decrease plasmolysis, but generally damage the plasmalemma and possibly
intracellular organelles. The latter may upon thawing cause the release of
enzymes into solution, whereby they become active. Mechanical damage to
cell walls, i.e., in vegetable tissues, is also possible; this may result in a
product of poor, e.g., sloppy, texture upon thawing. Fast freezing, which
leads to the formation of more and smaller ice crystals, often tends to reduce
freezing damage. During frozen storage, temperature fluctuations can cause
partial thawing and refreezing, generally resulting in an increase of crystal
size and increased freezing damage.
There is a vast literature on preventing or minimizingfreezing injuryto
living organisms. Most organisms can protect themselves by a number of
metabolic adaptations to freezing conditions, a subject outside the scope of
this book. Killing of vegetative bacteria by freezing can occur to some
extent, but it is never complete. Bacterial spores are very resistant to
freezing.
Cryoprotectants. These are substances that reduce undesirable
effects caused by freezing. Addition of nonionic solutes reduces ionic
strengthof freeze-concentrated solutions containing ionic substances. Since
a high ionic strength can be very damaging to proteins, addition of solutes
may be quite a useful measure. At a given temperature, the magnitude of the
water activity (as well as that of the osmotic pressure) is fixed in a partly
frozen system (Figure 16.12). The higher the initial molar solute
concentration, the smaller the proportion of water freezing at a given
temperature, hence the lower the concentration of the ionic species. This is
illustrated in Figure 16.14, and it is seen that the salt concentration can be
considerably reduced at a given temperature. The lower the molar mass of
the cryoprotectant, the greater its efficiency for a given mass fraction.
FIGURE16.13 Development of an extracellular ice crystal (black) and the
resulting plasmolysis occurring in a piece of tissue upon slow cooling. Highly
schematic. (Adapted from H. T. Meriman. Federation Proc. 22 (1963) 81.)