Room-Temperature Aerosol Dehydration of Green Fluorescent Protein.

Rapid Room-Temperature Aerosol Dehydration (RTAD) is a novel, scalable drying technology for powderization and thermal stabilization of pharmaceutical drug products. Compared to conventional spray drying processes, typically using droplets of 10-200 micron in diameter generated by high-shear spraying, RTAD uses much smaller droplets with diameter 0.1 to 20 microns produced in modified liquid atomization processes. These fine droplets evaporate rapidly within 10-100 ms at room temperature conditions, thereby reducing drying-induced stresses for thermally sensitive biologics. In this study, we employed Green Fluorescent Protein (GFP) as a model biological molecule to optimize the RTAD system design and process parameters. We experimentally investigated the effects of droplet size, multiphase flow patterns in the drying chamber, and usage of polysorbate 20 as a model surfactant on GFP fluorescence after drying and powder reconstitution. The experiments demonstrated that the presence of surfactant in the formulation significantly influenced GFP fluorescence intensity, especially for smaller droplets. The numerical studies using Computational Fluid Dynamics simulations revealed that the intensity of GFP fluorescence in the produced dry powders was dependent on the patterns of multiphase flow in the drying chamber. Non-axisymmetric flows and closed circulating streamlines near the drying gas inlet resulted in considerably longer particle residence times and subjected GFP molecules to excess stress that negatively impacted the GFP fluorescence intensity. Through iterative optimization of the chamber design, process parameters and feedstock formulation, we achieved recovery of the GFP fluorescence intensity that exceeded 96% in the obtained dry powders. This work establishes GFP as a sensitive model biologic and its fluorescence intensity as a powerful tool to rapidly assess process efficiency and the ability to preserve bioactivity after dehydration. The study has broad implications for the design and scale-up of drying technologies, which can potentially transform the production of dry powder biopharmaceuticals.