Mark Pipp (6th hour), Kim Mitchell, Lindsay Mensch

Dr. Knittle
AP Biology - 4th hour
3 February 2015
Cell Respiration Lab
Introduction:
Cellular respiration is a metabolic process that involves the oxidation of glucose to
produce carbon dioxide, water, and energy. This reaction occurs in the mitochondria of each cell
and is a three-step process consisting of glycolysis, the citric acid cycle, and the electron
transport chain/oxidative phosphorylation. The main purpose of cellular respiration is to convert
the chemical energy of organic compounds to energy that all cells can use: ATP. This experiment
will measure the amount of oxygen consumed in germinating and non germinating (dried) peas
at two different temperatures, in order to measure their rates of respiration.

Hypothesis:
Null: The pea seeds at 27oC will consume the same amount of oxygen as the pea seeds at
15oC.
Alternative: The peas at 27oC will consume more oxygen than the pea seeds at 15oC
because the enzymes involved in respiration are more active at higher temperatures.

Procedure:
1. Fill two small tubs with water, one with 15oC water and the other
with 27oC water. Make sure to maintain the temperature of each tub with ice or
hot water throughout the experiment.
2. For first respirometer: Fill a 100 mL graduated cylinder with 50
mL of water and add 25 germinating peas to determine the volume of the peas by

water displacement. Record the volume of the peas, empty the graduated cylinder
and set aside the 25 peas for respirometer 1.
3. For second respirometer: Refill the graduated cylinder to 50 mL
with water and put in 25 dried peas. Add glass beads until the water is displaced
to the same volume as the larger germinating peas. Empty the graduated cylinder
and set aside the 25 peas with the beads for respirometer 2.
4. For third respirometer: Refill the graduated to 50 mL with water
and put in enough glass beads to get the volume equivalent of the germinating
peas. Empty the graduated cylinder and set aside the beads for respirometer 3.
5. Repeat steps 2, 3, and 4 to get a second set of peas and beads for
the second temperature.
6. Assemble six respirometers. Get six vials, six rubber stoppers, and
six pipets. At the bottom of each vial, place a small piece of absorbent cotton
saturated with 15% KOH. Make sure no KOH gets on the sides of the vials. Place
a small piece of nonabsorbent cotton on top of saturated cotton to prevent the
alkaline solution from later touching the test subjects. Be sure to use the amount
of cotton in each vial.
7. Put the germinating peas, non germinating peas with beads, and
only beads into respirometers 1, 2, and 3, respectively, and repeat for
respirometers 4, 5, and 6 with second set of peas and beads. Insert rubber stopper
and pipette into top of vials.
8. Apply petroleum jelly around the base of each pipette where it
enters the rubber stopper. This will prevent unwanted water from entering the
vials.
9. To keep the pipettes out of the water during the 7 minute
equilibrium period, make a masking tape sling across each tub. Vials 1, 2, and 3

should be immersed in the 27oC water and vials 4, 5, and 6 should be immersed in
the 15oC water.
10. Completely immerse the six respirometers into the water. (A small
amount of food coloring can be added to the end of each pipette before they are
immersed so reading is easier). Remember to keep water temperature constant.
11. After a 3 minute equilibration period, record the position of the
water every 5 minutes for 20 minutes to the nearest .01 mL.

Observations:
Our class observed that some of the germinated peas had begun to sprout. Also, some of
the dry peas were gooey and sticky after the experiment, which may have been due to KOH
leaking from under the nonabsorbent cotton and reacting with the peas. If a group experienced a
leak during the experiment, the inside of the respirometer would be noticeably damp and dewy.

Data:

See attached data sheet for results measured by our group.

Class Average of Adj. Diff.

Time

Germinated Pea

Dried Pea Seeds & Glass

Germinated Pea

Dried Pea Seeds & Glass

(min.)

Seeds at 27C (mL)

Beads at 27C (mL)

Seeds at 15C (mL)

Beads at 15C (mL)

0.23

0.02

0.09

0.03

10

0.37

0.19

0.02

15

0.65

0.03

0.33

0.02

20

0.83

0.07

0.42

0.02

25

0.98

0.05

0.53

0.03

30

1.15

0.05

0.56

0.02

Results:
As determined by the class data averages, the germinated peas at 27oC consumed the
most oxygen, at an average rate of .041 mL/min, The germinated peas at 15oC consumed .020

mL O2/min. on average, which is less oxygen than those at 27oC but more than the dried peas and
glass beads at both temperatures. The dried peas at 27oC consumed oxygen at an average rate of .
0022 mL/min and at 15oC, the dry peas consumed oxygen at a rate of .0020.

Conclusion:
The averages of our class experimental data yielded results that were parallel to our
alternative hypothesis. The peas at the higher temperature respired more than those at the lower
temperature. Enzymatic activity is directly proportional to temperature, which accounts for the
increased respiration at 27oC. Two other trends also became evident as we looked at the
germinated peas versus the dried pea seeds. One of these trends was that the germinated peas
consumed more O2 than those that were not germinating. The dried peas were dormant - meaning
they were not actively growing - so they only needed to respire at a very low rate. The
germinated ones, however, needed to convert the stored chemical energy in the seed to the usable
form of ATP in order to grow.
Another evident trend was that the dried peas did not consume a significant amount of
oxygen over the thirty minutes of the experiment. Essentially, there was no change in the volume
of oxygen consumed. These non germinating peas were not dead, so they had to be performing
respiration, even at a very low rate; the lack of change in oxygen consumed suggests that the
respirometers we used were not sensitive enough to detect minute changes in volume. In fact, the
pipettes only gave accuracy to the hundredths place.
Several potential errors could have occurred, which include inconsistent water bath
temperature during the lab and human calculation errors. Despite these possibilities, the data we
uncovered can be explained logically and scientifically. Overall, our data was consistent with our
alternative hypothesis and scientific knowledge.

