Cellular Respiration

Blake Lockwood

Introduction:
The human body has to have energy in order to perform the functions that allow life. This energy comes from the process of cellular respiration. Cellular respiration releases energy that the body can use in the form of ATP from carbohydrates by using oxygen. Cellular respiration is not just one singular reaction, it is a metabolic pathway made up of several reactions that are enzyme mediated. This process begins with glycolysis in the cytosol of the cell. In glycolysis, glucose is split into two threecarbon compounds called pyruvate, producing a small amount of ATP The final two steps of cellular respiration occur in the mitochondria. These final two steps are the electron transport system and the Krebs Cycle. The overall equation for cellular respiration is C6H12O6 + 6O2 -> 6CO2 + 6H2O + 686 kilocalories of energy per mole of glucose oxidized. There are three ways to measure the rate of cellular respiration. These three ways are by measuring the consumption of oxygen gas, by measuring the production of carbon dioxide, or by measuring the release of energy during cellular respiration. In order to measure the gases, the general gas law must be understood. The general gas law state: 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, and T is the temperature of the gas (in K). The gas law also shows concepts about gases. If temperature and pressure are kept constant, then the volume of the gas is directly proportional to the number of molecules of the gas. 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. If the number of gas molecules and the temperature remain constant, then the pressure is inversely proportional to the volume. If the temperature changes and the number of gas molecules is kept constant, then either pressure of volume will change in direct proportion to the temperature.

In this experiment, the rate of cellular respiration will be measured by measuring the oxygen gas consumption by using a respirometer in water. This experiment measures the consumption of oxygen by germinating and non-germinating at room temperature and at ice water temperature. The carbon dioxide produced in cellular respiration will be removed by potassium hydroxide (KOH). As a result of the carbon dioxide being removed, the change in the volume of gas in the respirometer will be directly related to the amount of oxygen consumed. The respirometer with glass beads alone will show any changes in volume due to atmospheric pressure changes or temperature changes.

Hypothesis:
The germinating peas will have a higher rate of respiration, than the beads and nongerminating peas.

Materials:
This lab requires two thermometers, two water baths, beads, germinating and nongerminating peas, beads, six vials, twelve pipettes, 100 mL graduated cylinder, scotch tape, tap water, ice, KOH, absorbent and non-absorbent cotton, six washers, six rubber stoppers, scotch tape, and a one mL dropper.

Methods:
Start the experiment by setting up two water baths, one at room temperature and the other at 10 degrees Celsius. Then, find the volume of twenty-five germinating peas. Next, put 50 mL of water in a graduated cylinder and put twenty-five non-germinating peas in it. Then, add beads until the volume is the same as twenty-five germinating peas. Next, pour our the peas and beads, refill the graduated cylinder with 50 mL of water, and add only beads until the volume is the same as the twenty-five germinating peas. Repeat these steps for another set of peas and beads. Also, put together the six respirometers by gluing a pipette to a stopper and taping another pipette to the pipette for all six respirometers. Then, put two absorbent cotton balls, several drops of KOH, and half of a piece of non-absorbent cotton into all six vials. Next, add the peas and beads to the appropriate respirometers. Place one set of respirometers into the room temperature water bath and the other set in the ice water bath. Elevate the respirometers by setting the pipettes onto masking tape and allow them to equilibrate for five minutes. Next, lower the respirometers into the water baths and take reading at 0, 5, 10, 15, and 20 minutes. Record the results in the table.

Results:

Table:

Beads Alone

Germinating Peas

Dry Peas and Beads

Reading Reading at time Diff. at time X X Initial 13.2 0 to 5 11.0 2.2

Diff.

Corrected Reading Diff. at time Diff. Corrected Diff. X 12.9 2.2 0.0 11.1 1.8 0.4 0.3 0.5 0.4

12.7 10.5

5 to 10 10

3.2

9.0

3.7

0.5

10.0

2.8

10 to 15 9.2

4.0

8.0

4.7

0.7

9.4

3.5

15 to 20 9.1

4.0

7.5

5.1

1.2

9.3

3.6

Initial 14.0

13.5

14.0 0.3 0.3 0.3 0.3

0 to 5 13.3

0.7

12.1

1.4

0.7

13.6

0.4

5 to 10 12.9

1.1

11.0

2.5

1.4

13.2

0.8

10 to 15 12.6

1.4

10.0

3.5

2.1

12.9

1.1

15 to 20 12.2

01.8

9.0

4.5

2.7

12.5

1.5

Questions
1. In this activity, you are investigating both the effect of germination versus nongermination and warm temperature versus cold temperature on respiration rate. Identify the hypothesis being tested in this activity. Increasing the temperature could increase the oxygen consumption. Germinating peas have a higher respiration rate than non-germinating. 2. This activity uses a number of controls. Identify at least three of the controls, and describe the purpose of each control. One control was the respirometer with only beads in it because it didnt use respiration. Another control was that the water temperatures were constant. The final control was that there was the same amount of KOH in each vial. 3. Graph the results from the corrected difference column for the germinating peas and dry peas at both room temperature and at 10 degrees Celsius. On graph paper. 4. Describe and explain the relationship between the amount of oxygen gas consumed and time. The oxygen gas consumed increased fairly constantly in respect to time. 5. From the slope of the four lines on the graph, determine the rate of oxygen gas consumption of germinating and dry peas during the experiments at room temperature and at 10 degrees Celsius. Recall that rate = _y over _x. Condition Show Calculations Here Rate in mL oxygen gas/minute 0.1

