AV: Osmosis and Diffusion

LAB 3

MOVEMENT OF MOLECULES ACROSS CELL MEMBRANES

Cells are the basic structural and functional units of every living organism. All cells consist of a ground substance, the cytoplasm, surrounded by a cell membrane. The cytoplasm is a thick, semitransparent fluid consisting of about 90% water. Suspended or dissolved in the cell water are solid particles which vary in size and composition.

Small molecules such as those of salts or sugars that dissolve in water are called solutes. The dissolving substance (water) is termed the solvent. The water and the solutes comprise a solution. Larger molecules such as proteins do not dissolve in the water but remain suspended in it because they have like charges and repel each other. These large molecules are called colloids and combined with water form a colloidal suspension.

The plasma membrane is the outer limiting membrane of the cell. Although some cells have a cell wall surrounding them, the cell membrane is the site of interaction between the cell and its environment. The plasma membrane is comprised of a bilayer of phospholipid containing protein molecules located at different levels.

The plasma membrane regulates the movement of molecules into and out of the cell. Some molecules such as water, oxygen, and carbon dioxide are able to pass across freely. Other larger molecules require carrier molecules to traverse the cell membrane or are unable to penetrate it at all. Because the plasma membrane allows some molecules to pass through freely while preventing other molecules from doing so, it is said to be selectively permeable or differentially permeable. Now let us examine some of the processes by which molecules move across the plasma membrane.

Question 1: Describe the cytoplasm of the cell.

Question 2: What are the components of a solution?

Question 3: What are colloids?

Question 4: What prevents the colloids within the cytoplasm from settling out?

Question 5: Describe the structure of the plasma membrane.

Question 6: What is the function of the plasma membrane?

Question 7: What is meant by the term "selectively permeable"?

LAB OBJECTIVE:

To gain an understanding through observation of the processes of osmosis, dialysis, diffusion, and Brownian movement.

To predict the response of plant and animal cells to hypotonic, isotonic, and hypertonic environments.

To practice using the vocabulary associated with the phenomena described in these exercises.

I. Osmosis

Water can move freely across the plasma membranes of most cells and will do so when there is a difference in concentration inside and outside. The diffusion of water across a selectively permeable membrane is called osmosis. Based upon the content of water and solute, cells may be placed in three types of solutions or environments.

Solutions which contain a higher concentration of water and a lower concentration of solute than the cytoplasm are called hypotonic solutions. Animal cells placed in a hypotonic solution will swell and in some cases rupture. The rupture of a cell due to osmosis is called plasmoptysis. Plant cells placed in a hypotonic solution will swell and become turgid but will not rupture due to their outer cell wall.

Solutions which contain a higher concentration of solute and less water than the cell cytoplasm are called hypertonic solutions. Plant and animal cells placed in hypertonic solutions lose water and shrink. The loss of water due to osmosis is called plasmolysis.

Solutions which contain the same concentration of water and solutes as the cell cytoplasm are said to be isotonic solutions. Cells placed in an isotonic solution will neither shrink nor swell since there is no net gain or loss of water.

Question 8: Define osmosis.

Question 9: What are hypotonic solutions?

Question 10: Define plasmoptysis.

Question 11: Why do plant cells not rupture in hypotonic solutions?

Question 12: What are hypertonic solutions?

Question 13: Define plasmolysis.

Question 14: What are isotonic solutions?

Procedure:

1. Using a cork borer, cut two plugs from a potato. The two plugs should be approximately the same size.

2. Using a pan balance, carefully weigh each piece of potato to the nearest tenth of a gram and write down on separate slips of paper the weight of each piece of potato. Place the pieces of potato on the slips of paper showing their weights and return to your desk.

3. Using a ruler, measure the length of each piece of potato to the nearest millimeter and record the lengths of the potato pieces on the slips of paper that show their weights.

