Controlled water content for evaluation of denaturation temperature of freeze-dried enzymes

Abstract Water content and storage temperature play an important role in determining the thermal stability and activity retention of biomolecules, mainly in freeze-dried proteins; for this reason, the study of the effect of both properties on the thermal stabilization of proteins is paramount to the biopharmaceutical industry. A parameter used to evaluate thermal stability of proteins is the denaturation temperature ( T d ), reported extensively for a large number of proteins. Although it is known that T d values are strongly dependent on moisture, water content is not necessarily considered or deliberately controlled during denaturation measurement for solid samples. In the present work a method to evaluate T d under controlled water (ca. 6.5%) was implemented and tested for solid lysozyme to overcome the limitations encountered during the determination of T d . The implemented method can be used for quality control purposes for solid protein products since it allows an objective comparison of samples differing in provenance.