What is the Effect of Pollutants on

Aquatic Life?
The purpose of this experiment was to discover how pollutants of varying concentrations harm aquatic life.
I became interested in this topic when looking through a list of ideas. I thought it would be interesting to
see what pollutants were the most harmful to aquatic environments. I am also concerned about pollution of
our water because I enjoy many water sports including skiing and swimming.
The information gained from this experiment will help biologists, fish and wildlife agencies, and the public
learn how pollution harms the environment. This information will help determine which pollutants cause the
most damage to aquatic life and need to be cleaned out of the water. This data will help educate people on
how important it is to care for our water systems because such small amounts of these pollutants can
impact the environment.
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Hypothesis
My hypothesis is that in all four pollutants, the highest number of deaths, during both the chronic and
acute tests, will occur in the highest concentrations. I believe that the most harmful pollutants will be the
antifreeze and motor oil.
I base my hypothesis on information from the Environmental Protection Agency web site, which stated the
following: "Chemical compounds, such as oil and gasoline resulting from spillscan poison fish and other
marine organisms. Research has shown that by-products from the biological breakdown of petroleum
products can harm fish and wildlife" I also base my hypothesis on statistical information from the
Association of Washington Cities City News that automotive fluids are the most common pollutant found in
storm drains and creeks.

Experiment Design
The constants in this study are:
* The type of beta cup used as the environment
* The water, ultra-pure water

* The species of daphnia, pulex

* The equipment used to measure the water and the pollutants
The manipulated variable will be the type and amount of pollution used. The pollutants will include lignin
sulfonate, used motor oil, antifreeze and milk.
The first acute test the concentrations used, in percent by volume, were:
* 50%, 20%, 10%, 5%, 1% and control for all four pollutants.
Based on the lethal results of the first acute test, I decided to lower the concentrations and re-run the
acute tests.
The second acute test concentrations used, in percent by volume, were:
* Milk - 20%, 15%, 10%, 5%, 1% and control.
* Antifreeze - 5%, 4%, 3%, 2%, 1%, and control.
* Lignin sulfonate - 1%, 0.5%, 0.3%, 0.2%, 0.1%, and control.
* Motor oil - 1%, 0.5%, 0.3%, 0.2%, 0.1%, and control.
The chronic concentrations used, in percent by volume, were:
* Milk - 5%, 3%, 1%, 0.5%, 0.25%, and control.
* Antifreeze - 5%, 3%, 1%, 0.5%, 0.25%, and control.
* Lignin sulfonate - 0.25%, 0.2%, 0.15%, 0.1%, 0.05% and control
* Motor oil - 0.1%, 0.08%, 0.06%, 0.04%, 0.02%, and control
The responding variable will be the rate of death of the daphnia pulex in each concentration during the
acute test. For the chronic test, the responding variables will be the rate of death, and the possible
increase in population of the daphnia pulex.
To measure the responding variable during the acute test, I will count the number of daphnia in each cup
after a 24-hour period and again at 48 hours. The chronic test runs for seven days. On day seven, I will
count the number of daphnia pulex to see if there was an increase or decrease in population. This will
measure the stress on the daphnia. If they are stressed they will not reproduce; this will impact the
population count.

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Materials
Quantity

ItemDescription

8 meters

5mm aquariumair tubing

Aquarium air pumps

Gang valve assemblies

Plastic tee connectors

120

Discard-A-Stoneair diffusers

26

Tubing nipples

16 liters

Ultra-purewater

1.536g

NaHCO3 (Sodium Bicarbonate)

0.96g

CaSO4.2H2O (Hydrated Calcium Sulfate)

0.96g

MgSO4 (Magnesium Sulfate)

0.064g

KCI (PotassiumChloride)