*4. Obtain two small beakers from the front desk. With a wax pencil, label one beaker "sugar solution" and fill it three‑fourths full with a concentrated sugar solution. Place one of the potato plugs in the beaker containing the sugar solution. Record the weight and length of the plug in Table 1.

5. Label the second beaker "tap water", and fill it three‑fourths full with tap water. Place the second potato plug in the beaker of tap water. Record the weight and length of this potato plug

in Table 1.

6. Allow the potato plugs to remain in the tap water and sugar water for one hour and then finish this experiment.

7. Remove the potato plug from the sugar solution and gently blot the excess solution with a tissue. Weigh the potato plug and measure its length. Record the new weight and length in Table 1.

8. Remove the potato plug from the tap water, blot it, weigh it, and measure its length. Record the new weight and length in Table 1.

9. Determine the difference between the two weights recorded and the two lengths recorded for each potato piece and record them in Table 1.

* When finished return the sugar solution to the container provided.

Table 1

Osmosis

	First Weight	Second Weight	Difference in Weight	First Length	Second Length	Difference in Length
Plug in Sugar Solution
Plug in Tap Water

Question 15: What changes in weight and length were noted in the potato plugs?

Question 16: How does this experiment illustrate osmosis?

Question 17: Which solution was hypotonic to the potato cells?

Question 18: Which solution was hypertonic to the potato cells?

II. Dialysis

The cell cytoplasm contains a great variety of solute particles and colloids. The plasma membrane surrounding the cytoplasm is selectively permeable and allows some of the smaller solute molecules to diffuse freely across the plasma membrane. Larger solutes and colloids cannot diffuse across the cell membrane. These particles remain inside unless carrier molecules are present to transport them across the plasma membrane. Dialysis is the process by which small molecules are separated from larger ones by diffusing across a semipermeable membrane. This principle is utilized in the artificial kidney or dialysis machine which replaces the normal function of the human kidneys.

Question 19: Define dialysis.

Question 20: How may molecules too large to diffuse across the plasma membrane get across it?

Question 21: Give a practical application of dialysis.

Procedure:

1. Obtain a piece of plastic membrane about four inches long. Run tap water over it and rub it between your palms until it opens up into a tube.

2. Tie a knot in one end of the tube to form a sack open at one end. Be careful not to tear the bag.

3. Using a funnel, partially fill the bag with a solution containing one per cent starch and ten per cent glucose. Be sure to leave enough room to tie a knot at the other end of the bag.

4. Carefully tie a knot in the other end of the sack as close as possible to the solution inside. Rinse the sealed bag with tap water.

5. Blot the excess water from the surface of the bag, weigh it to the nearest tenth of a gram, and record its weight.

6. Obtain a small beaker from the front desk and fill it about three‑fourths full with distilled water. Place the bag into the beaker and allow it to remain there for thirty minutes.

7. While the bag is in the beaker of distilled water, obtain four test tubes from the front desk. These tubes will be used to test the solution placed in the bag and the distilled water for the presence of starch and glucose. Label two of the tubes "inside solution" and the other two "distilled water". Label one tube in each pair "starch" and the other "glucose".

8. With a clean dropper, add two dropperfuls of the solution placed in the bag (remove from the bottle) to each of the two tubes labeled "inside solution". With a different clean dropper, add two dropperfuls of distilled water to the other two tubes.

9. Add two drops of iodine solution to each of the tubes containing samples to be tested for starch and gently shake to mix the contents. A blue‑black color indicates that starch is present.

10. Add two dropperfuls of Benedict's reagent to each of the tubes containing samples to be tested for glucose and gently shake to mix the contents. Place the tubes in a beaker of boiling water for five minutes. A yellow, orange, or red color indicates that glucose is present. Record the results of these tests (9 and 10) in the appropriate columns of Table 2. Use a + for the presence of starch or glucose and a ‑ for their absence. Retain these four tubes to compare with results of similar tests to be performed in a few minutes.

