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Samira Sackietey A.

P Biology Lab # 5

Cellular Respiration

Introduction: In this lab we studied the process of cellular respiration in which is an ATP producing catabolic process which involves a number of enzymes-mediated reactions. It first begins with Glycolysis, which takes place in the cytosol of the cell and glucose is split into 2 three-carbon molecules called pyruvate and in the process producing ATP. The rest of respiration takes place in the mitochondria of the cell and these processes are known as the electron transport chai and The Krebs cycle. The general equation used for respiration is: C6H12O6 + 6O2 -> 6CO2 + 6H2O + 686 kilocalories of energy per mole of glucose oxidized. There are three ways to measure respiration which are to see how much oxygen is consumed, the production of carbon dioxide, and the release of energy. So in this lab we measured the volume of oxygen consumption by germinating and non-germinating peas at two different temperatures. PV=nRT is the inert gas law. 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. T is the temperature of the gas in degrees K. This law tells us several important things 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. 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 are kept constant, then either pressure or volume or both will change in direct proportion to the temperature. So in this lab the carbon dioxide that is released will be removed by potassium peroxide and turn it to potassium carbonate. We remove carbon dioxide so that the change in the volume of gas in the respirometer will be directly proportional to the amount of oxygen that is consumed. So since this occurs water will move toward the region of lower pressure. During respiration, oxygen will be consumed and its volume will be reduced to a solid. The result is a decrease in gas volume within the tube, and a related decrease in pressure in the tube. The respirometer with just the glass beads will allow changes in volume due to changes in atmospheric pressure or temperature changes. So we should expect the respirometer with only germinating peas will have a larger consumption of oxygen and will have a larger amount

of CO2 that is converted into potassium carbonate than the respirometer with beads and dry peas and the respirometer with beads alone. Materials: Germinating Peas Tap Water Timer Non-germinating (dry) Peas Ice 5-mL Pipette Glass Beads 100 mL Graduated Cylinder 6 Vials 2 Thermometers Absorbent and Non- absorbent Cotton 6 Rubber Stopper 2 Baths KOH Masking Tape Weights

Procedure: 1. The two temperatures for the water baths should be room temperature and 10 degrees (C) below room temperature (25C). Use ice to lower the temperature of the second set of water baths. Be sure to record these temperatures in the titles for your data tables. 2. Another lab group will set up the same 3 respirometers as replicates at the same temp, other lab groups will do the same at the second temperature then the data will be pooled so class means for all the respirometers with the seeds can be graphed (the data from the respirometers containing only glass beads are to be used for volume adjustments only). 3. To assemble the respirometers obtain 3 vials, each with stopper and attached pipette be careful not to dislodge the pipette or break the seal between the pipette and the stopper since any leaks will result in data inaccuracies. Make sure your vials are dry on the inside. Place a small piece of absorbent cotton in the bottom of each vial (take care to use the same amount in all 3 vials). Taking care not to get any KOH on the sides of the vial, moisten the

cotton with 3 drops of 15% KOH using a thin stem pipet. Be sure to replace the lid on your KOH so it doesnt absorb CO2 from the air. Now add an equally small piece of nonabsorbent cotton (to protect your seed from the KOH) 4. Place the germinating seeds, dry seeds, and beads in vials 1, 2 and 3, respectively as pictured in insert the stopper fitted with the calibrated pipette. 5. Make a sling of masking tape attached to each side of the water bath to hold the pipettes out of the water during an equilibration period of 7 minutes. Place the respirometers in the water bath; use a weight at the junction of the pipettes and stoppers to hold the respirometers under water.

6. AFTER an equilibration period of seven minutes, remove the masking tape and lower the tips of the respirometers into the water bath take care NOT to lift the vials out of the water you dont want to change the temperature of the respirometers when lowering them. Make sure respirometers are fully submersed in the water baths. Water will enter the pipettes for a short distance and then should appear to stop if water continues to move in check for leaks in the respirometer. Work swiftly and arrange respirometers so that the pipettes can be read through the water. Keep your hands out of the water after the experiment has begun and strive to maintain a constant temperature in the water. 7. Allow the respirometers to equilibrate for three more minutes and then record, to the nearest 0.01mL, the initial position of water in each pipette (= time 0 value). Check the temperature and record it.