1200

Neonate daphniapulex

500mL

Distilled water

Triple beam gramscale

6.3grams

Finely groundtrout chow

2.6 grams

Dried yeast

0.5 grams

Dried alfalfa

2 3.78L

glass jars

293.50mL

Milk

221.50mL

Antifreeze

197.50mL

Motor oil

197.50mL

Lignin sulfonate

50mL graduated cylinder

1000mL graduatedcylinder

1mL syringes

Eyedropper

White plate

White plastic spoons

Hole Punch

Pair scissors

Computer

As Needed

Paper

As Needed

Pencil/Pen

As Needed

Film forpictures

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Procedures
Acute Test #1:
1. Clean out six beta cups and lids.
2. Punch a hole in each lid for the air hose.
3. Cut four 25cm lengths of air hose to run from the beta cup to a valve on gang valve.
4. Cut a seven-centimeter length of hose to attach from one valve on the gang valve to a T-valve. Then cut
two more seven-cm lengths to attach to each side of the T-valve. These lengths will be used for beta cups.
5. Attach a Discard-A-Stone air diffuser with an airline nipple to one end of each hose.
6. Insert the other end of the hose through the hole in the lid.
7. Attach all the hoses to the gang valve.
8. To attach the gang valve to the air pump, cut a seven-cm length of hose to attach to the pump and the
gang valve.
9. Add each concentration, in percent by volume, of pollutant: 50%, 20%, 10%, 5%, and 1%. Control is
100mL of ultra-pure water.
10. Add the remaining amount of milliliters of water to each cup to equal 100mL.
11. Repeat steps 1-10 to create a replicate of the test for cross-referencing.
12. Using the eyedropper, remove 120 neonate daphnia pulex from the breeding tank and place ten in each
cup.
13. Place the air diffuser flat on the bottom of the beta cup and replace the lid.
14. Plug in the air pump, and using the adjusting switches on the gang valve, regulate the airflow in each
beta cup.
15. To count the daphnia:
a) After 24 hours, remove one concentration cup at a time from the gang valve.
b) Gently pour one solution onto the white plate. This helps in viewing the daphnia.
c) Using the eyedropper, remove all living daphnia that are visible. Place and count them in the bowl of a
white plastic spoon.
d) Rotate the plate and watch for any other movement.
e) If after a few minutes you see no more live daphnia, put the solution back in the beta cup and re-add the
daphnia.
f) Record the number of live daphnia and any comments. Repeat steps a-e for all test concentrations.
g) Reattach the hose to the same position on the gang valve.
16. After 48 hours repeat step 15 to count the daphnia. This ends the acute test.
17. Repeat steps 1-16 for all four of the pollutants. Use fresh materials for each of the pollutants.

Acute Test #2:

After running all four pollutants in the first acute test, and discovering that they were extremely lethal, I
decided to lower the concentrations and re-run the acute tests. I will use the following concentrations in
percent by volume:
* Milk - 20%, 15%, 10%, 5%, 1% and control.
* Antifreeze - 5%, 4%, 3%, 2%, 1%, and control.

* Lignin sulfonate - 1%, 0.5%, 0.3%, 0.2%, 0.1%, and control.

* Motor oil - 1%, 0.5%, 0.3%, 0.2%, 0.1%, and control.
Chronic Test
1. Clean out six beta cups and lids.
2. Punch a hole in each lid for the air hose.
3. Cut four 25cm lengths of air hose to run from the beta cup to a valve on gang valve.
4. Cut a seven-centimeter length of hose to attach from one valve on the gang valve to a T-valve. Then cut
two more seven-cm lengths to attach to each side of the T-valve. These lengths will be used for beta cups.
5. Attach a Discard-A-Stone air diffuser with an airline nipple to one end of each hose.
6. Insert the other end of the hose through the hole in the lid.
7. Attach all the hoses to the gang valve.
8. To attach the gang valve to the air pump, cut a seven-cm length of hose to attach to the pump and the
gang valve.
9. Based on the results of the acute tests, I will use the following concentrations, in percent by volume, for
each pollutant:
* Milk - 5%, 3%, 1%, 0.5%, 0.25%, and control.
* Antifreeze - 5%, 3%, 1%, 0.5%, 0.25%, and control.
* Lignin sulfonate - 0.25%, 0.2%, 0.15%, 0.1%, 0.05% and control
* Motor oil - 0.1%, 0.08%, 0.06%, 0.04%, 0.02%, and control.
10. Add the remaining amount of milliliters of water to each cup to equal 100mL.
11. Repeat steps 1-10 to create a replicate of the test for cross-referencing.
12. Using the eyedropper, remove 120 neonate daphnia pulex from the breeding tank and place ten in each
cup.
13. Place the air diffuser flat on the bottom of the beta cup and replace the lid.
14. Plug in the air pump, and using the adjusting switches on the gang valve, regulate the airflow in each
beta cups.
15. To count the daphnia:
a) After seven days, remove one concentration cup at a time from the gang valve.
b) Gently pour one test solution onto the white plate. This helps in viewing the daphnia.
c) Using the eyedropper, remove all living daphnia that are visible. Place and count them into the bowl of a
white plastic spoon.
d) Rotate the plate and watch for any other movement.
e) Record the number of live daphnia and any comments. Repeat steps a-e for all concentrations and
controls of both sets of pollutant.
16. Repeat steps 1-15 for all four of the pollutants. Use fresh materials for each of the pollutants.
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Research Report
Introduction
This report will discuss the topics of clean water, waste water treatment, daphnia pulex, milk, antifreeze,
lignin sulfonate, oil, water environments, and biochemical oxygen demand. Water is essential and extremely

