AP Investigation #6: Cellular Respiration
What factors affect the rate of cellular respiration in multicellular organisms?
Background
Living systems require free energy and matter to maintain order, to grow, and to reproduce. Energy
deficiencies are not only detrimental to individual organisms, but they cause disruptions at the
population and ecosystem levels as well. Organisms employ various strategies that have been
conserved through evolution to capture, use, and store free energy. Autotrophic organisms capture
free energy from the environment through photosynthesis and chemosynthesis, whereas
heterotrophic organisms harvest free energy from carbon compounds produced by other organisms.
The process of cellular respiration harvests the energy in carbon compounds to produce ATP that
powers most of the vital cellular processes. In eukaryotes, respiration occurs in the mitochondria
within cells.
If sufficient oxygen is available, glucose may be oxidized completely in a series of enzyme-mediated
steps, as summarized by the following reaction:
More specifically,
The chemical oxidation of glucose has important implications to the measurement of respiration.
From the equation, if glucose is the energy source, then for every molecule of oxygen consumed, one
molecule of carbon dioxide is produced.
Suppose you wanted to measure the overall rate of cellular respiration. What specific things could
you measure? Which of these might be easier or harder to measure? In this laboratory investigation,
you will learn how to calculate the rate of cellular respiration by using a respirometer system. These
measure relative volume (changes in pressure) as oxygen is consumed by germinating plant seeds. As
oxygen gas is consumed during respiration, it is normally replaced by CO2 gas at a ratio of one
molecule of CO2 for each molecule of O2. Thus, you would expect no change in gas volume to result
from this experiment. However, in the following procedure the CO2 produced is removed by
potassium hydroxide (KOH). KOH reacts with CO2 to form the solid potassium carbonate (K2CO3)
through the following reaction:
Thus, as O2 is consumed, the overall gas volume in the respirometer decreases. The change in
volume can be used to determine the rate of cellular respiration. Because respirometers are sensitive
to changes in gas volume, they are also sensitive to changes in temperature and air pressure; thus,
you need to use a control respirometer.
As you work through Procedure 1, think about this question: What factors can affect the rate of
cellular respiration? In Designing and Conducting Your Investigation, you will design and conduct an
experiment to investigate at least one of your responses to this question or some other question you
have. Your exploration will likely generate even more questions about cellular respiration.
This investigation also provides an opportunity for you to apply and review concepts that you have
studied previously, including the relationships between cell structure and function (mitochondria);
enzymatic activity; strategies for capture, storage, and use of free energy; diffusion of gases across cell
membranes; and the physical laws pertaining to the properties and behaviors of gases.
A number of physical laws relating to gases are important to the understanding of how the apparatus
that you will use in this investigation works. The laws are summarized in the general gas law that
states:
PV = nRT
where P = pressure of the gas
V = volume of the gas
n = number of moles of the gas
R = the gas constant (its value is fixed)
T = temperature of gas
This law implies the following important concepts about gases:
1. If the temperature and pressure are kept constant, then the volume of the gas is directly
proportional to the number of molecules of the 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 is kept constant, then either the
pressure or volume (or both) will change in direct proportion to the temperature.
It is also important to remember that gases and fluids flow from regions of high pressure to regions
of low pressure.
Procedure 1: Rate of Cellular Respiration of Alaska Peas
The amount of O2 consumed will be measured over a period of time. Three respirometers should be
set up as follows:
Respirometer
1
2
3
1.
Temperature
Room Temperature
Room Temperature
Room Temperature
Contents
Germinating Seeds
Dry Seeds + Beads
Beads
Respirometer 1: Obtain a 100 ml graduated cylinder and fill it with 50 ml of H20. (Remember to
be as accurate as possible…..squat down to view the meniscus at eye level.) Drop 15 germinating peas in
the graduated cylinder and determine the amount of water that was displaced (which is
equivalent to the volume of the peas). Record the volume of these
Germinating Pea Volume
= _________ ml
peas. Remove these peas and place them on a paper towel. They will
be used in respirometer 1.
2. Respirometer 2: Refill the graduated cylinder with 50 ml of H20. Drop 15 dry peas (nongerminating) into the graduated cylinder and then add enough glass beads to attain a volume
equivalent to that of the germinating peas. Remove these peas and beads and place them on
a paper towel. Take a few seconds to dry these peas as much as possible with the paper towel.
These peas and beads will be used in respirometer 2.