Observations: (Data) Table 5.1: (Group Data) Measurement of O Consumption by soaked and dry seed at room Temperature (25C) Table 5.1 10 Celsius Time (min) 0 5 10 15 20 Individual Data Germinating Peas Reading Diff. * Corrected at time X Diff. .85 ----.60 .25 .25 .39 .46 .46 .25 .60 .60 .09 .76 .76

Beads Alone Reading Diff.* at time X .84 --.84 0 .84 0 .84 0 .84 0

Dry Peas and Beads Reading Diff.* Corrected at time X Diff. .92 ----.90 .02 .02 .88 .04 .04 .86 .06 .06 .85 .08 .08

Class Average Data: 10 degrees Time 5 10 15 20 25 degrees Time 5 10 15 20 Beads alone .09 .015 .015 .02 Beads Alone 0 0 0 0 Germinating Peas .059 .115 .18 .25 Germinating .22 .44 .62 .81 Dry Peas and Beads .0225 .05 .06 .0325 Peas and Beads .07 .35 .45 .06

Graphs: Group Data:

The Effect of the Rate of Oxygen Consumemed by different seeds at 25C on the Time of Respiration
Oxygen Consumption in mL 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 5 10 Time (min) 15 20 Beads Alone Germinating Peas Dry Peas and Beads

Class Data:

The Effect of the Rate of Respiration on the Time Respiration

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 5 10 Time (min) 15 20 Oxygen Consumed (mL)

Beads Alone 10 C Germinating Peas 10C Dry Peas and Beads 10C Beads Alone 25C Germinating Pease 25C Dry Peas and Beads 25C

Results: In the end of our experiment we found see that the rate of respiration for the germinating peas was indeed higher than that of the rate for the dry peas. This was due to the fact that the germinating peas were doing a much rapid rate of cellular respiration and the dry peas were doing a much less rate because the dry peas require a much less energy so they carried slower process of respiration. This therefore allowed less oxygen to be consumed. Error Analysis: There were many factors that may have contributed to error in are results and data such being that the temperature of the water was not maintained at a constant. Also, moving the vials can cause inaccuracies, not having the same amount of cotton in the vials, allowing the KOH to come into come into contact with the peas, vials may have also been improperly sealed, and the equilibrium period may have varied also distorting results. Conclusion Questions: 1. In this activity you are investigating both the effect of germination versus non-germination and warm temperature versus cold temperature on the respiration rate. Identify two hypotheses being tested in this activity. The two hypotheses being tested are that the rate of respiration occurs at a faster rate for respirating peas than it does for non-germinating peas and whether the temperature has a direct effect on the rate of respiration 2. This activity uses a number of controls. What conditions must stay constant? Why? Some of the controls that must stay constant are the temperature because that will yield stable results. Also, the same amount of KOH in each vial will allow for an equal chance for cellular respiration. Other controls include, identical equilibrium periods for each respirometer, precise time intervals, and the respirometer filled with beads especially because no respiration is taking place therefore it can be used to detect and correct any in accuracies during the experiment. 3. Graph the results from the corrected difference column for the germinating peas and the dry peas at both room temperature and at 10C.

The Effect of the Rate of Respiration on the Time

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 5 10 Time (min) 15 20 Oxygen Consumed (mL)

Beads Alone 10 C Germinating Peas 10C Dry Peas and Beads 10C Beads Alone 25C Germinating Pease 25C Dry Peas and Beads 25C

4. Describe and explain the relationship between the amount of O consumed and time. The amount of O consumed increased as time went on.

5. Condition Germinating Peas/10C Germinating Peas/ Room Temperature Dry Peas/10 C Dry Peas/ Room Temperature Show Calculations Here (.115-.18)/5 (.85-.60)/5 (.225-.05)/5 (.92-.90)/5 Rate (mL O/minute) .013 mL O/min . 048 mL O/min .035 mL O/min .004 mL O/min

6. Why is it necessary to correct readings from the peas with the readings from the beads? Its necessary to correct the readings because the corrected difference takes away any random movement taking place that may be due to air or molecules. 7. Explain the effect of germination (versus non-germination) on pea seed respiration. The germinating peas have a high metabolic rate therefore they consume more oxygen for growth and the non-germinating peas were although alive were also dormant so they needed less oxygen to survive.

8. Graph 5.2:

3 mL O Consumed 2 1 0 0 10 20 30 40 50

Temperature(C) Once the temperature reaches a certain point the enzymes for cellular respiration will denature inhibiting respiration. 9. What is the Purpose of KOH in this experiment? The purpose of using KOH in this experiment is that the CO produced by respiration will be removed by KOH which when both CO and KOH come in contact with one another will turn CO into potassium carbonate (Solid). Since this occurs then the change in volume of gas in the respirometer will relate directly to the amount of O consumed throughout the entire experiment. 10. If you used the same experimental design to compare the rates of respiration of a 25g reptile and a 25g mammal at 10C, what results would you expect? Explain your reasoning. We would expect that the rate of respiration would be higher in the mammal since its warm-blooded and has a constant temperature while, the reptile is cold-blooded and will remain at 10C. 11. If the respiration in a small mammal were studied at both room temperature (21C) and 10C, what results would you predict? Explain your reasoning.

I predict that the rate of respiration would be higher at 21C because its necessary for the mammal to sustain a higher temperature. The rate of respiration will decrease at 10C since the mamma must retain its body heat a lower temperature. 12. Explain why water moved into the respirometers pipettes. Water moved into the pipettes because, while the peas underwent respiration they consumed oxygen and carbon dioxide was released which reacted with KOH decreasing the amount of gas in the vial which allowed for water to partially enter the vial.