Anaerobic respiration (Fermentation) Lab (35 points)
N. Kolar
Yeast are classified as facultative anaerobes, which means they are capable of
both aerobic and anaerobic respiration. When oxygen is unavailable, yeast carry out
fermentation, a type of anaerobic respiration (without oxygen). The difference between
aerobic and anaerobic respiration lies in how the NADH + H+ produced in glycolysis is
converted back to NAD+. In aerobic respiration the hydrogens (electrons) from NADH
+H+ are passed to oxygen in the electron transport chain yielding 3 ATPs per NADH +
H+ while in fermentation the hydrogens (electrons) are passed on to acetaldehyde to form
ethanol yielding no ATPs per NADH + H+ as follows:
Step 1 - Pyruvate (from glycolysis) → Acetaldehyde + CO2
Step 2 - Acetaldehyde Ethanol & NADH + H+ NAD+
Humans have made use of the byproducts of fermentation for centuries: CO2
to make bread rise and ethanol in beer and wine making. From the yeast's “perspective”
both CO2 and ethanol are waste products and in fact ethanol is toxic, killing the yeast
organisms when it reaches a concentration between 14-18%. This is why the percentage
of alcohol in wine and beer doesn't exceed approximately 16%. In order to produce
beverages with higher concentrations of alcohol (liquors), the fermented products must
be distilled.
Fermentation produces only 2 ATPs per glucose molecule (via glycolysis).
Aerobic respiration produces 36 - 38 ATPs from a molecule of glucose. Thus the ability
of yeast to live in the absence of oxygen comes at a price - fermentation produces 19-fold
fewer ATPs per glucose molecule than does aerobic respiration.
Purpose - Part 1
* In this exercise the effect of the following compounds on fermentation rate in
bakers’ yeast (Saccharomyces cerevisiae) as measured by CO2 production
will be observed.
Make a hypothesis: (put on written lab sheet)
Procedure:
* Using fermentation tube……BE EXTREMELY CAREFUL NOT TO
BREAK!!!
1. Label your fermentation tube (initials and class period) with colored tape.
2. Fill the fermentation chamber with water to find out how many ml of water your
fermentation chamber will hold…..should be approximately 25 ml (record the exact
amount). Then pour the water out.
3. Gently pour 12-13 ml of sugar solution (2% sucrose) into the chamber and then
add 12-13 ml of the 2% Baker’s yeast solution.
4. Gently mix the contents of the chamber by placing your thumb on the open end of
the fermentation chamber and slowly invert the chamber a couple of times. NO
BUBBLES should be within the chamber!
4. Make certain that you tip the fermentation tube when you fill it so the blind
end (end that is closed) is completely filled with the sugar/yeast solution.
5. Insert a foam plug into the outlet of each fermentation chamber.
6. Put your labeled fermentation chamber into the class tray on the back lab table.
7. I will incubate all fermentation chambers at 29ºC for increments of 10 minutes
of time. (make your data table and answer questions to your lab while waiting).
8. Check the fermentation chambers and record the amount of accumulated CO2
(mm) every 10 minutes for 60 minutes. Measure the height in millimeters on
your data table in the column of accumulated CO2 (mm).
9. I will remove the box of fermentation chambers from the incubator to take
measurements.
10. Place your fermentation chamber on a flat surface and gently swirl the contents
to release any gas trapped in the base by the settling yeast organisms. Then
measure the amount of CO2 produced; measure from the bottom of the bubbles in
the blind end of the tube to the top of the blind end. To read the measurements it
might be helpful to place an index card on the top of the blind end of the
fermentation tube making it parallel to the table top so it hits the ruler. This will
give you an accurate reading rather than just ‘eye balling it’. See illustration
below:
Clean up:
• At the end of the experiment very carefully and thoroughly rinse out your
fermentation tubes. Do not use soap.
• Rinse out and save the foam plugs.
Laboratory Report:
• Write up this exercise as a laboratory report.
• This would of course include graphing of your data (included a data table of time vs.
CO2 (ml) produced) and the evaluation of your to determine rate of yeast respiration for
this particular substrate. This is important information for you to keep because we will
be using this information as a comparison to the next lab that you create using various
treatments on yeast for CO2 production.
Hypothesis
Write your hypothesis in the space below. Remember that a hypothesis is a testable
model/explanation based on your prior knowledge about the experiment topic (2 pts)
Prediction
Write your prediction of the outcome of the experiment in the space below. (2 pts)
Your prediction will be what you predict will be the outcome of your experiment.
Data (10 pts)
Using 2% Sucrose/Yeast solution Air space of CO2 produced After Time allotted:
10 min
20 min
ml
ml
30 min
40 min
ml
ml
50 min
ml
60 min
ml
Questions
Answer in the space provided.
1. Describe what is sucrose is; also write chemical formula? (2 pts)
2. What was the independent variable in your experiment? (1 pt)
24 hrs.
ml
48 hrs.
ml
3. What was the dependent variable in your experiment? (1 pt)
4. What would be a control in this experiment? Describe one. (2pt)
5. What was the specific source of energy in the fermentation chamber that the yeast
used for respiration? (1 pt)
6. How would changes in the incubation temperature influence this investigation?
(Explain ‘why’ in terms of your knowledge of reaction rates and respiratory
enzyme function.) (3 pts)
7. Did the yeast produce different amounts of carbon dioxide over same intervals of
time? If so, why do you think this happened? (3 pts)
8. Was your prediction correct?
Did the data support your hypothesis?
Explain.
Would you revise your hypothesis?
If so, why? (3 pts)
9. Was there any error in this investigation? If so, list and explain at least two major
sources of error. If not, write down areas which could have caused error to the
experiment. (2 pts)
10. If you were to continue your investigation of respiration of yeast, what would you
test next? Give a brief description of the experiment you would do. (3pts)