Diffusion
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Submit the exercises for diffusion by Monday, March 11 at midnight.

Introduction to Diffusion:

    Diffusion occurs when molecules move through some kind of medium. For example, when you put food coloring in water, the food coloring spreads throughout the water—even if you only put in one little droplet. We can say that the food coloring diffused through the water. Another example is when you are cooking food and the smell of the food spreads from the oven throughout the kitchen, and even throughout your home. In this case, the odor diffused through the air. In these two examples, the molecules that diffuse are the food coloring or the odorant molecules, and the medium through which they diffuse is water or air. Molecules can diffuse through liquid (ex: water), gaseous (ex: air), or even solid media (see Exercise 1).

    In both examples above, the molecules diffused away from their original source (either the food coloring droplet or the food in the oven). Why do they move away? Molecules always diffuse along their concentration gradient. That means, that they diffuse from where they are in high concentration to where they are in low concentration (Figure 1).  Once they have diffused to even out the concentration everywhere, the concentration gradient is now zero, and they don’t move in any particular direction anymore.

    As molecules move along their concentration gradient, if a membrane is in the way of their path, they may end up crossing that membrane in order to keep moving.  If the membrane is a cell membrane, that means that they will either move into or out of the cell (see right side of Figure 1).

Figure description: The dark molecule is in highest concentration at the center of all the arrows at the left. This molecule diffuses outward toward the areas of lower concentration (only diffusion toward the right is shown here). When the molecule encounters a membrane, then whether that molecule continues to diffuse along its concentration gradient depends on the permeability of the membrane.

Exercise 1: Diffusion of dye through a solid

Learning objectives: 

1. Discover that diffusion is a slow process.

2. Learn how to measure the rate of diffusion.

3. Gain an understanding for how diffusion rate determines the maximum size of cells.

    How does diffusion relate to cells? Diffusion is a relatively slow process. Since cells rely on diffusion to get oxygen (a small molecule), the slowness of diffusion forces cells to remain small. If cells were as big as baseballs, it would take many hours for oxygen to diffuse to the center of the cell and the center of the cell would die.

    Since most small molecules diffuse at roughly similar rates, we can use a measurement of diffusion to determine the maximum possible size for a cell. To do this experiment, we will observe the diffusion of a dye (potassium permanganate) through gelatin. We are using a brightly colored dye so that we can easily see and measure its rate of diffusion.  We put the dye on gelatin so that there is no movement of the diffusion medium (the solvent). If we put the dye in plain water there would be some unavoidable movement and stirring of the water. Stirring is not diffusion. Stirring tends to hide and mask the effects of diffusion.

    The main question that we are asking in this exercise is: How big can a cell get and still have its oxygen needs met by diffusion?

The Assumptions—We are making the following assumptions in this experiment:

1. Potassium permanganate diffuses at roughly the same rate as oxygen.

2. Cytoplasm can live for one minute without oxygen. Therefore, if a cell becomes too big for oxygen to diffuse to the center of the cell in one minute or less, the cell will die (as illustrated in this figure).

Procedure:

1. Obtain the test tube you made with gelatin.  Get a single crystal of potassium permanganate from the container you received (this is not always easy). Place the crystal on the gelatin. Note the time.

2. Wait one hour. Measure how far the potassium permanganate has diffused. Make your measurements in millimeters (mm). The distance diffused is the distance from the top of the gelatin to the farthest point the dye has traveled downward.

 

In one hour, potassium permanganate diffused  ___ mm.

 

 

3. Use the following equations to calculate the distance diffused in one minute in micrometers (m). You will you use your answer from equation A to carry out equation B.

Equation A:

_____ mm diffused in one hour =      ______  mm diffused per minute
60 minutes per hour

Equation B:

______ mm diffused per minute   X 1000 m per mm = 

______ m diffused per minute

Answer questions about this exercise on the submission form.

