Untitled

Abstract

Lyophilization is the most popular method for achieving improved stability of labile biopharmaceuticals, but a significant fraction of product activity can be lost during processing due to stresses that occur in both the freezing and the drying stages. The effect of the freezing rate on the recovery of herpes simplex virus 2 (HSV-2) infectivity in the presence of varying concentrations of cryoprotectant excipients is reported here. The freezing conditions investigated were shelf cooling (223 K), quenching into slush nitrogen (SN2), and plunging into melting propane cooled in liquid nitrogen (LN2). The corresponding freezing rates were measured, and the ice crystal sizes formed within the samples were determined using scanning electron microscopy (SEM). The viral activity assay demonstrated the highest viral titer recovery for nitrogen cooling in the presence of low (0.25% w/v sucrose) excipient concentration. The loss of viral titer in the sample cooled by melting propane was consistently the highest among those results from the alternative cooling methods. However, this loss could be minimized by lyophilization at lower temperature and higher vacuum conditions. We suggest that this is due to a higher ratio of ice recrystallization for the sample cooled by melting propane during warming to the temperature at which freeze-drying was carried out, as smaller ice crystals readily enlarge during warming. Under the same freezing condition, a higher viral titer recovery was obtained with a formulation containing a higher concentration of sugar excipients. The reason was thought to be twofold. First, sugars stabilize membranes and proteins by hydrogen bonding to the polar residues of the biomolecules, working as a water substitute. Second, the concentrated sugar solution lowers the nucleation temperature of the water inside the virus membrane and prevents large ice crystal formation within both the virus and the external medium.

Development of Thermostable Lyophilized Inactivated Polio Vaccine

“…The first design of experiment approach was based on the most commonly used lyoprotectants, i.e., sucrose, trehalose and mannitol, in combination with dextran, with and without NaCl (Table SI , supplemental data). These excipients are known to provide physical and biochemical stabilization as well as appropriate structural properties to the cake structure, during and after lyophilization ( 29 – 32 ).…”

Effect of Chemical Stabilizers on the Thermostability and Infectivity of a Representative Panel of Freeze Dried Viruses

“…In fact, decreases in viral titre correlate with demonstrable changes since the viral envelope is assumed to be associated with viral infectivity [ 11 ]. Lyophilization of a heat-labile lipid-enveloped virus such as Herpes simplex virus pointed out that high concentration (27% w/v) of stabilizers could achieve up to 80% recovery of the viral titre [ 23 ]. In our study, similar results were observed with HSV.…”

Structural Characterization of Frozen n-Heptane Solutions of Metal-Containing Reverse Micelles

“…Recently, we have investigated the structure of reverse micelles formed under extreme conditions such as high vacuum or cryogenic temperatures. Our interest in the latter case was inspired by the increasing industrial exploitation of cryogenic techniques in biomedicine for the long-term preservation of biological materials in combination with the ability of reverse micelles to model and/or to mime some fundamental aspects of biomembranes. , Moreover, investigations of the processes occurring during fast freezing of microheterogeneous systems could also be of theoretical and technological interest. Information in this field is mostly obtained by freeze-fracture TEM and more recently by cryoTEM techniques.…”