Impact of sucrose level on storage stability of proteins in freeze-dried solids: II. Correlation of aggregation rate with protein structure and molecular mobility*

The purpose of this study is to investigate the impact of sucrose level on storage stability of dried proteins and thus better understand the mechanism of protein stabilization by disaccharides in lyophilized protein products. Five proteins were freeze dried with different amounts of sucrose, and protein aggregation was quantified using Size Exclusion Chromatography. Protein secondary structure was monitored by FTIR. The global mobility was studied using Thermal Activity Monitor (TAM), and fast local dynamics with a timescale of nanoseconds was characterized by neutron backscattering. The density of the protein formulations was measured with a gas pycnometer. The physical stability of the proteins increased monotonically with an increasing content of sucrose over the entire range of compositions studied. Both FTIR structure and structural relaxation time from TAM achieved maxima at about 1:1 mass ratio for most proteins studied. Therefore, protein stabilization by sugar cannot be completely explained by global dynamics and FTIR structure throughout the whole range of compositions. On the other hand, both the fast local mobility and free volume obtained from density decreased monotonically with an increased level of sucrose in the formulations, and thus the local dynamics and free volume correlate well with protein storage stability. © 2008 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:3145–3166, 2009