Cryoprotectant

A cryoprotectant is a substance used to protect biological tissue from freezing damage (i.e. that due to ice formation). Arctic and Antarctic insects, fish and amphibians create cryoprotectants (antifreeze compounds and antifreeze proteins) in their bodies to minimize freezing damage during cold winter periods. Cryoprotectants are also used to preserve living materials in the study of biology and to preserve food products.

Mechanism

Cryoprotectants operate by increasing the solute concentration in cells. However, in order to be biologically viable they must easily penetrate cells and must not be toxic to cells.

Glass transition temperature

Some cryoprotectants function by lowering the glass transition temperature of a solution or of a material. In this way, the cryoprotectant prevents actual freezing, and the solution maintains some flexibility in a glassy phase. Many cryoprotectants also function by forming hydrogen bonds with biological molecules as water molecules are displaced. Hydrogen bonding in aqueous solutions is important for proper protein and DNA function. Thus, as the cryoprotectant replaces the water molecules, the biological material retains its native physiological structure and function, although they are no longer immersed in an aqueous environment. This preservation strategy is most often utilized in anhydrobiosis.

Toxicity

Mixtures of cryoprotectants have less toxicity and are more effective than single-agent cryoprotectants. A mixture of formamide with DMSO (dimethyl sulfoxide), propylene glycol, and a colloid was for many years, the most effective of all artificially created cryoprotectants. Cryoprotectant mixtures have been used for vitrification (i.e. solidification without crystal ice formation). Vitrification has important applications in preserving embryos, biological tissues, and organs for transplant. Vitrification is also used in cryonics in an effort to eliminate freezing damage.

Conventional cryoprotectants

Conventional cryoprotectants are glycols (alcohols containing at least two hydroxyl groups), such as ethylene glycol , propylene glycol, and glycerol. Ethylene glycol is commonly used as automobile antifreeze, and propylene glycol has been used to reduce ice formation in ice cream. Dimethyl sulfoxide (DMSO) is also regarded as a conventional cryoprotectant. Glycerol and DMSO have been used for decades by cryobiologists to reduce ice formation in sperm,^[1] oocytes,^[2] and embryos that are cold-preserved in liquid nitrogen.

Examples in nature

Insects

Insects most often use sugars or polyols as cryoprotectants. One species that uses cryoprotectant is Polistes exclamans. In this species, the different levels of cryoprotectant can be used to distinguish between morphologies.^[3]

Amphibians

Arctic frogs use glucose,^[4] but Arctic salamanders create glycerol in their livers for use as a cryoprotectant.

Food preservation

Cryoprotectants are also used to preserve foods. These compounds are typically sugars that are inexpensive and do not pose any toxicity concerns. For example, many (raw) frozen chicken products contain a sucrose and sodium phosphates solution in water..

Common cryoprotectants

DMSO Ethylene glycol Glycerol

2-Methyl-2,4-pentanediol (MPD) Propylene glycol Sucrose

See also

• Antifreeze protein
• Lyophilization
• List of emerging technologies
• Cryopreservation
• Cryostasis (clathrate hydrates)

References

1. Imrat, P.; Suthanmapinanth, P.; Saikhun, K.; Mahasawangkul, S.; Sostaric, E.; Sombutputorn, P.; Jansittiwate, S.; Thongtip, N.; et al. (February 2013). "Effect of pre-freeze semen quality, extender and cryoprotectant on the post-thaw quality of Asian elephant (Elephas maximus indicus) semen". Cryobiology 66 (1): 52–59. doi:10.1016/j.cryobiol.2012.11.003.
2. Karlsson, Jens O.M.; Szurek, Edyta A.; Higgins, Adam Z.; Lee, Sang R.; Eroglu, Ali (February 2014). "Optimization of cryoprotectant loading into murine and human oocytes". Cryobiology 68 (1): 18–28. doi:10.1016/j.cryobiol.2013.11.002.
3. J.E. Strassmann; R.E. Lee Jr.; R.R. Rojas & J.G Baust (1984). "Caste and sex differencesin cold-hardiness in the social wasps, Polistes annularis and P. exclamans". Insectes Sociaux 31 (3): 291–301. doi:10.1007/BF02223613.
4. Larson, D. J.; Middle, L.; Vu, H.; Zhang, W.; Serianni, A. S.; Duman, J.; Barnes, B. M. (15 April 2014). "Wood frog adaptations to overwintering in Alaska: New limits to freezing tolerance". Journal of Experimental Biology. doi:10.1242/jeb.101931.