Abstract
Dry preservation of biologics in sugar glasses is regarded as a promising alternative to conventional cryopreservation. Evidence from various studies has suggested that there is a critical range of water content beyond which the viability of preserved biologics can be greatly compromised. In this study the viability of T-cells was determined as a function of end water content after microwave-assisted drying in trehalose solutions. Hydrogen-bonding and clustering phenomena in trehalose solutions of the same moisture content were also evaluated using molecular dynamics simulation. Post-rehydration viability decreased dramatically within the range of 0.1–1 gH2O/gdw. Molecular modeling revealed that as the water content approached 0.1 gH2O/gdw the matrix formed a large interconnected trehalose skeleton with a minimal number of bound water molecules scattered in the bulk. The diffusion coefficients of trehalose oxygen atoms most distant from the glycosidic linkage fluctuated around 7.5 × 10−14 m2/s within the range of 0.02–0.1 gH2O/gdw and increased again to ~1.13 × 10−13 m2/s at 0.01 gH2O/gdw and below due to the loss of water in the free volume between trehalose molecules. These insights can guide the optimal selection of final moisture contents to advance dry preservation methods.
Highlights
Despite the utility of sugar glasses for biopreservation purposes, there seems to be a critical range of water content beyond which the viability of certain preserved biological samples is greatly compromised
What is the critical water content at which all water molecules are hydrogen-bonded to solutes or other macromolecules? What is the basis for the non-monotonic relationship between molecular mobility and water content? Molecular dynamics studies of aqueous trehalose systems to date have not concentrated on the low water content composition range
To provide further and direct evidence for the existence of the critical water content range when biological samples are subject to dehydration conditions, we carried out viability studies on Jurkat Clone E6-1 cells that had been dried in trehalose solutions to various end water contents by employing a microwave-assisted drying approach
Summary
Despite the utility of sugar glasses for biopreservation purposes, there seems to be a critical range of water content beyond which the viability of certain preserved biological samples is greatly compromised. It has been observed that the gel to liquid crystalline phase transition temperature (Tm) of phospholipids does not change significantly until the water content falls below ~0.25 gH2O/gdw[13] This critical level, the Tm can increase sharply as a result of a denser packing of the lipid headgroups because the water molecules are depleted from the vicinity of the headgroups and the lipid membranes can be destabilized[13]. Drying methods such as microwave-assisted[16,17], spin[1], or air[11] drying, are able to deplete macromolecules and membranes of their hydration shell water[13,18] This allows an extremely low water content to be achieved and, as a result, the glass transition temperature of the sugar-water matrix can be elevated above the temperature of drying. To provide further and direct evidence for the existence of the critical water content range when biological samples are subject to dehydration conditions, we carried out viability studies on Jurkat Clone E6-1 cells that had been dried in trehalose solutions to various end water contents by employing a microwave-assisted drying approach
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