Pressure-activated disassembly of cryoprotectant supramolecules in isochoric freezing

The efficacy of cryopreservation is fundamentally limited by the difficulty of achieving high intracellular concentrations of cryoprotective agents (CPAs) without inducing osmotic injury or chemical toxicity during loading. This Short Communication advances a thermodynamic hypothesis proposing that intracellular cryoprotection could be achieved through the in situ generation of cryoprotective solutes via pressure-activated disassembly of supramolecular complexes composed of cryoprotectant monomers or oligomers. The physical trigger for this disassembly is the hydrostatic pressure that arises intrinsically during isochoric (constant-volume) freezing: as ice forms, the fixed-volume constraint produces a substantial pressure increase. We propose that the stability of these assemblies is governed by the Helmholtz free energy, and that isochoric pressure shifts the free-energy landscape to favor the dissociated state for assemblies with a negative reaction molar volume. Because the pressures generated during isochoric freezing reach levels known to destabilize supramolecular complexes, this mechanism offers a plausible route for generating CPAs precisely during the freezing process. This approach would decouple cryoprotectant availability from membrane transport and synchronize protection with the onset of freezing. The purpose of this contribution is to articulate the thermodynamic basis and physical plausibility of this mechanism and to motivate future investigation of pressure-mediated preservation strategies.