Germinating Peas/10C Germinating Peas/ Room Temperature Dry Peas/10C Dry Peas/Room Temperature

(1.2-0.7)/5

(2.7-2.1)/5

0.12

(-.4-0)/5 (-.3-(-.3))/5

-0.08 0

6. Why is it necessary to correct the readings from the peas with the readings from the beads? The gas changes in the beads were only due to pressure and temperature, and not gas consumption, so the beads act as a control. 7. Explain the effect of germination (versus non-germination) on pea seed respiration. Germinating peas consumed more oxygen than non-germinating. 8. Below is a sample graph of possible data obtained for oxygen consumption by germinating peas up to about 8 degrees Celsius. Draw in predicted results through 45 degree Celsius. Explain your predictions.

The amount of oxygen consumed will steadily increase until the temperature reaches a point at which the enzymes become denatured. 9. What is the purpose of KOH in this experiment? The purpose of the KOH was to remove the effect of carbon dioxide from the readings. 10. Why did the vial have to be completely sealed around the stopper? The vial had to be completely sealed so that gases couldnt escape and water couldnt leak into the respirometer.

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 10 degrees Celsius, what results would you expect? Explain your reasoning. The reptile would use less oxygen because it is cold-blooded and wouldnt be as active at a colder temperature as the mammal would. 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 respiration of the small mammal would be higher at 10 degrees Celsius because it would need more energy to keep its normal body temperature. 13. Explain why water moved into the respirometers pipettes. Water moved into the respirometers pipettes because pressure decreased when the amount of oxygen was decreased. 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? Set up five respirometers containing beads, non-germinating peas, peas that have germinated for one day, peas that have germinated for two days, and peas that have germinated for three days. Measure the water readings in intervals of five minutes for twenty minutes. The peas that have been germinating for three days will have the highest rate of respiration and the beads will have the lowest rate of respiration. 15. According to your graph, what happens to the rate of oxygen consumed by germinating peas over time? What does this indicate to you? The rate of oxygen consumption is fairly constant. 16. How did the KOH affect the water movement in the respirometer? It allows more water into the pipette. 17. Which of the two pea types, germinating or non-germinating, consumes the most oxygen? Why? Germinating peas consume more oxygen because they are growing and are more active than non-germinating peas. 18. What was the effect of temperature on pea respiration? Warmer temperatures allow for the peas to respire at a faster rate. 19. During aerobic respiration, glucose is broken down to form several end products. Which end products contain the carbon atoms from glucose? The hydrogen atoms from glucose? The oxygen atoms from glucose? The energy stored in the glucose molecules? Carbon dioxide contains the carbon, water contains

the hydrogen, both carbon dioxide and water contain the oxygen, and ATP contains the energy. 20. What is fermentation? What are the two types of fermentation? What organisms use fermentation? Fermentation is a catabolic process that makes a limited amount of ATP from glucose without an electron transport chain and that produces an end-product such as ethyl alcohol or lactic acid. The two types of fermentation are alcoholic and lactic acid fermentation. Plants use alcoholic while animals use lactic acid. 21. Draw a Venn diagram showing how respiration and fermentation are similar and how they differ.

22. What are the three pathways involved in the complete breakdown of glucose to carbon dioxide and water? What reaction is needed to join two of these pathways? What are the substrates and products of this reaction and where does it take place? The three pathways are glycolysis, the electron transport chain, and the Krebs Cycle. The reaction of the pyruvate joining with CoA enzyme and NAD to produce acetyl CoA, NADH, and carbon dioxide. The acetyl CoA goes to the Krebs Cycle and NADH to the electron transport chain in the mitochondria. 23. Write the letter of the pathway that best fits each of the following processes. Pathway

a. Glycolysis b. Krebs Cycle c. Electron Transport System Process 1. Carbon dioxide is given off b. 2. Water is formed c. 3. PGAL a. 4. NADH becomes NAD+ c. 5. Oxidative phosphorylation c. 6. Cytochrome carriers c. 7. Pyruvate a. 8. FAD becomes FADH2 b. 24. Calculate the energy yield of glycolysis and cellular respiration per glucose molecule. Distinguish between substrate-level phosphorylation and oxidative phosphorylation. Where does the energy for oxidative phosphorylation come from? 36 ATPs are formed per glucose moelcule. Four of the ATPs are formed from substrate level and 32 from oxidative. Oxidative Substrate Glycolysis Transition Krebs Total 2 0 2 4 4 6 22 32 6 6 24 36 Total

Error Analysis:
Some of the errors that could have occurred during the experiment were water leaking into the respirometers, gas escaping the respirometers, water temperature in the bath changing, and mathematical mistakes.

Discussion and Conclusion:

This lab showed that germinating peas consumed more oxygen at a faster rate than the non-germinating peas and the beads did. The non-germinating peas and the beads didnt consume hardly any oxygen at all. It also showed that the respiration rate of germinating peas was faster than the respiration rate of nongerminating peas. Finally, this experiment showed that respiration rates increase as the temperature increases. This shows that temperature and respiration rates are directly proportional to each other.