AP Biology updated Jan. 2010 Cellular Respiration Lab Cellular Respiration – Adapted from Laboratory 5 in AP Student Manual Introduction: Cellular respiration is the release of energy from organic compounds by metabolic chemical oxidation in the mitochondria within each cell. Cellular respiration involves a series of enzyme-mediated reactions. The equation below shows the complete oxidation of glucose. Oxygen is required for this energy-releasing process to occur. C6H12O6 + 6O2 -----> 6 CO2 + 6 H2O + 686 kilocalories of energy /mole of glucose oxidized By studying the equation above, you will notice there are three ways cellular respiration could be measured. One could measure the: 1. Consumption of O2 (How many moles of oxygen are consumed in cellular respiration?) 2. Production of CO2 (How many moles of carbon dioxide are produced by cellular respiration?) 3. Release of energy during cellular respiration. Objectives calculate the rate of cell respiration from experimental data. relate gas production to respiration rate; and test the effect of temperature on the rate of cell respiration in nongerminated versus germinated seeds in a controlled experiment. In this experiment, the relative amount (ppm) of CO2 produced by germinating and nongerminating (dry) peas at two different temperatures will be measured. Overview: 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. When seeds become moist, and water penetrates into them, enzymes begin the process of cellular respiration to provide the new plant with the fuel that it needs for the building of new material. Cellular respiration is the process of oxidizing food molecules, such as sugar, into carbon dioxide and water. Therefore, an increase in (CO2) concentration inside the flask with the germinating seeds is evidence that cellular respiration is occurring as the seeds germinate. In this experiment you will measure carbon dioxide production during germination. You will measure the change in gas production 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. 1 AP Biology updated Jan. 2010 Cellular Respiration Lab The amount of carbon dioxide will be measured in __ minute intervals (tbd), over a period of 20 minutes. Four respirometers should be set up as follows: Respirometer Temperature Contents 1 Room Germinating seeds 2 Room Dry Seeds 3 0 10 C Germinating Seeds 4 0 Dry Seeds 10 C FIRST: write down your hypotheses! What do you predict will happen and why? Materials Computer with USB ports PASPORTtm USB link PASPORTtm CO2 Gas sensor 250 mL Erlenmeyer flasks Pea seeds (dry and germinated) Cold water bath thermometers Procedure 1. Prepare a room-temperature station (approx. 25 degrees Celsius) and a cold-water bath (approx. 10 degrees Celsius). 2. Count out 100 germinating peas and 100 nongerminating peas and put into two separate 250 mL Erlenmeyer flasks. To assemble respirometers, insert the stopper with the CO2 sensor into each flask mouth to get a tight seal. 3. Using the same procedure as in the previous step, to assemble 2 more respirometers*. These will go into the 10-degree bath. 2 AP Biology updated Jan. 2010 Cellular Respiration Lab *This can be done in two sets, at different temperatures, if space or time is limited. 8. Connect the CO2 sensors to the USB ports of a computer using the USB link and allow the respirometers an equilibration period of 1 minute before taking measurements. Respirometers 1 and 2 should be kept at room temperature bath, and the other two should be in the 10 degree bath. 9. After equilibration, take an initial CO2 measurement, after which a reading should be taken every 5 minutes for a period of 20 minutes. Check the temperature and record all data. This collective data should be displayed in table form in your report (Table 5.1). A graph (Graph 5.1) should display these data. Results: a. Table 5.1: Measurement of CO2 Production by Germinating and Dry Pea Seeds at Room Temperature (___0C) and 100C in ppm. b. Graph 5.1 : the results for respiration of the peas at both room temperature and 10 0C. c. Table 5.2 : From the slope of the four lines on the graph, determine the overall rate of CO2 Production of germinating and dry peas during the experiments at room temperature and 100C. Recall that rate = Δ Y/ Δ X. See below. Table 5.2 Rate of CO2 Production Condition Germinating Peas/100C Show Calculations Here The house is red and sometimes blue Germinating peas /Room The house is red and somue Temperature sometimes blue Rate in ppm CO2 / min The house is red and sometimes blue The house is red and sometimes blue Dry peas/100C The house is red and sometimes blue The house is red and sometimes blue Dry Peas /Room Temperature The house is red and sometimes blue The house is red and sometimes blue The house is red and sometimes blue The house is red and sometimes blue 3 AP Biology updated Jan. 2010 Cellular Respiration Lab For DISCUSSION (may be done on separate paper) 1. VOCABULARY- define the following: Adenosine diphosphate (ADP) Adenosine triphosphate (ATP) Carbon dioxide (CO2) Cellular respiration Germination Dormancy 2. In this investigation, you are investigating both the effect of germination versus nongermination and warm temperature versus cold temperature on respiration rate. From the data collected (tables and graph), can you accept or reject your hypotheses? Why/why not? 3. This activity uses a number of controls. Identify at least three of the controls, and describe the purpose of each control. 4. Analyze the graphed results for respiration of the peas at both room temperature and 10 0C. a. What is the independent variable? b. What is the dependent variable? 4 AP Biology updated Jan. 2010 Cellular Respiration Lab 5. Describe your graph, in as much detail as possible. 6. Table 5.2 shows CO2 production of germinating and dry peas during the experiments at room temperature and 100C. Looking at this, and the slope of the line of Graph 5.1, during which time interval was gas production the greatest? The least? How might different variables affect this? 7. Describe and explain the relationship between the amount of CO2 produced and time. 8. Summarize the effect of germination (versus nongermination) on peas seed respiration. 9. Why did the respirometer have to be completely sealed around the stopper? What are some possible errors that could take place in this experiment? 10. If you used the same experimental design to compare the rates of respiration of a 25g. reptile and a 25g. mammal, at 100C, what results would you expect/ Explain your reasoning. 11. If respiration in a small mammal were studied at both room temperature (210C) and 100C, what results would you predict? Explain your reasoning. 12. Read through Lab 5 in the AP Student Laboratory Manual. (a) Note at least three differences between that lab setup and the way that we did our lab. (b) Did we achieve the same objectives, using different methods? Explain. 5