AP
Lab 12
Dissolved
Oxygen and Aquatic Primary Productivity
Objectives-
Before
doing this laboratory exercise, you should understand:
-the biological
importance of carbon and oxygen cycling in ecosystems
-how primary productivity
relates to the metabolism of organisms in an ecosystem
-the physical and
biological factors that affect the concentration of dissolved oxygen in aquatic
ecosystems
-the relationship between
dissolved oxygen and the processes of photosynthesis
and respiration as they affect primary productivity
After
completing the laboratory exercise, you should be able to:
-measure primary productivity based on changes
in dissolved oxygen in a controlled
experiment .
-investigate the effects of changing light
intensity on primary productivity in a controlled experiment
Introduction
Oxygen is critical for the life processes of
nearly all organisms. In the aquatic
environment, oxygen must be in solution in a
Oxygen enters water via diffusion from the air
and photosynthesis by Aquatic plants. Physical factors such as the temperature
and salinity of the water and the partial pressure of oxygen in the air
influence the rate at which oxygen enters the water. In the absence of mixing
by winds, currents, tides or other flows, the only way that oxygen is distributed
through water is by diffusion. At 20° C, oxygen diffuses 300,000 times more
slowly in water than in air. In contrast to the relatively uniform distribution
of oxygen in air, spatial distribution of oxygen in water can be highly
variable. Oxygen can be consumed at lower regions of an aquatic environment
faster than it can be replaced from the surface, resulting in gradients of
oxygen concentration and/or anaerobic conditions in some parts of a body of
water.
Biological processes such as photosynthesis and
respiration can also significantly affect dissolved oxygen concentrations.
Photosynthesis usually increases the oxygen concentration in water. Respiration
requires oxygen and will usually decrease dissolved oxygen concentration.
Respiration by microorganisms can be particularly influential in bodies of
water, because populations of microbes can increase quickly. Warm temperatures
usually accelerate microbial growth, increasing the demand on dissolved oxygen.
Water
Pol1ution
Water pollutants can decrease dissolved oxygen
concentration, usually and Dissolved by stimulating microbial growth and
thereby increasing the demand Oxygen for oxygen for microbial respiration. Many
organic pollutants, such as sewage, can be directly metabolized by microbes in
oxygen-requiring processes. Therefore a large influx of sewage stimulates
growth of microbes that metabolize it, with a subsequent decrease in dissolved oxygen.
Decomposition of dead organisms is also carried
out by microbes through oxygen requiring processes. Any pollution event that
kills large numbers of organisms(such as spills of herbicides or pesticides) results
in proliferation of decomposers and the use of oxygen for decomposition.
Although the effect is not direct, fertilizer pollution can diminish dissolved
oxygen in the same way. The pollution first stimulates over proliferation of
aquatic plants, which may first produce additional oxygen, but eventually the
plants will die and require decomposition. Fish kills as -the result of
fertilizer pollution are often the result of oxygen starvation that occurs as
large masses of dead plant material are decomposed.
Exercise
12-A
In this exercise, you will determine the effect
of temperature on dissolved oxygen. You will start with a single water sample,
divide it into three portions, and let each portion equilibrate at a different temperature.
Then you will measure dissolved oxygen. Water samples are not usually
saturated; the amount of oxygen dissolved in a water sample is often only part
of what the water could hold. You will use a graphical device called a
nomograph to determine what percent of saturation is represented by the levels
of dissolved oxygen you measure in the different temperature samples.
Primary
Productivity
The fertility of any body of natural water
depends on the productivity of the green plants within it. The primary
productivity of an ecosystem is defined as the rate at which sunlight is stored
by plants in the form of organic materials (carbon-containing compounds). Aquatic
green plants use carbon from the carbon dioxide dissolved in the water for
carbohydrate synthesis according to the basic equation for photosynthesis:
light
6 CO2 + 6 H2O ---------------------> C6H12O6+
6 O2
Chlorophyll
Primary productivity can be determined by
measuring the rate of carbon dioxide utilization, the rate of formation of
organic compounds, or the rate of oxygen production. In this exercise you will determine
productivity by following oxygen production. For each milliliter of oxygen
produced approximately 0.536 grams of carbon has been assimilated.
Determining productivity from oxygen production
data can be complicated by the fact that
plants both produce and use oxygen. Plants produce oxygen and glucose through
photosynthesis, which requires light. Plants use the glucose they manufacture
as an energy source through respiration just as animal cells do. Like animal respiration,
plant respiration requires oxygen.
Plants respire constantly, but photosynthesize
only when light is available. The balance of the two processes, respiration and
photosynthesis, determines whether the plant is a net consumer or producer of
oxygen and indicates the net productivity.
