Osmosis and Diffusion: How biological membranes are important This page is a lab preparation guide for instructors.

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1 Osmosis and Diffusion: How biological membranes are important This page is a lab preparation guide for instructors. **All solutions and dialysis bags can easily be prepared prior to lab start to maximize time in the lab. ** Experiment 1- Materials per group of 4 students: (4) 13-cm lengths of dialysis tubing, to be soaked in water. (Ensure you tie a knot into on end prior to soaking the materials), dental floss/string (to tie tops of dialysis bags after being filled), 1 ml plastic pipette, china marker, (4) 250 ml beakers, scissors Materials for the class: Water source/dh2o, balance/scale, paper towels, 15% and 30% sucrose solutions Experiment 2- Materials per group of 4 students: (2) 13-cm lengths of dialysis tubing, to be soaked in water. (Ensure you tie a knot into on end prior to soaking the materials), dental floss/string (to tie tops of dialysis bags after being filled), (2) 1 ml plastic pipettes, china marker, (2) 400 ml beakers, scissors, spot plate or plastic petri dish for visual test. Materials for the class: Water source/ dh2o, balance/scale, paper towels, 10% soluble starch solution, diluted iodine solution (I2KI) Experiment 3- Materials per group of 4 students: (2) Clean class microscope slides and coverslips, compound light microscope Materials for the class: Elodea plant in tap water, (2) bottles of distilled water/dh2o, 20% NaCl (sodium chloride)

2 Osmosis and Diffusion: How biological membranes are important For these labs you will be working with many chemicals and equipment. Ensure you are careful while handling the chemicals and equipment. Clean all glassware properly. Write all information, results, and end of lab questions in your lab notebook. All labs should be written in the appropriate format listed in the course syllabus. Consult the lab aids or professor if you require assistance. Living cells are constructed primarily of water that is retained by a selectively permeable membrane, known as the phospholipid bilayer. This membrane will only allow substances such as water, oxygen, carbon dioxide and small nonpolar ions to diffuse across the membrane freely. These small molecules are known as the solute. The water itself that the molecules dissolve in are the solvent. Together they will account for the total solution. A solution can be described as being hypertonic, hypotonic or isotonic to the cell (plant or animal) that is placed into that solution. A hypertonic solution is one that has many solutes in comparison to its solvent concentration. A hypotonic solution has low amounts of solutes and carries more water. An isotonic solution has an equal distribution of both solute and solvent. The cell that is placed into each of these solutions will undergo a change based on the presence or absence of the concentration gradient. Diffusion is the movement of molecules from a region of higher concentration to a region of lower concentration. This is known as the concentration gradient. It is created when an unequal distribution of molecules vs. osmotic pressure attempt to create an equilibrium. Osmosis is the movement of water from a region of higher concentration to a region of lower concentration. Don t get osmosis and diffusion confused. Osmosis in a high concentration simply means there are little solutes and water will move from that higher location to an area with more solutes to create an equilibrium. The same concept applies in diffusion when solutes move from a higher concentration into an area of lower concentration. These molecules and water gradients will always attempt to create a perfect equilibrium. This lab includes the following experiments; 1. Effect of Solute Concentration on Osmosis 2. Effect of Solute Size on Diffusion and Selective Permeability of Membranes 3. Effects of Concentration in Plant Cells Experiment 1: Effect of Solute Concentration on Osmosis (20 min. setup/60 min. complete) Introduction: The purpose of this experiment is to understand solute concentration and the effects it has on the rate of osmosis. Utilizing a dialysis bag that represents the phospholipid, semipermeable membrane, you will be able to witness osmosis over a period of 60 minutes by weighing the bag and observing physical evidence of osmosis taking place. Procedure: 1. Make a table in your lab notebook like the one listed below and write your hypothesis about what will occur with each bag: Page 1 of 4

3 Bag Bag Contents Beaker Contents 1a dh 2O dh 2O 1b 15% sucrose dh 2O 1c 30% sucrose dh 2O 1d dh 2O 30% sucrose 0 min. 15 min. 30 min. 45 min. 60 min. Total Weight Change (g) 2. Soak four sections of dialysis tubing provided in water for a few seconds. This will allow them to open up at the open end in order for you to fill it with the listed contents above. 3. Label four 250 ml beakers with a china marker to correspond with the bag contents from your table listed above. (1a, 1b, 1c, and 1d). 4. Fill each beaker with the corresponding contents above to the 100 ml indicator. 5. Using a small pipet, take each content and fill the bags one at a time with 10 ml as listed above. (After filling each bag ensure you twist the bag and tie it with a strip of dental floss. This will prevent the contents of each bag from leaking out of the top. Also ensure you don t have any excess air in the bag, this will affect your results). 6. After filling the bags, rinse the outside with water and blot dry with a paper towel completely. 7. Now weigh each one and record the data for the 0 min. section of your table. 8. Place each bag into their corresponding labeled beakers. 9. At each given time interval remove the bags and dry them in the same manner as before and weigh them. Record the information and place them back into their containers. 10. After your final weighing clean all equipment and discard of dialysis tubing in the trash. 11. Answer the following questions in your lab notebook: a) Was your original hypothesis for each solute in its given concentration solvent correct? Why or why not? b) Describe what you observed in each bag. c) Was the direction of net movement of water in bags 2b-2d going into or out of each bag? d) What bag gained the most weight? Why did this happen? e) If a 30% solution is separated from a 50% sugar solution by a selectively permeable membrane, what direction will there be movement of water? Explain. f) Based on your observations in this exercise, do you think that dialysis membrane is permeable to sucrose? Why or why not? Explain. g) How does osmosis differ from diffusion? How are osmosis and diffusion alike? Experiment 2: Effect of Solute Size on Diffusion and Selective Permeability of Membranes (15 min/60 min complete) Introduction: The purpose of this experiment is to demonstrate the effects that a solutes size will have on the ability to pass through a selectively permeable membrane. You already know from the exercise 1 that osmosis is the movement of water. Water, H2O, is a smaller molecule so it can pass freely across the dialysis tubing. This experiment will test the rate of diffusion on larger molecules with a selectively permeable membrane. Page 2 of 4

