Cellular Respiration

Cellular Respiration Lab

Purpose | Hypothesis | Predictions | Materials | Procedure  Observations | Analysis | Conclusions | Error Analysis

Purpose:

To observe aerobic and anaerobic respiration of yeast in an enclosed, fluid environment, and in that way learn about cellular respiration as a whole.

Hypothesis:

The yeast, a facultative anaerobe, will first respire aerobically, but soon there will be no O2 left in the enclosed flask, so the yeast will respire anaerobically.  ATP will be made via substrate-level phophorylation during glycolysis, which requires no O2.  To regenerate the NAD+ needed for glycolysis, the yeast will undergo alchoholic fermentation, which produces CO2 .  An added amount of gas inside the flask will cause the balloon to grow.

Predictions:

We believe that the yeast will soon begin to respire anaerobically through alcoholic respiration, releasing CO2 but taking in no O2.  Therefore, the balloon, filled with the CO2 that is released as a by-product of alcoholic respiration, will inflate, and possibly pop.

Materials:

  • 50 mL Erlenmeyer Flask
  • teaspoon yeast
  • 40 mL apple cider
  • Latex balloon
  • Spoon

Procedure:

  • Place teaspoon of yeast in Erlenmeyer Flask, by filling a spoon to the point where the yeast is about level with the top of the spoon.
  • Add 40 mL of apple cider to the 50 mL Erlenmeyer Flask.
  • Cover the flask tightly with a balloon
  • Observe and make predictions
  • Let sit for 24 hours, then make more observations.

Observations:  

When we had added the apple cider and shaken the flask, the yeast went to the bottom of the flask, making a light beige band at the bottom of the cider.  The sediment clouded the cider and made it murky, and tiny bubbles began to rise to the top of the cider.  Some granules of yeast remained stuck to the flask above the level of the cider.  Most of the yeast was still at the bottom after a couple of minutes, but a level had formed at the top also, creating a floating light band as well.

Day 2:  The liquid was even more opaque and tan in color, and most of the yeast had settled at the bottom as a lighter powder.  There were still bubbles of gas rising from the concentration of yeast to the top of the cider.  There was a layer of yeast that was attached to the side of the flask above the cider level that had not been apparent yesterday.  This layer appeared to be thick, wet, and covered almost all free space on the side of the flask that was above the cider level.  The balloon was standing on its own will and was full of air, but was not fully inflated.  At its widest, the balloon was 15 cm in circumference.

Day 3:   The balloon and flask appear to be almost identical to their look during the second day, with the balloon of approximately the same size and yeast still concentrated on the bottom and on the inside of the flask above the level of the cider.  The only difference was that there were no longer bubbles rising to the top of the container.  The balloon had actually grown slightly, its circumference now 16.2 centimeters.

Day 4:  There were no differences in the flask, but much of the air had diffused from inside the balloon.  It was still inflated and standing upright but was not as thickly filled.

 

Circumference of Ballon (cm)

Original

0

After 1 day

15

After 2 days

16.2

After 3 days

Less

Analysis: 

This chart and the graph shows the change in the daily circumference of the balloon.  There was a very sharp rise from the uninflated balloon of the first day and the 15 cm circumference of the second day.  A small rise, 8%, occurred between the first day and the second day, when the balloon had a transverse circumference of 16.2 cm. There was a 20% drop between the second day and the third day (13 cm.).

 

Circumference of Ballon (cm)

Original

0

After 1 day

15

After 2 days

16.2

After 3 days

Less

 

Conclusions: 

