Desiccation sensitivity and cryopreservation of the embryonic axes of the seeds of two Acer species

Abstract

The most dangerous stage of cryopreservation of embryonic axes of maple seeds is tissue dehydration, as excessive desiccation leads to membrane lipid peroxidation and irreversible loss of semi-permeability. We attempted cryopreservation of the embryonic axes (EAs) from orthodox seeds of Acer platanoides (Norway maple) and recalcitrant seeds of A. pseudoplatanus (sycamore), two closely related native tree species. Two pretreatment steps were tested before cryostorage: air drying and cooling to −40 °C. The aim of our study was to determine the possible types of damages to the isolated embryonic axes and their viability after exposure to liquid nitrogen (LN2, −196 °C). Ice nucleation in tissues was assessed by differential thermal analysis (DTA). In embryonic axes of A. platanoides, ice crystallization did not occur when the water content (WC) was below 26.8 %, while in A. pseudoplatanus, the safe level of WC that prevented ice nucleation was below 35.0 %. During desiccation alone, the plasma membrane integrity and viability of A. platanoides EAs were only slightly affected, while in A. pseudoplatanus, an increase in membrane breakdown and a marked decrease in the viability of EAs were observed. However, when WC exceeded 30 %, we observed significant damage to the tissues of both Acer species after the EAs were cooled to −40 °C or after cryostorage in LN2. EAs of orthodox A. platanoides tolerated dehydration to 10 % WC, while recalcitrant A. pseudoplatanus to 25 % WC only. A very highly significant correlation was found for those species between the ice nucleation and WC, as determined from DTA analysis. Exposure of EAs to LN2 was successful (i.e., ≥50 % axes formed seedlings) for A. platanoides tissues desiccated to 10–15 % WC and for A. pseudoplatanus tissues desiccated to 15–20 % WC.