11. Remove the sac from the beaker. Blot the excess fluid from the surface of the bag, weigh it, and record its weight.

12. Obtain four more test tubes from the front desk. You will now test the solution inside the bag and the solution in the beaker for starch and glucose. Label the four tubes accordingly.

13. With a pair of scissors snip one end of the bag and with a clean dropper add two dropperfuls to two of the test tubes.

14. Add two drops of iodine solution to one of the tubes and shake to mix the contents.

15. To the other sample add three dropperfuls of Benedict's reagent and shake to mix the contents. Place the tube in a beaker of boiling water for five minutes.

16. Record the results on the tests on the solution from the bag in Table 2.

17. Repeat the tests for starch and glucose using a sample (same volume) taken from the solution in the beaker.

18. Record the results of the tests on the solution from the beaker in Table 2.

Table 2

Dialysis

	Starch	Glucose
Solution Placed in Bag
Inside Solution After 30 Minutes
Distilled Water
Outside Solution After 30 Minutes

Question 22: How does this experiment illustrate dialysis?

Question 23: Why were the starch molecules unable to diffuse out of the bag?

Question 24: What process caused the bag to gain weight?

III. Diffusion

Because of their kinetic energy, molecules tend to scatter until a uniform distribution is attained. The movement of molecules from areas of higher concentration to areas of lower concentration is called diffusion. The difference in the concentration between the two areas is termed the concentration gradient. Diffusion, therefore, is movement with or down a concentration gradient. The condition achieved when diffusing molecules become evenly distributed in their available space is called equilibrium. The rate of diffusion is affected by such factors as molecular weight and temperature. The lower the molecular weight and the higher the temperature, the greater the rate of diffusion. Respiration, nutrition, and secretion are cellular processes in which diffusion occurs.

Question 25: Define diffusion.

Question 26: What is a concentration gradient?

Question 27: List two factors which affect the rate of diffusion.

Question 28: List three cellular processes in which diffusion occurs.

Procedure:

1. One person from each lab table should obtain a petri plate containing agar.

2. Your lab instructor will place a crystal of potassium permanganate and a crystal of methylene blue about one and one‑half inches apart on the surface of the agar plate. The molecular weight of potassium permanganate is 158 and the molecular weight of methylene blue is 320.

3. Place the plate on the lab desk and allow it to remain undisturbed for one hour.

4. Measure the diameter of the circular areas produced by both crystals using a ruler marked in millimeters.

5. Determine the rate of diffusion (millimeters per hour) for each compound by taking one‑half of the diameter (radius) of the circular areas produced by both crystals.

Question 29: How does this experiment illustrate diffusion?

Question 30: Explain why the two crystals exhibited different rates of diffusion.

IV. Brownian Movement

The molecules of all types of matter exhibit some degree of movement. The molecules of gases and liquids move much more freely than those of solids. The energy responsible for molecular movement is called kinetic energy. When visible or microscopic particles are suspended in a liquid or a gas, they are bombarded by the molecules of the suspending medium. You may have noticed the movement of dust particles in a beam of light entering a dark room. The collision of the invisible air molecules with the visible dust particles causes them to vibrate in the beam of light. This random type of motion was first reported by Robert Brown in 1827, and has since been termed Brownian movement.

Procedure SLIDE ONE:

1. Place a drop of tap water on the surface of a clean microscope slide.

2. Remove the dropper from a container of India ink and mix the ink on

the tip with the drop of water. USE VERY LITTLE INK.

3. Place a coverslip over the suspension and examine it under the microscope using the 10X and 40X objectives.

4. Look for small black carbon particles that exhibit random movement.

PROCEDURE SLIDE TWO:

1. Place a drop of tap water on a clean microscope slide.

2. Add a leaf of Elodea to the drop and place a coverslip over it.

3. Examine the cells of the leaf using the 10X and 40X objectives.

4. Observe the smallest particles within the cells for a vibrating type of movement.

Question 31: Define Brownian movement.

Question 32: What is the source of energy for Brownian movement?

Question 33: Does Brownian movement occur in both living and nonliving matter?