Solid-State Rehydration-Induced Recovery of Bilirubin Oxidase Activity in Lyophilized Formulations Reduced during Freeze-Drying.

Abstract

Dehydration during freeze-drying often alters the native conformation of proteins, leading to a loss of activity. Solid-state rehydration of lyophilized protein formulations was studied to examine the recovery from structural damage caused by freeze-drying. Bilirubin oxidase was used as a model protein. The enzymatic activity of bilirubin oxidase decreased markedly when freeze-dried with polyvinylalcohol (PVA), but this loss in activity was partially recovered by subsequent solid-state rehydration of the lyophilized formulations. This recovery of activity upon solid-state rehydration was not observed in lyophilized formulations containing dextran that exhibited a similar loss of activity during freeze-drying. Thus, PVA appears to have a greater ability to act as a water substitute than dextran, resulting in less structural damage during dehydration. This in turn allows a recovery in activity upon solid-state rehydration.