Rotational Rheology of Bovine Serum Albumin Solutions: Confounding Effects of Impurities, Mechanistic Considerations and Potential Implications on Protein Formulation Development
To show and rationalize the confounding effects on the rotational/oscillatory rheology of surface active impurities in commercial protein formulations such as bovine serum albumin, BSA.
Bulk and interfacial rotational/oscillatory rheology were used to study the viscosity, complex viscosity, storage/elastic modulus, G’ and loss/viscous modulus, G”, as a function of time of aqueous formulations of BSA and their purified components.
Viscosity/time profiles at steady shear for different commercial BSA products and lots showed viscosity increase, decrease and time-independent profiles at low shear rates. All lots showed shear thinning. BSA monomer and dimers/aggregates, in general, showed similar profiles. Addition of low levels of surfactant or high shear rates rendered all solutions to be Newtonian-like. Interfacial viscosity studies paralleled those on the rotational rheometer. G’ > G” with viscosity increase and G’ < G” with viscosity decrease over time.
We provide a rational explanation for the time-dependent and source-dependent rheological behavior of aqueous formulations of commercially available BSA proteins based on the migration of protein and surface active impurities to the air/water interface within the rheometer plates leading to the formation and breakdown of protein networks. Highly purified proteins is warranted in rheological studies of protein drug product candidates.
KEY WORDSbovine serum albumin high molecular weight species/aggregates mechanism protein stability rotational/interfacial rheology surface active impurity
Bovine serum albumin
High molecular weight species
- Polysorbate 20
Reversed phase high-performance Liquid chromatography
Size exclusion high-performance Liquid chromatography
Small angle Neutron scattering
ACKNOWLEDGMENTS AND DISCLOSURES
The authors wish to thank Alice Beekman for her pioneering work on the rheology of protein and polymer formulations and who brought to our attention the possibility of proteins at surfaces contributing to the overall rheology she observed; in addition we thank her for her contributions to the manuscript. The authors are grateful to Dave Brems and Margaret Ricci for support of this research; to Da Ren for technical support in LC/MS analysis; to Lyanne Wong for her early work on protein rheology. The authors also thank Philip Rolfe and David Bohnsack for their contributions to this work.
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