Cryoprotection of yeast by alcohols during rapid freezing

We have investigated the cryoprotective effect of alcohols on Saccharomyces cerevisiae and other yeast under various freezing conditions. For S. cerevisiae, at a cooling rate of 3 degrees C min-1 methanol and ethanol acted as cryosensitizers. However, at a cooling rate of 200 degrees C min-1, both methanol and ethanol proved superior to all other cryoprotectants tested, including glycerol, dimethyl sulfoxide, lactose, trehalose, polyethylene glycol, and polyvinyl pyrrolidone. Propan-2-ol also demonstrated a small but measurable effect although propan-1-ol and butan-1-ol demonstrated no cryoprotective effect. A minimum cooling rate of 25 degrees C min-1 to elicit the cryoprotective effect of ethanol was necessary; below this rate it acted as a cryosensitizer. At cooling rates up to 650 degrees C min-1 substantial cryoprotective effect was still evident. Although the effect of ethanol was variable for other yeast genera tested, ethanol acted positively for all strains of S. cerevisiae. We hypothesize that the cryoprotective effect of alcohols during rapid cooling is a result of their ability to induce increased membrane permeability, allowing rapid water equilibration during extracellular freezing and avoidance of intracellular ice crystal formation.