Calorimetric properties of water and triacylglycerols in fern spores relating to storage at cryogenic temperatures

Storing spores is a promising method to conserve genetic diversity of ferns ex situ. Inappropriate water contents or damaging effects of triacylglycerol (TAG) crystallization may cause initial damage and deterioration with time in spores placed at −15 °C or liquid nitrogen temperatures. We used differential scanning calorimetry (DSC) to monitor enthalpy and temperature of water and TAG phase transitions within spores of five fern species: Pteris vittata, Thelypteris palustris, Dryopteris filix-mas, Polystichum aculeatum, Polystichum setiferum. The analyses suggested that these fern spores contained between 26% and 39% TAG, and were comprised of mostly oleic (P. vittata) or linoleic acid (other species) depending on species. The water contents at which water melting events were first observable ranged from 0.06 (P. vittata) to 0.12 (P. setiferum) g H2O g−1 dry weight, and were highly correlated with water affinity parameters. In spores containing more than 0.09 (P. vittata) to 0.25 (P. setiferum) g H2O g−1 dry weight, some water partitioned into a near pure water fraction that melted at about 0 °C. These sharp peaks near 0 °C were associated with lethal freezing treatments. The enthalpy of water melting transitions was similar in fern spores, pollen and seeds; however, the unfrozen water content was much lower in fern spores compared to other forms of germplasm. Though there is a narrow range of water contents appropriate for low temperature storage of fern spores, water content can be precisely manipulated to avoid both desiccation and freezing damage.