Synergistic cryoprotective effect of deaeration and polysorbate 80 on IgG denaturation during thin-film freeze-drying

Abstract

Polysorbate 80 (PS80) is employed as a cryoprotectant in biotherapeutic formulations to preserve biologics from denaturation during the freezing process. However, excessive quantities of PS80 can lead to stickiness in dry powder, thereby reducing production yields and altering the physical and aerodynamic characteristics of the powder. Additionally, degradation byproducts of PS80 pose a risk of destabilizing proteins in therapeutic formulations. Our study reveals the synergistic cryoprotective effect of combining deaeration with PS80. PS80 plays a protective role by competitively binding to ice/water and air/water interfaces, as well as binding directly to IgG molecules. Deaeration increases the cryoprotective effect of PS80 by reducing the surface area of air microbubbles, thereby reducing the required amount of PS80 for surface coverage and limiting available adsorption sites for IgG molecules, which allows the decrease in the necessary concentration of PS80 required to minimize IgG denaturation during the thin-film freeze-drying (TFFD) process. Even at a low concentration of 0.0025 % w/v, PS80 prevents the aggregation of IgG during the freezing step of TFFD but does not eliminate IgG denaturation during the drying step. PS80 concentrations ranging from 0.0025 % to 0.0400 % w/v exhibit no impact on the specific surface area (SSA) and aerodynamic properties (i.e., fine particle fraction (FPF), mass median aerodynamic diameter (MMAD)) of IgG dry powders. These IgG TFFD powders exhibit a fine particle fraction exceeding 70 % and a median mass aerodynamic diameter of approximately 2.3–2.5 μm, with an SSA of around 20 m2/g.