Exercise
12-B
In this laboratory exercise, you will be
measuring oxygen production by the alga Chlorella under different light conditions,
using the light and dark bottle method. In this method, the dissolved oxygen concentrations
of samples of ocean, lake, or river water or of laboratory algae cultures are
measured and compared after incubation in light and darkness. You will expose
samples to a range of light intensities, from full light to 2% of full light,
and measure the changes in dissolved oxygen concentration after overnight
incubation.
Total oxygen production in a sample bottle is the
sum of any increase in total oxygen plus the amount of oxygen consumed by
respiration during the incubation period. In the bottles kept in darkness, the change
in dissolved oxygen (DO) concentration from the initial concentration is a
measure of respiration, since photosynthesis cannot occur in the absence of
light. In the bottles exposed to light, both photosynthesis and respiration
occur; therefore, the change in DO concentration in these samples is a measure
of net productivity. The difference between the final DO concentrations in the
light bottle and the dark bottle is the total oxygen productivity and therefore
an estimate of gross productivity.
Exercise
12-A
The
Measurement of Dissolved Oxygen
Overview
The Winkler method is commonly used to measure
DO. In this method, a series of solutions is added to the water sample which
react with the dissolved oxygen in the sample 'to release free iodine. The
quantity of free iodine released is proportional to the amount of free
dissolved oxygen in the original sample. The amount of free iodine in the
sample can be determined by adding a starch indicator solution to the sample, which
turns blue in the presence of free iodine, then titrating with sodium
thiosulfate until a colorless endpoint is reached. The amount of sodium
thiosulfate needed to titrate the iodine is directly proportional to the
concentration of dissolved oxygen in the original sample.
Procedure
1. Label 3 BOD bottles, one 4° C, one 25°
C, and one 30° C.
2. Fill the BOD bottles. The most important
aspects to this process are: 1) not to trap any air in the bottle and 2) to
avoid introducing any turbulence, since turbulence will mix air into the
samples and increase the dissolved oxygen levels.
There are several
different ways to fill the bottles:
a) submerge the bottle in the sample, let
it fill, then cap it while it is still submerged
b) pour the sample very gently from a
beaker, creating as little turbulence as possible
c) use the 60 mL syringe (take special care
since jtis easy to introduce air by pushing the sample out fast)
d) use the 60 mL syringe with a piece of
tubing attached. This works well if the sample container has a narrow mouth or
is very deep.
If you use a beaker or syringe, tip the BOD
bottle as you fill it and let the sample run gently down the side. If you are
using tubing, place the end of the
tubing at the bottom of an upright BOD bottle and introduce sample gently. Fill
the bottle until it overflows significantly. This is to ensure there is no air
trapped in the bottle to give elevated oxygen readings. Cap bottles tightly
after filling.
3. After filling the three bottles, fix the
oxygen in each by the following procedure:
a) Uncap each bottle.
b) Add 8 drops of manganous sulfate
solution to each bottle.
c) Add 8 drops of alkaline potassium iodide
azide to each bottle.
d) Cap bottles and mix. A precipitate will
form. Allow the precipitate to settle to the shoulder of the bottle before proceeding.
e) Use spoon to add 1 gram (1 spoon) of sulfamic
acid powder to each bottle.
f) Cap and mix until reagent and
precipitate dissolve.
The samples are now fixed. This is an optional
stopping point. Samples can be stored at room temperature until convenient to
continue.
4. After fixing the oxygen in each bottle in step three,
determine the amount of dissolved oxygen in each one by the following method:
a) uncap the 4° C bottle and fill titration
tube to 20 mL line. Care should be
taken to be as precise as possible. Variations in filling from group to group
and from bottle to bottle will result in inconsistent data.
b) Fill titrator to the 0 line with sodium
thiosulfate. Read the volume across the concave edge of the plunger.
c) Add one drop at a time to sample,
swirling between each additional drop until the sample is a faint yellow color.
d) Remove the titration syringe and cap
without disturbing the syringe; Add 8 drops of starch indicator solution.
e) Replace the lid of the titration tube
and swirl the sample. The solution should turn blue. If the solution does not
turn blue, there is not a measurable amount of oxygen present, or too much
sodium thiosulfate was added. If so, pour out the sample, refill the titration
tube from the BOD bottle, and start the titration again (step b).
f) Continue the titration with the sodium
thiosulfate already in the syringe. Add one drop at a time, swirling the sample
after the addition of each drop, until the blue color disappears. If the blue
color does not disappear after the addition of the whole syringe of sodium
thiosulfate, refill and continue. When the titration is complete, add 10 ppm
from the first syringe to the amount added from the second syringe.
g) Read the syringe at the bottom of the
plunger. The number represents ppm DO which is equal to mg oxygen per liter of
water. Record data in Table 12.1
5. Repeat steps a-e above for the 25° C
sample and the 30° C sample.