4 Procedure: 1. Make a table in your lab notebook like the one listed below and write your hypothesis about what will occur with each bag in the beaker pertaining to the solute. Visually identify the color change in your results on a scale of (0, +, ++, +++) where 0 is no color change and +++ is the most color change. (i.e. white to brown would be +++ and white to yellow would be +) Beaker/Content Number 0 min. 15 min. 30 min. 45 min. 60 min. Beaker 2a: dialysis bag 10% starch Beaker 2a: Iodine contents 200 ml Beaker 2b: dialysis bag Iodine (I 2KI) Beaker 2b: 10% Starch contents 200 ml 2. Soak two sections of dialysis tubing provided in water for a few seconds. 3. Label two 400 ml beakers with a china marker to correspond with the bag contents from your table listed above. (2a, 2b) 4. Fill the beakers with the given contents above with 200 ml for each. 5. Using a small pipet, take each content and fill one bag with 10 ml of the starch solution and the other with 10 ml of the iodine solution. Ensure you use the same method for tying each bag as in Exercise 1 to prevent leaking. 6. After filling the bags, rinse the outside with water and blot dry with a paper towel. 7. Place bag 2a into your labeled beaker 2a containing iodine. Place bag 2b into your beaker labeled 2b containing starch. 8. Record in the time (0 min.) the initial color of the two dialysis bags and beaker contents in your table. 9. At each given time interval record the information about the color change of the beaker and the dialysis bags. 10. While waiting do a small visual test by placing the starch into a beaker and placing a drop of iodine into it. Observe the reaction. This will give you a visual reference as to what you are seeing over time in the two beakers. 11. After your final observation at 60 minutes, clean all equipment and discard of dialysis tubing in the trash. 12. Answer the following questions in your lab notebook: a) Was your original hypothesis for each dialysis bag in its beaker correct? Why or why not? b) Make a description about what you observed and why this happened. c) I2KI has a molecular weight of 420 and starch ranges from 20, ,000 because it is such a large and variable polysaccharide. What physical effect does this have on the dialysis tubing? Page 3 of 4

5 Experiment 3: Effects of Concentration in Plant Cells (15 min) Introduction: In the past experiments we looked at the effects of a selectively permeable membrane pertaining to the passage of molecules and water. Given what you know about the structure of a cell in animals vs. a plant, what effect will a solutes concentration have on a plants cell wall? Remember that a plant cell has a large vacuole that also has a selectively permeable membrane. The vacuole typically is used to store pigments, wastes, salts, minerals, nutrients and proteins needed for functioning. The vacuole takes up about 80% of the space in a plant cell. The turgidity of the cell wall surrounding the plant cell is made of cellulose, hemicellulose, and lignin. This is needed because the concentration of solutes is higher inside the vacuole than that of the surrounding cytoplasm in the cell. Plants also contain a plasma membrane as well. During plasmolysis a cell loses its plasma due to the hypertonic solution outside the cell. In this experiment you will discover the effect that the external solute concentration has on the structure of plant cells. Procedure: 1. With forceps, take two leaves from the tip of an Elodea plant. 2. Mount one leaf in a drop of distilled water (dh2o) on a microscope slide. 3. Mount the second leaf in a drop of 20% salt (NaCl) solution. 4. Place a coverslip over both leaves to protect the microscope. 5. Observe the leaf in distilled water first. Draw in your notebook the cells you are seeing and label them. (Chloroplasts, Cell wall, Nucleus, Cytoplasm) 6. Observe the leaf in 20% NaCl solution. Draw in your notebook the cells you are seeing and label them if possible. 7. After you are finished clean all slides and return all equipment to its proper storage area. 8. Answer the following questions in your lab notebook: a) Were the contents of the vacuole in the Elodea leaf in distilled water hypotonic, isotonic, or hypertonic compared to the dh2o? b) Was the 20% NaCl solution hypertonic, hypotonic, or isotonic in relation to the cytoplasm? c) If a hypotonic and hypertonic solution are separated by a selectively permeable membrane, in which direction will the water move? d) Name two selectively permeable membranes that are present within the Elodea cells and that were involved in the plasmolysis process. e) Is the solution in which the cells have been places hypotonic, hypertonic, or isotonic relative to the cytoplasm? How do you know? f) What happens at the cellular level when a wilted plant is watered and begins to recover from the wilt? g) In your notebook draw three separate boxes or circles. In these spaces draw one containing an isotonic, hypotonic, and hypertonic solution and what the cell looks like in each one. Page 4 of 4