title: Formation of protein sub-visible particles during vacuum degassing of etanercept solutions.
abstract: The main purpose of this manuscript is to describe a phenomenon in which vacuum degassing a reconstituted freeze-dried fusion protein etanercept formulation caused a significant amount of protein sub-visible particles (SbVP). Physical stability of etanercept was monitored by micro-flow imaging (MFI), dynamic light scattering (DLS), size-exclusion high pressure liquid chromatography (SE-HPLC) and far- and near-ultraviolet circular dichroism (far- and near-UV CD). One potential explanation of this phenomenon is that bubble collapses when the vacuum is applied, leads to substantial heat formation, and ultimately free radical formation. Subsequently, the effect of a free-radical scavenger (ascorbic acid, AA) on SbVP formation was also evaluated. Degassing of etanercept solution by applying vacuum caused substantial increase of SbVP, as detected by MFI and DLS. However, traditional techniques such as SE-HPLC could not detect any change. The addition of free-radical scavenger had minimal effect on SbVP formation, therefore the formation of free radicals was probably not the main cause for this effect.
authors: College of Pharmaceutical Sciences, Zhejiang University, 866 Yuhangtang Road, Xihu District, Hangzhou 310058, China; Division of Biopharmaceuticals, Zhejiang Hisun Pharmaceutical Inc, 46 Waisha Road, Jiaojiang District, 317000 Taizhou, China.
authors: Haibin
authors: H
authors: Wang
authors: Division of Biopharmaceuticals, Zhejiang Hisun Pharmaceutical Inc, 46 Waisha Road, Jiaojiang District, 317000 Taizhou, China.
authors: Hong-Jian
authors: HJ
authors: Zheng
authors: Division of Biopharmaceuticals, Zhejiang Hisun Pharmaceutical Inc, 46 Waisha Road, Jiaojiang District, 317000 Taizhou, China.
authors: Zhao
authors: Z
authors: Wang
authors: Division of Biopharmaceuticals, Zhejiang Hisun Pharmaceutical Inc, 46 Waisha Road, Jiaojiang District, 317000 Taizhou, China.
authors: Hua
authors: H
authors: Bai
authors: Department of Pharmaceutical Sciences, University of Colorado Denver, Aurora, CO 80045, United States.
authors: John F
authors: JF
authors: Carpenter
authors: College of Pharmaceutical Sciences, Zhejiang University, 866 Yuhangtang Road, Xihu District, Hangzhou 310058, China. Electronic address: chenshuqing@zju.edu.cn.
authors: Shuqing
authors: S
authors: Chen
authors: Division of Biopharmaceuticals, Zhejiang Hisun Pharmaceutical Inc, 46 Waisha Road, Jiaojiang District, 317000 Taizhou, China. Electronic address: wjfang@hisunpharm.com.
authors: Wei-Jie
authors: WJ
authors: Fang
mesh_terms: D002626:Chemistry, Pharmaceutical / Q000379:methods; D004355:Drug Stability; D000068800:Etanercept; D016166:Free Radical Scavengers / Q000737:chemistry; D005612:Freeze Drying / Q000379:methods; D007074:Immunoglobulin G / Q000737:chemistry*; D010316:Particle Size; D011208:Powders / Q000737:chemistry; D011506:Proteins / Q000737:chemistry*; D018124:Receptors, Tumor Necrosis Factor / Q000737:chemistry*; D012996:Solutions / Q000737:chemistry*; D014618:Vacuum
chemical_list: D016166:Free Radical Scavengers; D007074:Immunoglobulin G; D011208:Powders; D011506:Proteins; D018124:Receptors, Tumor Necrosis Factor; D012996:Solutions; D000068800:Etanercept
keywords: Degassing; Freeze-drying; Micro-flow imaging; Protein particle