title: Hydrogen Peroxide Induced Protein Oxidation During Storage and Lyophilization Process.
abstract: Although the impact of hydrogen peroxide (HP) on proteins in liquid solutions has been studied extensively, the impact during lyophilization has been largely overlooked. The purpose of this work was to investigate the effect of HP on lyophilized proteins and HP removal by lyophilization. A protein formulation at 5 mg/mL and its placebo were spiked with HP up to 5.0 ppm and then lyophilized. HP concentration, protein oxidation, and aggregation were monitored before and after lyophilization, as well as during storage at 25°C. The lyophilization process removed on average 94.1% of HP from protein formulation, but only 72.5% from the placebo. There were also significant increases in protein oxidization and aggregation. The oxidation increment correlated with the decrease of HP concentration in both the protein formulation and placebo at all temperatures. Protein oxidation at different freezing temperatures was also studied in follow-up studies. Data from these studies suggest that (1) HP has a significant impact on oxidation and aggregation of protein during lyophilization; (2) significant oxidation can occur even when the protein formulation is frozen; (3) the oxidized protein is more prone to aggregation during lyophilization process.
authors.affiliation: Biopharmaceutics Development, Sanofi Genzyme, Framingham, Massachusetts 01701. Electronic address: wcheng@shire.com.
authors.forename: Weiqiang
authors.initials: W
authors.lastname: Cheng
authors.affiliation: Biopharmaceutics Development, Sanofi Genzyme, Framingham, Massachusetts 01701.
authors.forename: Xiaoyang
authors.initials: X
authors.lastname: Zheng
authors.affiliation: Biopharmaceutics Development, Sanofi Genzyme, Framingham, Massachusetts 01701. Electronic address: mark.yang@genzyme.com.
authors.forename: Mark
authors.initials: M
authors.lastname: Yang
mesh_terms: D002626:Chemistry, Pharmaceutical / Q000379:methods* / Q000592:standards; D004356:Drug Storage / Q000379:methods / Q000592:standards; D005612:Freeze Drying / Q000379:methods / Q000592:standards; D006861:Hydrogen Peroxide / Q000737:chemistry / Q000378:metabolism*; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism / Q000592:standards; D019999:Pharmaceutical Solutions / Q000737:chemistry / Q000378:metabolism / Q000592:standards; D010865:Pilot Projects; D011506:Proteins / Q000737:chemistry / Q000378:metabolism* / Q000592:standards
chemical_list: D010446:Peptide Fragments; D019999:Pharmaceutical Solutions; D011506:Proteins; D006861:Hydrogen Peroxide
keywords: aggregation; hydrogen peroxide; lyophilization; oxidation; protein