Understanding Poly(vinyl alcohol)-Mediated Ice Recrystallization Inhibition through Ice Adsorption Measurement and pH Effects

The development of improved cryopreservative materials is necessary to enable complete recovery of living cells and tissue after frozen storage. Remarkably, poly(vinyl alcohol) (PVA) displays some of the same cryoprotective properties as many antifreeze proteins found in cold tolerant organisms. In particular, PVA is very effective at halting the Ostwald ripening of ice, a process that mechanically damages cells and tissue. Despite the large practical importance of such a property, the mechanism by which PVA interacts with ice is poorly understood, hindering the development of improved cryoprotective materials. Herein, we quantitatively evaluated ice growth kinetics in the presence of PVA at different pH conditions and in the presence of a range of neutral salts. We demonstrated that pH, but not salt identity, alters the ability of PVA to halt ice grain coarsening. These observations are consistent with hydrogen-bonding playing a crucial role in PVA-mediated ice recrystallization inhibition. The evolution of the size distribution of ice crystals with annealing was consistent with incomplete surface coverage of ice with PVA. Binding assay measurements of dissolved fluorescently labeled PVA in an ice slurry showed that PVA interacts with ice through weak adsorption (<9%) to the ice crystal surface, which stands in contrast to fluorescently tagged type III antifreeze peptide, which binds strongly (ca. 64%) under the same conditions.