluated of which the Ginstling-Brounshtein model fitted the data best. Extrapolation to and experimental verification of typical HME-related conditions, i.e. 5 min at 100°C and 125°C, showed no significant degradation, thus demonstrating the HME capabilities of buserelin. Mass spectrometric identification of the buserelin-related degradants formed under solid state heat stress was performed. Based upon the identity of these degradants, different degradation hypotheses were raised. First, direct β-elimination of the hydroxyl moiety at the serine residue, followed by fragmentation into an amide (pGlu-His-Trp-NH2) and pyruvoyl (pyruvoyl-Tyr-D-Ser(tBu)-Leu-Arg-Pro-NH-Et) peptide fragments, was postulated. Alternatively, internal esterification due to nucleophilic attack of the unprotected serine residue, followed by β-elimination or hydrolysis would yield pGlu-His-Trp, pGlu-His-Trp-NH2 and the pyruvoyl peptide fragment. Degradant pGlu-His-Trp-Ser-Tyr-NH2 is believed to be formed in a similar way. Secondly, direct backbone hydrolysis would yield pGlu-His-Trp and Tyr-D-Ser(tBu)-Leu-Arg-Pro-NH-Et peptide fragments. Moreover, the presence of Ala-Tyr-D-Ser(tBu)-Leu-Arg-Pro-NH-Et can be explained by hydrolysis of the Trp-Ser peptide bond and conversion of the serine moiety to an alanine moiety. Third and finally, isomerisation of aforementioned peptide fragments and buserelin itself was also observed.
authors.affiliation: Drug Quality and Registration (DruQuaR) Group, Faculty of Pharmaceutical Sciences, Ghent University, Harelbekestraat 72, 9000 Ghent, Belgium.
authors.forename: Matthias
authors.initials: M
authors.lastname: D'Hondt
authors.affiliation: Center for Biotechnology and Biomedicine, Institute for Bioanalytical Chemistry, Leipzig University, Deutscher Platz 5, 04103 Leipzig, Germany.
authors.forename: Maria
authors.initials: M
authors.lastname: Fedorova
authors.affiliation: Institute of Statistical Science, Academia Sinica, Academia Road 128, 11529 Taipei, Taiwan.
authors.forename: Chien-Yu
authors.initials: CY
authors.lastname: Peng
authors.affiliation: Drug Quality and Registration (DruQuaR) Group, Faculty of Pharmaceutical Sciences, Ghent University, Harelbekestraat 72, 9000 Ghent, Belgium.
authors.forename: Bert
authors.initials: B
authors.lastname: Gevaert
authors.affiliation: Drug Quality and Registration (DruQuaR) Group, Faculty of Pharmaceutical Sciences, Ghent University, Harelbekestraat 72, 9000 Ghent, Belgium.
authors.forename: Lien
authors.initials: L
authors.lastname: Taevernier
authors.affiliation: Center for Biotechnology and Biomedicine, Institute for Bioanalytical Chemistry, Leipzig University, Deutscher Platz 5, 04103 Leipzig, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Hoffmann
authors.affiliation: Drug Quality and Registration (DruQuaR) Group, Faculty of Pharmaceutical Sciences, Ghent University, Harelbekestraat 72, 9000 Ghent, Belgium. Electronic address: Bart.DeSpiegeleer@UGent.be.
authors.forename: Bart
authors.initials: B
authors.lastname: De Spiegeleer
mesh_terms: D018931:Antineoplastic Agents, Hormonal / Q000737:chemistry*; D002064:Buserelin / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D004355:Drug Stability; D006358:Hot Temperature; D006868:Hydrolysis; D007536:Isomerism; D007700:Kinetics; D008956:Models, Chemical; D015394:Molecular Structure; D011208:Powders; D055550:Protein Stability; D059748:Proteolysis; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry; D013678:Technology, Pharmaceutical / Q000379:methods
chemical_list: D018931:Antineoplastic Agents, Hormonal; D011208:Powders; D002064:Buserelin
keywords: Buserelin; Degradant identification; Dry heat stress; Kinetic models; Peptide; Solid state