By Lee Palmer, Upper School Science Department, The Bryn Mawr School
Abstract
In this laboratory exercise, you will design and conduct an experiment to evaluate the effect of the presence of biodegradable materials on dissolved oxygen levels.
Objectives
After completing this exercise, you will be able to:
 Understand the effect of biodegradable pollutants on water quality.
 Design and conduct an experiment.
 Interpret data.
 Suggest additional studies.
 Do serial dilutions.
Introduction
In aquatic systems, aerobic microorganisms will consume biodegradable material for energy, and in doing so will also take up oxygen from the environment as part of the cellular respiration process. Scientists use dissolved oxygen levels as an indication of contamination by such pollutants as sewage, agricultural runoff, organic industrial effluents, etc.
Pre-lab questions
1) List dissolved organic materials and possible sources that may decrease oxygen levels as a result of decomposition when present in water.
2) List possible organisms that may act as decomposers of dissolved organic material in aquatic systems (see on-line video posted on our web-site if you need help).
Hypothesis:
Which test tube will turn clear first and why?
Materials
 Biodegradable liquid
 Six test - at least 10 ml capacity
 10-ml graduated cylinder
 Test tube rack
 Six 1- ml pipettes
 Yeast
 Methylene Blue (in dropper bottlel)
Safety Measures
Methylene Blue may be harmful if swallowed, inhaled, and/or in contact with skin. It may cause severe eye irritation.

4) Using a graduated cylinder, remove 5 ml of milk from tube #1 and place it in tube #2. Mix the contents well.
5) Remove 5 ml from tube #2 and place it in tube #3. Mix the contents well.
6) Remove 5 ml from tube #3 and place it in tube #4. Mix the contents well.
7) Remove 5 ml from tube #4 and place it in tube #5. Mix the contents well, and then discard 5 ml of the solution.
8) Add 3 drops of methylene blue to each tube. Be careful to hold the dropper bottle upright so that the drops are uniform. Mix each tube well.
9) The contents of each tube should now be as below:
TEST TUBE
#
1
2
3
4
5
6 mL Milk and/or H
2
0
5
5
5
5
5
5
Methylene Blue
3 drops
3 drops
3 drops
3 drops
3 drops
3 drops
% Milk
100%
50%
25%
12.5%
6.25%
0%
Dilution
1:1
1:2
1:4
1:8
1:16
--

10) Noting the time, quickly add 1 ml of yeast mixture to each test tube. Invert each test tube 4 times and place in the test tube rack.
11) Carefully observe each tube and record the time at which the color change from blue to white is complete (no more color change).
12) When the color has changed from blue to white in each test tube, record the exact time at which the change is complete. The surface of each test tube will remain blue.
13) BEFORE YOU CLEAN UP, SHAKE ONE OF THE TEST TUBES THAT HAS TURNED WHITE. Record your observations.
Data
Include your raw data and a graph (using a spreadsheet) of the data in your submitted report.
TEST
TUBE #
4
5
6
1
2
3
MIXING
START TIME
(A)
TIME OF COLOR
CHANGE TO WHITE
(B)
TOTAL TIME FOR
COLOR CHANGE (B-
A)
Analysis Questions
1) What is the gas taken in by the microorganisms?
2) What is the gas given off by the microorganisms?
3) Where do microorganisms living in water get the oxygen that they use in decomposition?
4) Where do the green plants living in water get the carbon dioxide that they need?
5) What happened in the tube you shook after it had turned white (step 13)? Why?
6) Why is the oxygen in this experiment used up?
7) Which part of your experiment represents the decomposers?
8) Which part of your experiment represents the sewage dumped into the water?
9) In which test tube did you have the most "sewage"?
10) Graph your results using Excel. What are the labels for your axes?
11) What does this graph tell you about the relationship between the biodegradable waste in water and the amount of dissolved oxygen in the water?
12) What would be the effects of dumping a great deal of raw sewage into a river as regard to the dissolved oxygen in the same river?

Conclusion:




Was the hypothesis supported or refuted? Why or why not?
What were the sources of error in this experiment?
What might be done differently to improve the quality of the data?
Why eutrophication is important to study