The temperature outside my window is 11 deg. F, well below the freezing temperature of water. This is certainly evidenced by the fact that all the water I see is frozen – either ice or snow. I must also assume that the water I can’t see is frozen as well. This should include the water inside the cells of hibernating insects. Since insects have a high concentration of water inside their cells, and since insects are cold-blooded – their temperature matching that of their surroundings - one would expect that the water inside their cells would freeze at these low temperatures. However, if that should happen, it would be lethal to the insect. To see why, try these two simple experiments”
- On a cold winters day, go outside and breath on a window of your house. The moisture in your breath, on hitting the cold glass, will form ice crystals. These crystals, as you should be able to see, have very sharp edges. These sharp edges can wreak havoc on the delicate internal structures of the cell, possibly even rupturing the cell itself.
- Place an unopened soda can in the freezer and leave it to freeze. When the water in the can turns to ice, it expands, rupturing the can. When water in a cell freezes, it too expands and ruptures the cell.
So how is it that hibernating insects are not affected by freezing temperatures? They have two basic strategies to prevent this freezing damage; freeze tolerance and freeze avoidance.
In freeze tolerance, the water in the insect body does freeze, but before the freezing occurs, the water in the cells migrates to the spaces between the cells. Therefore, when the water freezes, the ice crystals that form do so in between the cells – not inside them where their sharp edges would damage the cells.
In freeze avoidance, the insects have a few methods to prevent the water in the cells from freezing at these low temperatures. Three of these methods are listed in this month’s nature story, “Nature’s Ornaments.”. One of them will be discussed here.
During the winter, many insects build up high concentrations of chemicals called cryoprotectants in the fluids in their bodies, both in the cells and the spaces between the cells. Think of a cryoprotectant as anti-freeze, like the anti-freeze that goes in the car radiator to prevent the radiator water from freezing. In fact, some insects actually use ethylene glycol – the same chemical that cars use for anti-freeze - as their cryoprotectant. However, the most common cryoprotectant used by insects is glycerin.
The higher the concentration of glycerin in water, the lower the temperature before the water will freeze. In some cases, the water inside the insect will remain liquid down to -40 deg. F!
Here is an experiment to demonstrate this:
In this experiment, I set up an ice cube tray with water. Each individual cube of the tray had a different concentration of glycerin mixed in with the water. The tray was placed in the freezer and left overnight. The next day, I removed the tray and recorded at what concentration the water remained liquid at the temperature my freezer was set at.
- Glycerin (also called glycerol, bought at the pharmacy)
- Graduated cylinder – or something to measure milliliters of liquid
(I used a cup used for measuring dosages of medicine)
- Ice cube tray
- Thermometer (make sure it will measure temperatures below freezing)
For this experiment, my water samples were
100% water/0% glycerin
80% water/ 20% glycerin
70% water/ 30% glycerin
60% water/40% glycerin
50% water/50% glycerin
40% water/60% glycerin
Making a percent solution:
Chemists often make up solutions of chemicals in water as a percent solution. This is very easy to do, and if you always make up the solutions in your experiments this way, you will be sure that you are using the same technique throughout all your experiments.
In this case, the solute is the glycerin and the total solution is the water and glycerin together.
Therefore, for 25 ml of a 20% solution of glycerin:
- As a control, add 100% water to the first cube in the tray.
- In the second cube, add 25 ml of a 20% glycerin solution.
According to the math above, 5 ml of glycerin are needed to make 25 ml of total solution. So add 5 ml of glycerin to your container and bring the total volume to 25 ml by adding 20 ml of water.
- In the third cube, add 25 ml of a 30% glycerin solution.
- In the fourth cube, add 25 ml of a 40% glycerin solution.
- In the fifth cube, add 25 ml of a 50% glycerin solution.
- In the sixth cube, add 25 ml of a 60% glycerin solution.
- In the seventh cube, add 25 ml of a 70% glycerin solution.
- Place the ice cube tray in the freezer.
Note; mark your experiment cube tray so no one mistakenly uses your glycerin ice cubes for a drink!
- Place a thermometer inside your refrigerator to record it’s temperature.
- Check your ice cube tray the next day to see at what concentration the water does not freeze.
The purpose of this experiment is only to show that water, which normally freezes at 32 deg. F can be made to freeze at lower temperatures. It does not indicate the level of glycerin actually in an insect’s body. Remember, the water inside an insect’s cell is not pure water. There are many other chemicals in the cell which will affect the freezing point of the water without the glycerol added. Therefore, the amount of glycerin necessary in the actual insect cell may differ from your results.
- Other Explorations:
Glucose is another common cryoprotectant used in insects. Repeat the above experiment using glucose instead of glycerin.
*glucose can be bought at health food stores as Dextrose
*glucose comes as a solid.
The formula for making a percent solution from a solid is:
Therefore, for 25 ml of a 20% solution of glucose:
So, 5 grams of glucose are needed for a total volume of 25ml.
Add 5 grams of glucose to your container and add about 20 ml of water and mix. Once mixed, add more water until the total volume is 25 ml.
- Ocean water has a salinity of about 3.5%. Because of the salt, salt water freezes at about 28.4F. What does the concentration of salt need to be before it freezes at the temperature of your refrigerator?