C 6 H 12 O 6 + 6O 2 6CO 2 + 6H 2 O ATP Glucose Oxygen Carbon Water Dioxide

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1 CELLULAR RESPIRATION Pre-lab Questions 1. What types of cells undergo aerobic respiration? What are the reactants and products of each phase of respiration, and where in the cell do they occur? 2. In this lab we will be using germinate beans and peas. What does it mean to be germinated? Why do these peas do respiration and not photosynthesis? Introduction In this experiment, you will work with seeds that are living but dormant. A seed contains an embryo plant and a food supply surrounded by a seed coat. When the necessary conditions are met, germination occurs, and the rate of cellular respiration greatly increases. In this experiment you will measure oxygen consumption during germination. You will measure the change in gas volume in respirometers containing either germinating or nongerminating pea seeds. In addition, you will measure the rate of respiration of these peas at two different temperatures. Cells break down glucose molecules to produce [generate] ATP (adenosine triphosphate). ATP can then be used to run cellular processes that require energy. ATP is the energy source to run the cellular process. This process is called Cellular Respiration. Energy is used to break down glucose molecules and oxygen into carbon dioxide and water. In the Photosynthesis lab, you learned that carbon dioxide and water in combination with light energy produces glucose and oxygen. C 6 H 12 O 6 + 6O 2 6CO 2 + 6H 2 O ATP Glucose Oxygen Carbon Water Dioxide There are several methods to measure the rate of cellular respiration in an organism. One method would be to monitor changes in temperature because the process of respiration produces heat. Another way would be to measure either the oxygen consumption or carbon dioxide production. A Respirometer is a device used to take such measurement; it measures gas volume changes. From this information, the rate of cellular respiration can then be determined. During cellular respiration, two gases are changing volume. Oxygen is being consumed and carbon dioxide is diffusing out of the cells and released. The respirometer must try to measure both gases at the same time. In order to accomplish this, Potassium Hydroxide (KOH) is added to the device. KOH absorbs carbon dioxide and forms Potassium Carbonate and water. CO 2 + 2KOH K 2 CO 3 + H 2 O Carbon Potassium Potassium Water Dioxide Hydroxide Carbonate The potassium carbonate produced is a solid precipitate. Any carbon dioxide produced will be immediately converted into a solid and therefore the respirometer will then be able to measure only one variable the consumption of oxygen. In the experimental apparatus if water temperature and volume remain constant, the water will move toward the region of lower pressure. During respiration, oxygen will be consumed. Its volume will be reduced, because the carbon dioxide produced is being converted to a solid. The net result is a decrease in gas volume within the tube, and a related decrease in pressure in the tube. The vial with glass beads alone will permit detection of any changes in volume due to atmospheric pressure changes or temperature changes. The amount of oxygen consumed will be measured over a period of time. Background Information

2 A number of physical laws relating to gases are important to the understanding of how the apparatus that you will use in this exercise works. The laws are summarized in the general gas law that states: PV = nrt where P is the pressure of the gas, R is the gas constant (its value is fixed) V is the volume of the gas, T is the temperature of the gas (in K0) n is the number of molecules of gas This law implies the following important concepts about gases: 1. If temperature and pressure are kept constant, then the volume of the gas is directly proportional to the number of molecules of gas. 2. If the temperature and volume remain constant, then the pressure of the gas changes in direct proportion to the number of molecules of gas present. 3. If the number of gas molecules and the temperature remain constant, then the pressure is inversely proportional to the volume. 4. If the temperature changes and the number of gas molecules are kept constant, then either pressure or volume (or both) will change in direct proportion to the temperature. 5. It is also important to remember that gases and fluids flow from regions of high pressure to regions of low pressure.

3 3 Vials 6-8 Cotton Balls 1 bottle 15% KOH 1 bottle red food coloring ~30 germinated peas ~30 non-germinated peas ~25 blue plastic beads RESPIROMETER SET UP Each group will need the following: 1 large tray for water 3 Volumeters (rubber stopper with serological pipet inserted) Marker Pen Goggles Nitrile gloves Wooden sticks Long handle Forceps Masking Tape Thermometer Gloves and goggles should be worn at all times during this experiment. Potassium Hydroxide (KOH) is a strong caustic base. Follow the safety instructions given to you by your lab instructor for disposal of the KOH saturated cotton and peas. Timer Procedure 1. Label the Vials # s Place one absorbent cotton ball in the bottom of each vial. Use a wooden stick or long forceps to push the cotton to the bottom of the vial. 3. Open the 15% KOH bottle and saturate the cotton with a few drops of potassium hydroxide. Use the same number of drops for each vial. 4. Place one dry non-absorbent cotton ball on top of the saturated cotton ball. 5. Find the volume of 25 germinating peas by filling a 100mL graduated cylinder 50mL and measuring the displaced water. Place these peas in vial #1 6. Fill the graduated cylinder with 50mL water, 25 non-germinating peas, and add enough glass beads to attain an equal volume to the germinating peas. Place these in vial #2. 7. Using the same procedure as in the previous two steps, find out how many glass beads are required to attain the same volume as the 25 germinating peas. Place this in vial #3. 8. Insert a rubber stopper with a serological pipet into each vial. Make sure the pipet has a tight fit.. 9. Turn the pipet so that the numbers can be easily read when the vials lay on their sides in the water tray. 10. Put a drop of red food coloring into the tip of the serological pipet. 11. Completely fill a tray with water. (may be done already for you) 12. Add metal washers around the neck of the repirometer to weigh it down in the water 13. GENTLY lay the 3 vials into the tray of water. It is important to be SLOW and GENTLE or the red food coloring will shoot out the tip of the pipet. 14. Make sure all vials and volumeters (rubber stoppers and serological pipet) are under water. Add more washers as necessary. 15. Take a reading of where the red food coloring is located in the serological pipet. 16. Record the position of the red food coloring in each pipet at the end of 5, 10, 15, 20 and 25 minutes. Place data in Table # When complete, remove the respirometers from the water.

