A Novel Method of Natural Cryoprotection

Correlating measurements from differential scanning calorimetry, freeze-fracture freeze-etch electron microscopy, and survival of twigs after two-step cooling experiments, we provide strong evidence that winter-hardened Populus balsamifera v. virginiana (Sarg.) resists the stresses of freezing below -28 degrees C by amorphous solidification (glass formation) of most of its intracellular contents during slow cooling (</=5 degrees C per hour). It is shown that other components of the intracellular medium go through glass transitions during slow cooling at about -45 degrees C and below -70 degrees C. This ;three glass' model was then used to predict the results of differential scanning calorimetry, freeze-fracture freeze-etch electron microscopy, and biological experiments. This model is the first definitive explanation for the resistance of a woody plant to liquid N(2) temperatures even if quench cooling (1200 degrees C per minute) begins at temperatures as high as -20 degrees C and warming is very slow (</=5 degrees C per hour). It is also the first time high temperature natural intracellular glass formation has been demonstrated.