[203, 209]. They are usually responsible for initial ice nucleation
events in volumes larger than ~10 μm diameter water droplets
prepared as aerosols or emulsions to study homogeneous
nucleation [212].
The phase diagram of a cryoprotectant solution can be divided
into heterogeneous and homogeneous nucleation zones (Fig. 3; for
more details, see [105]). These zones give rise to at least three
distinct types of vitrification.
Vitrification using solute concentrations insufficient to prevent
passage through the homogeneous nucleation zone will
include nucleation at an innumerably large number of points in
the solution [199] (e.g., !2500 nuclei/μm
3 in one case [213]).
Since in the homogeneous nucleation zone water is self-nucleating,
passage through this zone should be considered unstable vitrification. (In glass science, liquids in the homogenous nucleation temperature zone (T h > T > T G ) are said to be “doubly unstable”
because the zone can only be entered if temperature changes rapidly, which causes state properties of the liquid to depart from
thermodynmic equilibrium because of insufficient time for structural relaxation, and because the liquid is also liable to spontaneous crystallization by homogeneous nucleation [214]). Unstable
vitrification can be survived if warming is sufficiently rapid, as
discussed below.
Vitrification using a solute concentration high enough to
depress the T h curve to below the T G of the selected concentration
nominally avoids homogeneous nucleation, but ice formation is
normally still thermodynamically favored (T ( T m ), resulting in
what can be defined as metastable vitrification [105, 215, 216]. It is
metastable because although such a solution can supercool all the
way to the glass transition without ice nucleation necessarily occurring, the solution is not stable upon rewarming and will tend to
devitrify if the warming rate is too low. In this portion of the phase
diagram, ice nucleation events above T G will essentially be confined
to discrete locations where heterogeneous nucleators are present.
A third form of vitrification is stable vitrification, or “equilibrium vitrification” [19], which uses solute concentrations so high
that ice cannot exist in the solution [2, 105]. A practical approximation to the edge of the stable vitrification zone can be defined as
the concentration that is sufficient to make devitrification vanish
during slow warming [23, 105], but complete stability requires the
higher concentrations that are typical of those that preclude ice
growth during previous slow freezing (the “unfreezable concentration,” or C U [18, 89]).
Ice nucleation occurs by local reorientation of water molecules.
The nucleation rate is therefore driven primarily by the free energy
difference (thermodynamic driving force) between ice and liquid
water in solution. Rates of nucleation rise as temperature decreases,
reaching a maximum near the glass transition temperature
Principles of Vitrification
47
events in volumes larger than ~10 μm diameter water droplets
prepared as aerosols or emulsions to study homogeneous
nucleation [212].
The phase diagram of a cryoprotectant solution can be divided
into heterogeneous and homogeneous nucleation zones (Fig. 3; for
more details, see [105]). These zones give rise to at least three
distinct types of vitrification.
Vitrification using solute concentrations insufficient to prevent
passage through the homogeneous nucleation zone will
include nucleation at an innumerably large number of points in
the solution [199] (e.g., !2500 nuclei/μm
3 in one case [213]).
Since in the homogeneous nucleation zone water is self-nucleating,
passage through this zone should be considered unstable vitrification. (In glass science, liquids in the homogenous nucleation temperature zone (T h > T > T G ) are said to be “doubly unstable”
because the zone can only be entered if temperature changes rapidly, which causes state properties of the liquid to depart from
thermodynmic equilibrium because of insufficient time for structural relaxation, and because the liquid is also liable to spontaneous crystallization by homogeneous nucleation [214]). Unstable
vitrification can be survived if warming is sufficiently rapid, as
discussed below.
Vitrification using a solute concentration high enough to
depress the T h curve to below the T G of the selected concentration
nominally avoids homogeneous nucleation, but ice formation is
normally still thermodynamically favored (T ( T m ), resulting in
what can be defined as metastable vitrification [105, 215, 216]. It is
metastable because although such a solution can supercool all the
way to the glass transition without ice nucleation necessarily occurring, the solution is not stable upon rewarming and will tend to
devitrify if the warming rate is too low. In this portion of the phase
diagram, ice nucleation events above T G will essentially be confined
to discrete locations where heterogeneous nucleators are present.
A third form of vitrification is stable vitrification, or “equilibrium vitrification” [19], which uses solute concentrations so high
that ice cannot exist in the solution [2, 105]. A practical approximation to the edge of the stable vitrification zone can be defined as
the concentration that is sufficient to make devitrification vanish
during slow warming [23, 105], but complete stability requires the
higher concentrations that are typical of those that preclude ice
growth during previous slow freezing (the “unfreezable concentration,” or C U [18, 89]).
Ice nucleation occurs by local reorientation of water molecules.
The nucleation rate is therefore driven primarily by the free energy
difference (thermodynamic driving force) between ice and liquid
water in solution. Rates of nucleation rise as temperature decreases,
reaching a maximum near the glass transition temperature
Principles of Vitrification
47