4 DATA TABLE #2 CALCULATION OF OXYGEN CONSUMPTION Respirometer 1 Germinating Seeds Time Interval 0 min 3 min 10 min 15 min 20 min 25 min Reading (ml) Volume Change ml (reading time 0) Respirometer 2 Non-Germinating Seeds Time Interval 0 min 3 min 10 min 15 min 20 min 25 min Reading (ml) Volume Change ml (reading time 0) Respirometer 3 Glass Beads Only Time Interval 0 min 3 min 10 min 15 min 20 min 25 min Reading (ml) Volume Change ml (reading time 0) Graph: Plot the independent variable on the X axis (time) and the dependent variable on the Y axis (change in reading on the pipet). Graph a line for: Germinating Peas Non-Germinating Peas Beads **Use a line of Best Fit so that you can calculate the slope**

5 Questions: 1. Calculate the RATE of oxygen consumption for each respirometer by calculation the SLOPE of the line from your graph. Be sure to include units! 2. What was the purpose of the: a. Beads: b. KOH 3. Explain why the water moved into the pipet. 4. Why was it necessary to absorb the carbon dioxide? 5. Which respirometer in this experiment was the Control? Why? 6. If you performed the lab without soaking the cotton with KOH, what results would you predict? Why? 7. If you used the same experimental design to compare the rates of respiration of a 25 g reptile and a 25 g mammal, at 100C, what results would you expect? Explain your reasoning. 8. If respiration in a small mammal were studied at both room temperature (210C) and 100C, what results would you predict? Explain your reasoning.

6 CELLULAR RESPIRATION - INQUIRY Introduction Now that you have learned how to measure the rate of cellular respiration in germinating seeds, you have a tool for exploring questions on your own. Think about the process of cellular respiration. When does it occur? Are there any situations when living cells are not respiring? Why might some living cells respire more than others? Are there differences between major groups of organisms in how fast they respire? What is the difference, if any, in the rate of cellular respiration between germinating seeds and nongerminating seeds? Does the temperature of germinating seeds affect the rate of cellular respiration? Do plant seeds consume more oxygen at higher temperatures than at lower temperatures? Do germinating seeds just starting to germinate consume oxygen at a greater rate than seeds that have been germinating for several days (age dependence)? Do small seeds of spring flowers, weeds, or grasses respire at a different rate from seeds from summer, fall, or winter plants? Can the same respirometer system be used to measure the rate of respiration in small invertebrates, such as insects or earthworms? Step 1 Design an experiment to investigate one of your own questions about cellular respiration or one of the questions above using respirometers. When identifying your design, be sure to address the following: What is the essential question being addressed? What assumptions are made about the question(s) being addressed? Can those assumptions be verified? Will the measurements you choose to make provide the necessary data to answer the question under study? Did you include a control in your experiment? What are possible sources of error in the experiment(s)? Give your teacher a list of materials needed and a rough outline of your procedure ASAP so that materials can be procured in time for the experiment. Step 2 Make a hypothesis, which should include a prediction about the effect of the factor(s) you chose to investigate on the rate of cellular respiration. Step 3 Conduct your experiment(s) and record data and any answers to your questions. Step 4 Record your data using appropriate methods, such as the example table provided in Procedures. Then graph the results to show the effect of the factors/variables you investigated on the rate of cellular respiration. Calculate the rate(s) of cellular respiration for each factor/variable. Lab-write up DUE and worth pts 1. Make a title that reflects our independent and dependent variables. 2. List the materials and methods in your experiment. 3. Results: Include a data table, graph(s), and 1-2 sentence summaries of graph(s). Also, show your calculations for finding the rate of respiration in your experimental variables. 4. ANALYSIS:

7 Spend some time analyzing your results. Relate the results from your experiment to the experiment you conducted the day before. How was the rate of cellular respiration affected by the experimental variable(s) you chose as compared to the control(s)? 5. CONCLUSION: Write a conclusion for this lab. Was your initial hypothesis about the effect of your factor on the rate of cellular respiration supported? Why or why not? What did you learn from this lab? What were some challenges you had in performing your experiment? Did you make any incorrect assumptions? If so, explain. This would be a good time to include some research about cellular respiration. Cite all references used in your research. Class Presentations will be and worth pts Notes: