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RESPIRATION |
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Introduction 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. In this experiment you will measure
oxygen consumption during germination. You will measure the change in gas
volume in respirometers containing either
germinating or non-germinating pea seeds. In addition, you will measure the
rate of respiration of these peas at two different temperatures. 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 à 6 CO2 + 6 H2O
+ 686 kilocalories of energy / mole of glucose 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. In this experiment, the relative volume
of O2 consumed by germinating and nongerminating
(dry) peas at two different temperatures will be measured. |
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Materials |
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Background
Information 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, · V is the volume of the gas, · n is the number of molecules of gas · R is the gas constant (its value is
fixed) · T is the temperature of the gas (in K0) 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. In this experiment, the CO2 produced
during cellular respiration will be removed by potassium hydroxide (KOH) and will form solid
potassium carbonate (K2CO3) according to the following reaction. CO2 + 2 KOH à K2CO3 + H2O Since the carbon dioxide is being
removed, the change in the volume of gas in the respirometer
will be directly related to the amount of oxygen consumed. 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. At least six respirometers should be set up Using the procedure below. |
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Procedure 1. Prepare a room-temperature bath
(approx. 25o C) and a cold-water bath (approx. 10º C). 2. Find the volume of 25 germinating peas
by filling a 100mL graduated cylinder 50mL and measuring the displaced water. 3. Fill the graduated cylinder with 50mL
water, 25 nongerminating peas, and add enough glass
beads to attain an equal volume to the germinating peas. 4. 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. 5. Repeat steps 2-4. These will go into
the 10ºC bath. 6. To assemble 6 respirometers,
obtain 6 vials, each with an attached stopper and pipette. Number the vials.
Place a small wad of absorbent cotton in the bottom of each vial and, using a
dropper, saturate the cotton with 15% KOH (potassium
hydroxide). It is important that the same amount of KOH be used for each respirometer. 7. Place a small wad of dry, nonabsorbent
cotton on top of the saturated cotton. 8. Place the first set of germinating
peas, dry peas & beads, and glass beads in the first three vials,
respectively. Place the next set of germinating peas, dry peas & beads,
and glass beads in vials 4, 4, and 6, respectively. Insert the stopper with
the calibrated pipette. Seal the set-up with silicone or petroleum jelly.
Place a weighted collar on each end of the vial. Several washers around the
pipette make good weights. 9. Make a sling of masking tape attached
to each side of the water baths. This will hold the ends of the pipettes out
of the water during an equilibration period of 7 minutes. Vials 1, 2, and 3
should be in the room temperature bath, and the other three should be in the
10 degree bath. 10. After 7 min,
put all six set-ups entirely into the water. A little water should enter the
pipettes and then stop. If the water continues to enter the pipette, check
for leaks in the respirometer. 11. Allow the respirometers
to equilibrate for 3 more minutes and then record the initial position of the
water in each pipette to the nearest 0.01mL (time 0). Check the temperature
in both baths and record. Record the water level in the six pipettes every 5
minutes for 20 minutes. |
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Analysis
of Results 1. Identify the hypotheses being tested
in this activity. 2. This activity uses a number of
controls. Identify at least three of the controls, and describe the purpose
of each control. 3. Prepare a table of your data. 4. Graph the results from the corrected difference column for the
germinating peas and dry peas at all of the temperatures (There will be 2
lines on the graph per temperature). 5. Describe and explain the relationship
between the oxygen consumption and time. 6. From the slope of the four lines on
the graph, determine the rate of oxygen consumption of germinating and dry
peas during the experiments at 25°C
and 10°C. 7. Why is it necessary to correct the
readings from the peas with the readings from the beads? 8. Explain the effect of germination
(versus nongerminating) on pea seed respiration. 9. What is the purpose of KOH in this
experiment? 10. If you used the same experimental
design to compare the rates of respiration of a 25 g reptile and a 25 g mammal, at 10°C, what results would
you expect? Explain your reasoning. 11. If respiration in a small mammal were
studied at both room temperature (21°C)
and 10°C, what results would
you predict? Explain your reasoning. |
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