osmosis and diffusion experiment Introduction The cell or plasma membrane, because of its molecular composition, is selectively permeable. It also acts to keep the useful cell contents inside while allowing the waste products of cell metabolism to pass to the outside. Two transport systems operate to determine what passes across the plasma membrane: an active transport system and an passive transport system. In the practical today, you will be looking at passive movement processes viz. osmosis and diffusion. You will be using red blood cells to investigate osmosis. Blood is classified as a connective tissue. As such, it consists of two components, a fluid component called plasma and a cellular component. The separation of the cells from the plasma can be measured and is called the Packed Cell Volume (PCV). Plasma will equal approximately 55% of blood volume and the cellular component about 45%. These values will vary slightly with gender e.g. 48% in males and 42% in females. Learning Objectives. 1. To demonstrate diffusion and osmosis and differentiate between the two processes. 2. To define the terms hypertonic, isotonic and hypotonic. 3. To consider the significance of your findings in terms of the workings of the human body. Links: This practical links with the teaching on cells. References: Your anatomy and physiology textbook, such as Marieb (2010) will include a section explaining osmosis and diffusion. See p75 Cell physiology. A useful website explaining diffusion and osmosis with helpful animations: http://www.biologycorner.com/bio1/diffusion.html# WARNING! Chemicals are hazardous to the health of the user and others if inappropriately handled. Read all relevant substance safety data sheets (SSDS) before performing the experiment. PRECAUTIONS Avoid bodily contact with chemicals and solvent. Use in a well ventilated area. Wear disposable gloves and safety goggles. Equipment Glass beakers 7 test tubes Disposable gloves Test tube rack Protective goggles 5 pasteur pipettes Marker pen Printed page of a book Materials Potassium permanganate crystals SSDS No. P1 0.9% sodium chloride SSDS No. S5a 10% sodium chloride SSDS No. S5a 20% glucose 5% glucose 0.5% glucose Distilled water Detergent Horse blood SSDS No. H1 Method 1. DIFFUSION i) Before you start, construct a suitable table on the next page on which to record your observation. In order to construct the table, you will need to read through the following method and work out how many columns and rows you need. ii) Beakers filled with water can be found in the fume cupboard, There will be two sets of three beakers. One set of beakers will contain different concentrations of potassium permanganate (0.003 gms/200mls, 0.03gms/200mls, 0.3gms/200mls) in water at room temperature. The concentration will be clearly stated on each beaker. The other set of beakers will contain 3 crytals of potassium permanganate in water at different temperatures. The temperature will be stated on the beakers. iii) Observe what occurs to the crystals of potassium permanganate in each beakers at the time of the first addition of potassium permanganate. ii) Continue to observe the experiment for the length of the practical and make observations at time 0 minutes, 5 minutes, and then at 10 minute intervals for the first hour, with subsequent observations made at 20 minute intervals until the end of the practical time of 2 hours.. 2. OSMOSIS i) Again, construct tables in which to record your results for the following experiments. In order to construct the tables successfully, you will need to read through the following method and work out how many tables you need, and for each table, how many columns and rows you need e.g. you have 6 test tubes and will need a header row, 7 rows in total. You must also give each table a title. You may decide to present all the results on one table. ii) Mark the test tubes in the rack in front of you with the pen provided: (A ? F). iii) Prepare them by adding the following solutions using a Pasteur pipette: Tube A: add 2 ml 0.9% sodium chloride solution. Tube B: add 2 ml 10% sodium chloride solution. Tube C: add 2 ml distilled water. Tube D: add 2ml 20% glucose solution Tube E: add 2ml 5% glucose solution Tube F: add 2ml 0.5% glucose solution iv) Using a clean Pasteur pipette, add 2 drops of blood to test tube A. Mix gently by inversion (i.e. while wearing gloves, place your thumb over the top of the tube and tip gently upside down. Do not shake). v) Hold the test tube in front of a printed page to see if you can read the print easily through the solution. Record the clarity of print seen through the test tube. vi) Repeat this procedure for the remaining test tubes B to F. Record the clarity of print seen in each case. vii) Return to test tube A. Add 3 drops of detergent to it and observe the effects. Hold the test tube up to a printed page and see if you are now able to read the print through the solution. Think hard about what has happened and why. 2. The solutions used in this experiment were isotonic, hypertonic and hypotonic to the cells. Define what these terms mean and decide which of the solutions that were used could be either isotonic, hypertonic and hypotonic. 3. What happened to the clarity of the solution in test tube A when you added detergent? Can you explain what the detergent is doing to the red blood cells to make the solution become clear and why? (Think about why you use detergent to wash up and what a cell membrane is made of). Blood components and composition: Observe the demonstration whereby blood will be placed into a glass capillary tube. The tube will be sealed with plasticine and placed within a centrifuge. The centrifuge will be switched on and begin to spin the tube samples, the blood cells will be forced to the outside end of the tube by the centrifugal force. The percentage of cells to plasma can then be determined. Note the result from your blood sample. How does this compare with ?normal values??