Design Your Own Experiment:

Our experiment will test respiration rate in pea plants at various developmental stages. It
will be performed in the same respirometers set up in the aforementioned lab (including the
KOH saturated cotton to absorb CO2 and nonabsorbent cotton). Respiration will be measured by
determining the volume (in mL) of oxygen consumed. Instead of testing the effect of temperature
on the rate of cellular respiration, the independent variable will be different stages of growth for
the plants. A pea seedling (a plant that has a developing root system and a few leaves) will be
placed in the first respirometer. A few germinated pea seeds (equal in volume to the seedling in
respirometer 1) will be placed in the second respirometer. In the third respirometer, we will place
glass beads in a volume equal to the seedling, which will act as a control. The experiment will
proceed as the previous one did, where measurements will be recorded every 5 minutes for 30
minutes.
We predict that the experiment would show results in which the germinated pea seeds
consumed a greater amount of oxygen. This is expected because they are heterotrophs, living off
of energy stored within the seed; respiration is their only means of obtaining usable energy. Since
the seedlings have leaves, they are capable of photosynthesis and not dependent on cellular
respiration alone. Since the KOH in the seedlings respirometer will take in some of the carbon
dioxide, small amounts of the gas from the air will enter the stomata and be converted to oxygen
in photosynthesis. This will result in little or zero net change in the readings of the respirometer;
the CO2 released and absorbed by the cotton saturated in base will be replaced by the O2 formed
by photosynthesis.

Questions:

2. This activity uses a number of controls. Identify at least three of the controls, and
identify the purpose of each control.
One experimental control is the temperature, which ensures that the results won't be
affected by pressure changes within the respirometers. Since they are nonliving, and therefore do
not perform respiration, the glass beads are also a control for the peas, which do respire. This
guarantees that the vials do not simply lose air, but that in fact oxygen is consumed by the
process cellular respiration. Lastly, the volume of the peas and beads in all the respirometers was
held constant to keep pressure in each vial the same.
4. Describe and explain the relationship between the amount of O consumed and time.
During the first 15 minutes of the experiment, oxygen was consumed quickly because it
was still highly concentrated in the respirometer. Then, from the 15 to 25 minute marks, the
oxygen consumption started to decrease slightly since oxygen became less abundant. Oxygen
consumption decreased greatly during the last 5 minutes since there was not much oxygen left.
6. Why is it necessary to correct the readings from the peas with the readings from the
beads?
The glass bead readings show that oxygen was lost without the process of respiration at
work. To show the actual oxygen consumption of the dried and germinated peas, the bead
readings needed to be subtracted from the pea readings.
7. Explain the effect of germination (versus nongermination) on pea seed respiration.
The germinated peas were growing to become autotrophic plants, so they needed to
perform increased respiration to utilize stored energy. They used more oxygen than the dried
peas, since they were not germinating or growing.

8. Below is a sample graph of possible data obtained for oxygen consumption by

germinating peas up to about 8C. Draw in predicted results through 45C. Explain your
prediction.

As temperature increases, the rate of respiration will increase (i.e. oxygen consumption
will increase). However, there will come a point where the temperature will become too great for
the pea seeds and their proteins will denature. At that point, oxygen consumption will decrease.
9. What is the purpose of KOH in this experiment?
KOH was an important part of the experiment because it reacted with respirations waste
product, CO2, to form the solid K2CO3. This was essential, because otherwise, there would have
been little or no net movement in the pipettes, since there would be no pressure change from the
conversion of O2 to CO2. In other words, there would be no change in the reading if KOH were
not involved in the experiment.
10. Why did the vial have to be completely sealed around the stopper?
If the vial were not completely sealed, water could leak into the respirometer and disrupt
the experimental results by changing the pressure/volume.

11. If you used the same experimental design to compare the rates of respiration of a 25 g.
reptile and a 25 g. mammal, at 10C, what results would you expect? Explain your
reasoning.
The mammal would respire more because it is an endotherm and needs to thermoregulate.
It can maintain its body temperature by respiring (and therefore generating heat). The reptile
would perform less respiration because it is an ectotherm. It performs respiration mainly to
obtain energy for movement and other cellular processes.
12. If respiration in a small mammal were studied at both room temperature (21C) and
10C, what results would you predict? Explain your reasoning.
The respirometer of the mammal would show more oxygen consumption at 10oC because
it would need to work harder (i.e. respire more) to maintain its body temperature than it would at
room temperature. The colder the environment, the more heat it needs to generate to account for
heat loss and stay warm.
13. Explain why water moved into the respirometers' pipettes.
As oxygen was used, pressure in the respirometer decreased, which allowed water to
move into the pipette.
14. Design an experiment to examine the rates of cellular respiration in peas that have been
germinating for 0, 24, 48 and 72 hours. What results would you expect? Why?
The experiment would involve the four respirometers in one bath that is held at a constant
temperature. There would also be a control respirometer with only beads in it to show how much
oxygen was lost without the involvement of respiration. Once the pea seeds have been sealed in
the respirometers (which contain KOH, absorbent cotton, and nonabsorbent cotton), data will be
collected every five minutes for thirty minutes. We would expect results to show higher rates of

respiration in the peas that have been germinating longer because they are trying to grow to
become plants. The peas that have not yet germinated will yield results showing no respiration.
Since they are dormant, significant amounts of respiration will not be detected over the course of
the experiment.