3. Respirometer 3: Refill the graduated cylinder with 50 ml of H20. Determine how many glass
beads would be required to attain a volume equivalent to that of the germinating peas.
Remove these beads and place them on a paper towel. These beads will be used in
respirometer 3.
4. To assemble the respirometers, obtain three vials, each with an attached stopper and pipette.
Place a small piece of absorbent cotton in the bottom of each vial and, using a dropper,
moisten the cotton with 15% KOH. The cotton is to be moistened, not dripping wet. It is
estimated that you should use approximately 1.0-1.5 ml of KOH. Make sure that you don’t get
KOH on the sides of the respirometer, as this will kill the peas if they come in contact with the
KOH. It is imperative that the amount of cotton and KOH be the same for each respirometer.
Place a small wad of nonabsorbent cotton on top of the KOH-soaked absorbent cotton.
5. On top of the non-absorbent cotton, place the 15 germinating peas in respirometer 1, the dry
peas and glass beads in respirometer 2, and the glass beads in respirometer 3. Insert the
stopper fitted with the calibrated pipette. Place a weighted washer on the end of each vial (to
hold the pipette down in the water).
6. Make a sling of masking tape attached to each side of the water bath to hold the pipettes out
of the water during the equilibration period of 5 minutes.
7. After this equilibration period, immerse all respirometers entirely in the water bath. Water will
enter the pipettes for a short distance and then stop. If the water continues to move into a
pipette, check for leaks in the respirometer. Move the weighted washer to a place that
prevents the pipette from floating in the water (preferably either at the tip or near the vial).
Rotate the respirometers so that you can read the numbers through the water. After this time,
your hands are not allowed to touch the water (and the respirometers). If the water or
respirometers are shifted at any time during the experiment, your results will be affected.
8. Allow the respirometers to equilibrate for 3 more minutes, after your hands are done
manipulating the vials in the water. At the end of this three minutes, begin the experiment
with the first reading at Time 0. To do this, record, to the nearest 0.01 ml the position of the
water line in each pipette. Every 5 minutes, for 20 total minutes, take readings of the water’s
position in each pipette and record in the table. When reading the pipette, be careful that you
are recording the right number….pay attention to where zero is on the pipette.
Table 1: Measurement of O2 Consumption by Soaked and Dry Pea Seeds at Room Temperature
Using Volumetric Methods
Time
(min)
Respirometer 3:
Beads Alone
Reading at
time X
Difference*
Respirometer 1: Germinating Peas
Reading at
time X
Difference*
Corrected
difference
**
Respirometer 2: Dry Peas + Beads
Reading at
time X
Difference*
Corrected
difference
**
0
5
10
15
20
*Difference= (initial reading at time 0) – (reading at time X)
**Corrected difference = (initial pea reading at time 0 – pea reading at time X) – (initial bead reading at time 0 – bead reading at time X)
Analysis (in your lab notebook)
1. Graph the results from the corrected difference column for the germinating peas and dry peas
at room temperature. You should create a line graph, give it an appropriate title, place your
independent and dependent variables on the correct axes, and include a key.
2. Determine the rate of O2 consumption of the germinating and dry peas. Create a data table to
Δ𝑦
show these rates. Show your calculations. Recall that rate = Δ𝑥
3. Describe and explain the difference in oxygen consumption over time in both types of peas.
Also compare the rates of respiration in your answer. Use evidence from the experiment in
your answer.
4. Explain why water moved into the respirometers’ pipettes.
Designing and Conducting Your Investigation
Now that you have learned how to measure the rate of cellular respiration in germinating seeds, you
now have the opportunity to explore questions about cellular respiration you might have. 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 non-germinating 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)?



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Do small seeds of spring flowers, weeds, or grasses respire at a different rate from seeds
from summer, fall, or winter plants?
Is there any relationship between the mass of the organism and its rate of respiration?
Do seeds from monocot plants respire at different rates from dicot plants?
Can the same respirometer system be used to measure the rate of respiration in small
invertebrates, such as insects or earthworms?
Design an experiment to investigate one of your own questions about cellular respiration. When
identifying your design, be sure to address the following:
 What is the essential question being addressed?
 What measurements will you choose to take? Will they 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?
 Identify your dependent and independent variables.
 Identify your hypothesis, which should include a prediction about the effect of the factor(s) you
chose to investigate on the rate of cellular respiration.