科研之友
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摘要
The succinimide intermediate generated during deamidation of asparagine-containing peptides and proteins has been implicated as having a role in the formation of multiple types of degradants in addition to hydrolysis products, including racemization products and, more recently, amide-linked, nonreducible protein and peptide aggregates. The formation of alternative degradants may be particularly important in solid-state formulations. This study quantitatively examines the role of the succinimide intermediate in hydrolysis, racemization, and covalent, amide-linked adduct formation in amorphous lyophiles. The degradation of a model peptide, GlyPhel-AsnGly, and its l- or d-succinimide intermediates were examined in lyophiles containing hydroxypropyl methylcellulose and varying amounts of excess GlyVal. Disappearance of the starting reactants and formation of up to 10 degradants were monitored when lyophiles were exposed to either 27 degrees C/40% relative humidity (RH) or 40 degrees C/75 RH using a stability indicating high-performance liquid chromatography method. Terminal degradant profiles were the same when the starting reactant was either GlyPhel-AsnGly or its succinimide intermediate. Nucleophilic attack occurred preferentially at the a-carbonyl of the succinimide intermediate at ratios of approximately 2:1 for both water and the N-terminus of GlyVal as the attacking nucleophiles. A mechanism-based kinetic model analysis indicates that hydrolysis, racemization, and covalent, amide-linked adduct formation all proceed via the succinimide intermediate.
- 出版日期2012-9