Exercise 2: Diffusion through an artificial membrane

Learning objectives:

1. Discover that molecules will diffuse through membranes.

2. Discover that the size of molecules will affect their ability to cross membranes.

3. Examine how concentration gradients determine the direction of diffusion.

How does this relate to cells? Cells are surrounded by a membrane (the plasma membrane). Small molecules (O2, CO2) enter and leave cells by diffusing through this membrane. Concentration gradients determine which way the molecules move (in or out of the cell). But keep in mind that what we are doing today is with an artificial membrane. The detailed way that biological membranes work is slightly different. For example, large molecules do enter and leave cells, but they are not able to cross the membrane without help from the cell.

The lab exercise.

    In this exercise, you will set up a concentration gradient for two molecules: starch and salt. You will put both of these molecules into an artificial cell, which you will make with an artificial membrane and clips.

Procedure:

1.  Wet one strip of dialysis tubing in water to open the strip up into a tube.   The dialysis tubing just looks like a one-inch wide piece of plastic, but when wet, you will be able to open it up with your fingers.

2. Take a piece of thread and tie a very tight knot at one end of the tubing.  Make this as tight as you can.  Really, this is important.  If it is leaky, the experiment won't work.  If you are good with knotting, you could knot this first end of the dialysis tubing by making a knot in the tubing itself (that won't work for the second end).  You will end up with something that looks sort of like one of the items in this (awful) drawing of mine.

3.  Mix your starch solution by adding water in the starch test tube up to the 15 mL mark, then shake.

4.   Add approximately 3 milliliters (mL) of starch solution to the tube.  You have some plastic pipettes that are marked for milliliters.  The exact amount is only important because you want to add the same amount of salt.

5. Make your 40% salt solution by adding just over 1/4 cup of salt to 2 cups of water.

6.   Add 3 mL of salt solution to the tube (the same amount as you had added for starch).

7.  Tie off the other end of the tube with a very tight knot of threadThis is the hardest part to do alone without spilling.  You should do this over the sink-- just in case.  If you spill it, you should still have plenty of solutions to refill it.  Your filled tubing is now an artificial cell.

8.  Rinse your artificial cell under water to remove any solutions that may have spilled onto the outside of the cell.

9.   Place your artificial cell in a container large enough for it to lay flat along the bottom... but not too large, because that might interfere with your results.

10.  Pour water over the artificial cell until it just covers the top of the artificial cell.  You want to minimize the water, so don't even let there be 5 mm of water over the tubing.

11. Your experimental setup should now look like this figure to the right.

12. Let the artificial cell sit in the water for 20 - 30 minutes. After that time, test to see if diffusion has occurred.

Testing to see if diffusion has occurred:

   You will need your artificial cell in a beaker for the next experiment!!!! You may take a small amount of water from the beaker or cell, but try to keep the setup reasonably intact!!!!

Procedure:

  1. Use your pipette to get some water from the solution outside your artificial cell.
  2. Put 2 drops of the water in each of two spots on your spot plate.  Be sure to test this water in the spot plate and not right in the container with your artificial cell.
  3. To test for the presence of starch use the iodine solution (like you used last week). If there is starch present, the iodine will turn the water blue or purple-black. If there is no starch present, iodine turns the water yellowish.
  4. To test for the presence of salt, use silver nitrate. If there is salt in the water, adding silver nitrate results in the formation of a cloudy white precipitate.   Silver nitrate will stain-- not immediately, but when exposed to light for a little bit.  So be very careful not to spill any on your skin or clothes (it won't hurt you, but it will stain you).

Answer questions about this exercise on the submission form.

Exercise 3: More diffusion through an artificial membrane

We can now use your experimental setup to do another interesting experiment. In the last experiment we used iodine to test for the presence of starch. In this experiment, we will do the opposite, and use starch to test for the presence of iodine.

Procedure:

1. Use your same artificial cell in a beaker setup from the last experiment.

2. Add 3 mL of iodine solution to the water in the beaker (basically, enough to turn your outside solution slightly yellow).

3. Now wait about 20 minutes.

Answer questions about this exercise on the submission form.

 

2006 STCC Foundation Press, content by Dawn A. Tamarkin, Ph.D.

Last changed: January 21, 2007