The role of water in the cryopreservation of seeds

ZARITZKY NOEMI

Water Stress in Biological, Chemical Pharmaceutical and Food Systems

SPRINGER

Lugar: New York; Año: 2015; p. 231 - 244

The preservation of plant biodiversity avoids the risk that species, and plant varieties may become extinct, producing a definitive loss of their genetic variability. Seed storage is the most effective and efficient method for the ex situ preservation of plant genetic resources and cryopreservation is proposed as the safest technique for germplasm conservation. The objective of this presentation is to discuss the role of water and the behavior of orthodox and non orthodox (recalcitrant and intermediate) seeds, when they are submitted to cryopreservation techniques.Orthodox seeds can withstand dehydration to around 0.052 gH2O g-1dry material (5% wet mass basis, wmb) and their longevity increases as seed moisture content and storage temperature decreases. Orthodox seeds can be stored at freezing temperatures without harm, as there is insufficient water for lethal ice-crystals to form. Different processes and mechanisms can be considered as responsible for the acquisition of desiccation tolerance, ensuring survival of the desiccated condition in orthodox seeds: intracellular de-differentiation; presence of antioxidant systems;  changes in metabolism, accumulation of protective molecules, including late embryogenic accumulating/abundant proteins , sucrose, and certain oligosaccharides; formation of intracellular glasses, occurrence and operation of repair mechanisms during rehydration, etc. The presence of some of the mechanisms/processes, or their absence or partial expression, is considered in the context of the varied responses to dehydration shown by nonorthodox seeds.Recalcitrant seeds are characterized by high water contents at maturity (0.43 to 4.0 gH2O g-1 dry basis, 30 to 80 % wmb) and an inability to survive drying below 0.35-0.43 gH2O g-1 dry basis (20- 30% water content wmb); they are short lived, commonly surviving for only a few days or weeks after reaching maturity and cannot be stored at freezing temperatures.Intermediate seeds have properties somewhat in between those of orthodox and recalcitrant seeds. Some intermediate seeds may be stored at sub-zero temperatures, but many are injured by freezing temperatures; seeds of tropical origin do not store well below 10°C. Seeds having intermediate or recalcitrant storage behavior are cold-sensitive; consequently, they cannot be stored in standard seed gene banks at -20 ºC. Seeds of Coffea and Citrus species have intermediate storage characteristics and cannot be stored in liquid nitrogen without a previous partial dehydration process. The basis of the optimal hydration status for cryopreservation of intermediate oily seeds is discussed in detail using different experimental data obtained in our laboratory: seed sorption isotherms, presence of freezable water (determined by DSC analysis), lipid content and seed germination percentage after immersion in liquid nitrogen (seed viability). Case-study results indicated that the optimal desiccation relative humidity for seed tolerance to liquid nitrogen exposure, ranged between 64 and 75% for Citrus sinensis and Citrus paradisi, corresponding to seed water contents of 10-15%wmb. This optimal hydration status coincides with the unfreezable water content of the seeds determined by DSC. In conclusion the controlled dehydration process before liquid nitrogen exposure constitutes a satisfactory method by which non-orthodox oily seeds withstand cryopreservation processes.