6. Using the nomograph of oxygen
saturation, estimate the percent saturation of dissolved oxygen in your samples
and record this value in the table below; line up the edge of a ruler with the temperature
of the water on the top scale and the dissolved oxygen on the bottom scale,
then read the percent saturation from the middle scale. Record the data in
Table 12.1.
Questions
to Answer
1. How does temperature affect solubility
of oxygen in water?
2. If you were to design an experiment to
determine how salinity affects the solubility of oxygen in water, what would
the controls be and what would the variable be?
Exercise
12-B
The
Measurement of Primary Productivity
Overview
The productivity per square meter of a water
column within an aquatic ecosystem can be measured by the Light-Dark Bottle
Winkler method. When using this method, temperature should be held constant so
that only one variable is being tested. Each student group will measure initial
sample, a dark sample, a plain light sample, and four samples , wrapped with
different numbers of plastic screens. The screens filter the light available in
the bottle, simulating the effect of increasing depth in a pond.
Number of Screens Percent Light
0 100%
1 65%
3 25%
5 10%
8 2%
Procedure
Day
One
1. Work in groups of up to 5 students
depending on class size. Obtain seven BOD bottles and rinse them well.
2. label the bottles Initial, Dark, 0, 1,
3, 5, and 8.
3. Fill the bottles with pond water mixure
as described in Exercise 12-A.
4. Wrap the Dark bottle with aluminum foil.
5. Bottle 0 should not be wrapped. This
will be your full light bottle.
6. Wrap bottles 1, 3, 5, and 8 with the
corresponding number of screens.
7. Fix the Initial bottle as in Exercise
12A steps 1-3
a) The sample is now fixed. NOTE: If your
sample contains a heavy algae load, the algae will form a black precipitate
which will not go into solution. This will not affect your results.
8. Lay the remaining bottles on their sides
under a fluorescent or grow light, seam side down, and leave overnight.
Day Two
1. Obtain bottles from light source.
2. Fix the samples by following steps as in
Exercise 12A steps 3 a-f. After fixing
each bottle determine the amount of dissolved oxygen by following the steps 4
a-g.
3. Enter the amount of dissolved oxygen
(DO) for each bottle in Table 12.2. Multiply the dissolved oxygen concentration
in mg/L by 0.698 to convert it to mL/L, the units most commonly used to report dissolved
oxygen levels.
Table 12.2
|
Bottle |
DO (mg/L) |
Multiplier |
DO (mL/L) |
|
Initial (experimental control) |
|
0.698 |
|
|
Dark |
|
0.698 |
|
|
Light Bottle |
|
0.698 |
|
|
1 screens |
|
0.698 |
|
|
3 screens |
|
0.698 |
|
|
5 screens |
|
0.698 |
|
|
7 screens |
|
0.698 |
|
5. Calculate the respiration rate by the
following equation:
Respiration
= [Initial (mL O2/L)
-Dark (mL O2/L)]
time
in hours
Respiration
rate = __________________
6. Calculate the net productivities of the
remaining cultures by the following equation. Enter the data in Table 12.3.
Net Productivity = [Light(mL O2/L)
-Initial(mL O2/L)]
time
in hours
7. Assume the algae in each bottle consumed
the same amount of oxygen through respiration as did the algae in the dark bottle.
Each culture of algae therefore had a gross oxygen production equal to the net
oxygen production plus the amount consumed in respiration:
Gross
Productivity = [(Light -Initial) +
(Initial -Dark)]
time
in hours
=
[Light Bottle -Dark Bottle]
hours
8. Calculate the gross productivity of each bottle and enter it
in Table 12.3. Use mL O2/L
as units for dissolved oxygen and hours for time.
Table 12.3
|
Bottle |
Net productivity |
Gross productivity |
|
No screens |
|
|
|
1 screens |
|
|
|
3 screens |
|
|
|
5 screens |
|
|
|
7 screens |
|
|
9. Calculate class averages for the
different light intensities and calculate average gross and net productivities.
Record these data in Table 12.4.
Table 12.4 CLASS AVERAGE GROSS AND NET OPRODUCTIVITIES
Initial Sample Average DO
= ____________________________
Dark Sample Average DO = ____________________________
Average Respiration rate
= _____________________________
|
Bottle |
Ave DO (mg/L) |
Ave Net |
Ave Gross |
|
No screens |
|
|
|
|
1 screens |
|
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|
|
3 screens |
|
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|
|
5 screens |
|
|
|
|
7 screens |
|
|
|
10. Was growth in any of the sample bottle
limited by the availability of light? ____________
Use data collected from the lab to justify your answer.
11. Pot the average gross and net
productivities (mL O2/L)/hr as a function of light intensity (%).
Which is the independent variable?
Which is the dependent
variable?
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