Control of protein particle formation during ultrafiltration/diafiltration through interfacial protection.
This study investigates the mechanism of protein particle formation during ultrafiltration/diafiltration (UF/DF), finding that agitation drives particle formation by promoting protein-interface adsorption and desorption. Low conductivity and the presence of surfactant reduced the level of particle formation in small-scale stirring studies, and the same trends were observed in pumping and UF/DF. Polysorbate 80 (PS80) and hydroxypropyl-β-cyclodextrin (HPβCD) reduced particle formation in UF/DF by factors of 15 and 4, respectively. Measurements of conformational stability, colloidal stability, and surface tension demonstrated that PS80 protects against particle formation by preventing protein-interface adsorption, low conductivity improves the colloidal stability of the protein, and the mechanism of action of HPβCD remains unclear. This work demonstrates that interfacial adsorption-desorption of the protein during UF/DF is the principal cause of particle formation, that the level of surfactant-free particle formation depends on the colloidal stability of the protein, and that the inclusion of surfactant greatly reduces in-process particle formation during UF/DF.
Department of Purification Process Sciences, MedImmune, Gaithersburg, Maryland, 20878.
Daniel J
DJ
Callahan
Bradford
B
Stanley
Yuling
Y
Li
D000073738:2-Hydroxypropyl-beta-cyclodextrin; D000327:Adsorption; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000378:metabolism; D002919:Chymotrypsinogen / Q000737:chemistry* / Q000378:metabolism; D003102:Colloids; D003956:Dialysis / Q000009:adverse effects*; D004355:Drug Stability; D004795:Enzyme Stability; D005079:Excipients / Q000737:chemistry; D007074:Immunoglobulin G / Q000737:chemistry* / Q000378:metabolism; D009113:Muramidase / Q000737:chemistry* / Q000378:metabolism; D010047:Ovalbumin / Q000737:chemistry* / Q000378:metabolism; D010316:Particle Size; D011136:Polysorbates / Q000737:chemistry; D055550:Protein Stability; D012995:Solubility; D013500:Surface Tension; D013501:Surface-Active Agents / Q000737:chemistry; D014462:Ultrafiltration / Q000009:adverse effects*; D047392:beta-Cyclodextrins / Q000737:chemistry
D000911:Antibodies, Monoclonal; D003102:Colloids; D005079:Excipients; D007074:Immunoglobulin G; D011136:Polysorbates; D013501:Surface-Active Agents; D047392:beta-Cyclodextrins; D000073738:2-Hydroxypropyl-beta-cyclodextrin; D010047:Ovalbumin; D002919:Chymotrypsinogen; D009113:Muramidase
cyclodextrins; monoclonal antibody; processing; protein aggregation; stability; surfactants