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

Abstract

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.

Production, structure–function relationships, mechanisms, and applications of antifreeze peptides

“…The coverslip is placed in heptane, which had been cooled to –80 °C with dry ice, and then transferred to a –6 °C glass box cooled by circulation. After being kept at a constant temperature for 30 to 60 min, the micrographs were recorded (Burkey, Riley, Wang, Hatridge, & Lynd, 2018; Li et al., 2017). This method was transferred at low temperatures, so the operation is understandably difficult.…”

Designing the next generation of cryoprotectants – From proteins to small molecules

“…As a result, other researchers have synthesized various polymers and polyampholytes which have been assessed for IRI properties. The TH and IRI activity of various polymers (Figure ) containing poly‐ l ‐histidine 12 , poly‐ l ‐hydroxyproline 13 , poly‐ l ‐lysine 14 , poly‐ l ‐asparagine 15 , polyacrylic acid 16 , polyvinylpyrrolidone 17 , polyethylene glycol 18 , and polyvinyl alcohol 19 have been reported . Similar to the AFGPs, it was observed that larger polymers exhibited increased IRI activity .…”

A Review of the Material Characteristics, Antifreeze Mechanisms, and Applications of Cryoprotectants (CPAs)

“…Contrary to the strong, irreversible adsorp-tion of AFP, the interaction between PVA and ice is weak and reversible. The PVA in solution that adsorbed on ice may be less than 9% [28]. When the concentration of PVA is only 5 × 10 −7 mol/L, it can still effectively inhibit ice recrystallization [29].…”