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Anti Freezers: Protectors which binds to ice and combat cold



What are ice-binding proteins (IBPs)?

IBPs are proteins produced by cold-adapted microorganisms and commonly referred to as antifreezes. These proteins lower the freezing point of water and prevent the recrystallization, ice nucleation, and formation process by adsorbing to the ice crystals. The discovery of ice-binding proteins (IBPs) synthesized by cold-adapted microorganisms as a strategy of overcoming low temperatures resulted in an increasing interest in their specific roles. Their properties, such as lowering the freezing point of water or protection from recrystallization during storage, opened up a prospect of IBPs becoming valuable tools in commercial applications.

Where are they found, and what is their role in nature?

These unique molecules are produced by psychrophilic organisms, including bacteria, fungi, algae, certain plants, insects, and fishes, living in cold environments. The temperature on earth ranges from -81°C to 50°C, and approximately 80% of the surface is exposed continuously or partially to temperatures below 5°C. Psychrophiles are organisms that can survive in these extreme conditions. They have made certain modifications to avoid damage by crystallization by developing these IBPs, which protect their cells and help in food acquisition.

What are their types? How do they work?

They are classified into two groups, the first is antifreeze proteins (AFPs), and the second is ice-nucleating proteins (INPs). AFPs are further classified into three groups based on their thermal hysteresis (TH) and ice recrystallization inhibition (IRI) values. Their mechanism is explained further.

Figure 1. Various types of IBP


1) Antifreeze Proteins (AFPs): These acts by two mechanisms

  • Thermal Hysteresis(TH): It is the difference between melting point(MP) and freezing point(FP) by micro curvature (a small or bland curve) on the ice surface. The water molecules are thermodynamically more difficult to join to a curved ice surface than to the flat one. This property is described by the Kelvin effect. AFPs decrease FP below MP(which is the same for normal water, i.e., 0°C) in a non-colligative manner. It also increases MP insignificantly. AFPs, by binding to ice, generate a gap between FP and MP, aka TH so that ice melting and further freezing are protected. Also, change occurs in the morphology of crystals. Ice crystals take shapes ranging from circular, hexagonal, bipyramidal, to needle-shaped. Different AFPs have different structures, so changes in binding sites and thus mechanisms are seen.

          Figure 2. Mechanisms of IBP

  • IRIs: The process of creation of larger ice crystals via the Kelvin effect is called recrystallization. It occurs as temperature decreases and causes damage at a cellular level. IRIs prevent the nucleation step in which small ice crystals start coalescing into larger ones. This occurs at higher concentrations of AFPs and is an important cryoprotective mechanism.

2) INPs: These are high molecular weight proteins that act as ice nucleating nuclei and start crystallization of ice from 0°C to -2°C, in between the intercellular spaces and thus avoid the freezing of internal organelles. INPs act as nuclei of ice crystals by exhibiting a surface structure that is similar to the surface of ice crystals.

There are various hypotheses about the binding of IBP to ice surfaces. The first one is based on research by Deveries and Raymond (1977), who assumed that AFP binds irreversibly following hydrogen bonds between -OH groups of side chains of threonine residues in IBP. Other aspects like the extent of hydrogen bonding, IBS planarity, structural match with ice lattice, methyl group inclusion was taken into consideration by Clatherate and assumed that water molecules form “cages” around the hydrophobic region in IBS and then freeze as the next layer of ice on its surface, resulting in binding AFP and ice.


  1. In food technology: Used for shelf-life prolongation, ice creams(allowed by USFDA), and other bakery and dairy products like yogurt, to improve frozen meat and dough quality.
  2. In agriculture: For protecting temperature-sensitive plants and animals, avoid frosting in cold climates and applied as a biofertilizer to increase plant growth at cold temperatures.
  3. For cryopreservation: For hypothermic storage, cryosurgery, transplantation, and transportation of biologicals.
  4. In material technology: For deicing of roads and aircraft, for freezing instruments.
  5. In the fuel industry: As an anti-freezing agent.
  6. Weather control: Used for seeding clouds for rain.
  7. Production of artificial snow:  Applied as active bacterial cells or protein powder solutions for the production of artificial snow(Snowmax, Genecor, Rochester USA). One example is Snowmax which is produced from inactivated microorganisms and is harmless to the health of humans, animals, and plants. The producer says that Snomax is made from a bacterium (Pseudomonas syringae)

Reference (Jun-21-A7)

About the Author: Shrikant Dnyanoba Sonwane is First Year M. Pharm  Pharmaceutics student of the Institute of Chemical Technology, Mumbai. He aims to study thoroughly about novel drug delivery systems. Currently, he is working on ‘Formulation and Development of Parenteral Microspheres’. He is very passionate about his research work and puts all the efforts to achieve his goal.

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