OPTIMIZATION AND SCALE-UP OF SOLVENT-EXTRACTION IN PURIFICATION OF HEPATITIS-A VIRUS (VAQTA(R))

Solvent extraction is a very powerful purification step in the prepara tion of VAQTA((R)), a highly purified, inactivated hepatitis A vaccine . Extraction of an aqueous product-containing protein solution with ch loroform through vigorous shaking causes irreversible denaturation of contaminant proteins at the interface. However, the hepatitis A virus (HAV) remains viable and soluble in the aqueous phase. Because three p hases (air, aqueous, and organic) are involved, and the mixing is carr ied out in individual bottles, there is very little theory available t o characterize this process, so it must be studied experimentally. Thi s extraction step was characterized by following the removal of a spec ific impurity from the aqueous phase as a representative marker for th e degree of protein precipitation. These experiments led to the identi fication and optimization of the important variables controlling the e xtraction step. They were found to be mixing time and size of vessel, with longer mixing times resulting in higher purity and larger bottle size leading to faster kinetics of impurity removal. These parameters are most likely related to solvent/aqueous interfacial area and the re sulting shear due to shaking. We conclude that, to scale up this type of mixing, the kinetics of impurity removal need to be determined expe rimentally for the systems and equipment under consideration. (C) 1997 John Wiley & Sons, Inc.