Our predictions that the balloon would inflate because the yeast would participate in cellular respiration were supported by our results.  Yeast is a facultative anaerobe, meaning that it can participate in aerobic respiration when possible, but when this is impossible, it respires anaerobically.  When we closed the balloon by attaching it to the flask, stopping the exchange of gases from the external environment, there was some O2 left in the flask.  The yeast received its O2 and the glucose needed for cellular respiration from a supply in the apple cider. The yeast respired aerobically until the supply of O2 was used up.  This began between the first and second days, shown by the fact that the balloon increased in size between these two days.  During aerobic respiration, the same amount of gas that is used in the form of O 2 (as the final electron acceptor of the electron transport chain) is produced in the form of CO2 (a by-product of the reactions of the Krebs cycle), so anaerobic respiration was required for the balloon to begin to fill.  During this time, the yeast grew, shown by its pastiness as it spread out in the flask.  When it respired anaerobically, it proceeded through the Kreb Cycle and chemiosmosis as well as glycolysis.  This was more efficient, producing about 38 ATP per glucose molecule consumed.  However, there was soon no remaining O2 in the flask, so the yeast had to turn to anaerobic respiration, or fermentation, to make ATP.  Because the balloon inflated and we saw bubbles rising to the top of the cider, it is clear that the yeast participated in alcoholic fermentation, because it is the only type of fermentation that produces CO2 as a by-product.  This conclusion was also supported by the fact that when we emptied the flask, almost two weeks later, it smelled very alcoholic.  The bubbles rising to the top of the cider were gas bubbles of CO2 escaping from the yeast and rising to the air.  In alcoholic fermentation, glycolysis, the first of three steps in cellular respiration, occurs as normal.  Glycolysis is a nine step process, with carbohydrate intermediates at each step, that breaks a glucose molecule into two molecules of pyruvic acid, and produces two molecules of ATP for use by the cell via substrate level phosphorylation and two molecules of NADH, a high energy electron carrier.  Normally, NADH would drop its two electrons and H+ ion at the electron transport chain, but this process of chemiosmosis does not occur during fermentation.  Chemiosmosis requires O2 as the final electron carrier in the electron transport chain.  The O2 is broken in half and is reduced with 2 electrons and 2 H+ ions to form H2O, water.  Without O2, yeast will not proceed to chemiosmosis or even the Kreb Cycle, a cycle that produces even more high-energy electron carriers.  Instead, it converts the pyruvic acid and 2 NADH that have been produced to 2 molecules of ethanol, a two carbon compound.  In this way, NADH is oxidized to NAD+ and is able to be reused.  Two molecules of CO2, one for each ethanol that is made, are released as by-products.  In our experiment, the yeast produced CO2 gas but was intaking no gas, so the level of gas in the sealed compartment was growing.  This higher level of gas caused the balloon to inflate.  The cider became alcoholic and began to kill the yeast, so all of the yeast had died after the third day. Most of the yeast had been poisoned and died after the second day, shown by the fact that the circumference of the balloon grew very little between the second and third days.  The little amount of yeast that was left slightly added to the level of gas in the balloon in the second day, but by the third day, all of the yeast was dead.  The pressure of the latex balloon pushing inwards caused air to diffuse outwards through small holes that were not visible to us.  This caused the balloon to shrink after the third day.  The process that we used in the apple cider is similar to a process used to make wine, only that one way gas valves export the buildup of CO2 caused by the yeast.

 This experiment taught us several things about respiration and fermentation.  We were able to observe first aerobic and then anaerobic respiration of yeast, and we saw how an export but lack of import of gases in anaerobic environments can add gas to an enclosed environment.  On a small scale, we were able to see how wine and beer are made, and for the first time, we were able to use knowledge gained from the reading to made completely accurate predictions.

Error Analysis: 

In a fairly simple experiment such as this one, we seemed to have made very few errors.  We may have been less than perfect in our measurements of circumference and mL, and we know that we were less than exact about our measurements of teaspoons of yeast, but this did not seem to drastically change our results.  Contamination with other substances might have affected the metabolism of the yeast and therefore affected this experiment as well.  Another problem was the pores in the balloon.  We know that between the third and forth day, air escaped through these pores, but it is also possible that air might have entered, and added O2 might have slowed the beginning of fermentation. We also should have taken more measurements of the circumference of the balloon, perhaps measuring the longitudinal circumference and definitely measuring the transverse circumference in the first day.  If possible, we should have measured the alcohol level of the cider to see if the yeast truly engaged in alcoholic fermentation as we believe.  We also should have measured the thickness of the yeast layer to see whether or not the cellular respiration caused the yeast to grow.  Also possible mistakes involved mishandling of the flasks, but overall we were able to make successful predictions in an experiment that contained very few errors.

 

 

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