title: Characterization of potential degradation products in a PEGylating reagent 20 kDa monomethoxy polyethylene glycol propionaldehyde by RP-HPLC, APCI-MS and NMR.
abstract: Ensuring quality of PEGylating reagents is essential for the successful development and manufacturing of PEGylated biopharmaceuticals. However, little is known about how to maintain and verify the quality of PEG raw materials for PEGylated protein manufacturing. In this study, monomethoxy polyethylene glycol propionaldehyde (mPEG-aldehyde) was subjected to conditions that mimic accelerated stability conditions. Separation of trace-level degradation products in the presence of mPEG-aldehyde was achieved by derivatization with 2,4-dinitrophenylhydrazine (DNPH), followed by reversed phase high performance liquid chromatography with ultraviolet detection (RP-HPLC-UV) at 355nm. Structural characterization by atmospheric pressure chemical ionization mass spectrometry (APCI-MS) identified formaldehyde, acetaldehyde, crotonaldehyde, acrolein, benzaldehyde, and tolualdehyde as major degradation products or process-related impurities. The presence of formaldehyde and acrolein was confirmed by (1)H NMR in the forced degraded mPEG-aldehyde samples without derivatization of mPEG-aldehyde. Findings from this study imply that reactive impurities could form as a result of inappropriate mPEG-aldehyde handling or storage. Further, a rapid screening method based on reversed phase HPLC was shown to be an effective screening assay used for routine screening of mPEG-aldehyde to ensure consistent PEGylated protein product quality.
authors: Process and Product Development, Amgen, Inc., One Amgen Center Drive, Thousand Oaks, CA 91320, USA.
authors: Heidi
authors: H
authors: Zhang
authors: Process and Product Development, Amgen, Inc., One Amgen Center Drive, Thousand Oaks, CA 91320, USA.
authors: John
authors: J
authors: Wilson
authors: Process and Product Development, Amgen, Inc., One Amgen Center Drive, Thousand Oaks, CA 91320, USA.
authors: Jifeng
authors: J
authors: Zhang
authors: Process and Product Development, Amgen, Inc., One Amgen Center Drive, Thousand Oaks, CA 91320, USA. Electronic address: yingl@amgen.com.
authors: Ying
authors: Y
authors: Luo
mesh_terms: D000171:Acrolein / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D005557:Formaldehyde / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D013058:Mass Spectrometry / Q000379:methods; D010659:Phenylhydrazines / Q000737:chemistry; D011092:Polyethylene Glycols / Q000737:chemistry*
chemical_list: D010659:Phenylhydrazines; C088349:monomethoxypoly(ethylene glycol)aldehyde; D005557:Formaldehyde; C004787:2,4-dinitrophenylhydrazine; D011092:Polyethylene Glycols; D000171:Acrolein
keywords: APCI-MS; NMR; PEGylation; RP-HPLC; mPEG-aldehyde