important to our lives. We need to take care of this natural resource; otherwise it will be gone forever. I
hope you enjoy reading this report and learn something new about these topics.
Why Do We Need Clean Water?
Americans use rivers, lakes, and aquifers for drinking water. About half of the population uses
underground water sources for drinking water. The other half uses surface water. The water that is on
earth today is the same water that was on earth since the beginning of time. If we pollute this water, the
effects will be here forever; there will never be any new water. A survey in Money magazine found that
clean air and water are among the top ranked factors people consider in choosing a home.
The Great Lakes make up 95 percent of fresh, surface water in the United States. There are 3.5 million
miles of streams and rivers and 41 million acres of lakes. In Alaska alone, there are 170 to 200 million
acres of wetlands. In the rest of the US, there are 101 million acres of wetlands. All of this water is used
for tourism, transportation, recreation, and industry.
Americas commercial fishing and shell fishing industry relies on clean water to deliver products that are
safe to eat. Manufacturers use approximately 13 trillion gallons of water every year. This is more than the
amount of water that flows from the Mississippi into the Gulf of Mexico every day. The soft drink
industry uses over 12 billion gallons of clean water every year to produce their products. Think about it, if
the water reached a point where it could not be cleaned, you would have to drink dirty Dr. Pepper!
Crops grown over much of the country use water for irrigation. If the water isnt clean, the food the
nation depends on wont grow as well. If that dirty water gets into the food, then people everywhere will
consume it. This could make many people sick.
It is estimated that every year Americans take 1.8 billion trips, about seven trips per person, to water
destinations. The next time you think about dumping something into the water, think again. Do you really
want to be swimming in it?
Cleaning Water
It takes many different steps to clean water. Domestic sewage comes from peoples everyday activities.
Industrial wastewater varies depending on the type of industry. Wastewater travels from homes and
industries through underground pipes that are about 20 to 30 centimeters in diameter. Usually, these
sewer pipes flow to larger pipes under the city streets. Wastewater then flows to the treatment plant.
Once at the treatment plant, wastewater goes through primary treatment, secondary treatment, and
sometimes, advanced treatment. When the wastewater reaches the plant, it contains debris that could
damage or clog the machinery and pumps. The debris is removed by vertical bars or screens, and then is
burned or buried. With these materials removed, the water goes through a grinder where organic
materials are reduce to an appropriate size. Next, the grit chamber removes silt, sand, gravel, and
cinders. These materials are removed and disposed of in a sanitary landfill. Then, in a sedimentation tank,
called a clarifier, organic materials settle and are drawn off for further treatment and disposal. An
alternative to sedimentation is flotation, where air is forced into the water, which is then discharged into
an open tank. Here, the rising air bubbles make suspended solids rise to the surface where they are
removed. Solids removed from the settling tanks are called sludge. The sludge can receive more
microbiological digestion and be placed on sand beds for air-drying. Percolation and evaporation dry the
sludge, which is then used as a soil conditioner or fertilizer. The water is then sent to secondary
treatment.
Secondary treatment uses biological processes to remove dissolved material from the water. One type of
secondary treatment is the trickling filter. In this process, the wastewater is distributed over a bed or
column of porous medium. A film of microorganisms coats the medium and works as the removal agent. The

most common form of secondary treatment is called the activated sludge process. Here, microorganisms
and wastewater are mixed in a basin with air. These processes are followed by another sedimentation
tank to remove the microorganisms. A stabilization pond or lagoon is also a form of biological treatment.
Solids are decomposed in the bottom of the pond or lagoon.
Advanced wastewater treatment is required when the receiving water needs a higher degree of treatment,
or if the water is intended to be reused. Third stage treatment is used to remove nutrients like nitrogen
and phosphorus. Advanced treatment can also include additional steps to remove refractory pollutants.
Now, the water is clean enough to be discharged into the receiving water. The most common practice is
direct discharge into the stream or lake. This treated water is also used for groundwater recharge,
industrial processing, recreation, and irrigation of non-edible crops.
Water Environments
Freshwater life is the plants, animals and other life forms that are adapted to live in streams, rivers, lakes
and ponds. There is a huge variety of species living in these static and flowing environments.
Lotic habitats, or flowing water habitats, include rapid headwater streams, slower streams that have pools
and riffles, and the slow moving rivers of the floodplains and estuaries. The species in the fast flowing
streams have adaptations that let them keep their position in the fast current. Some species are
streamlined so there isnt as much resistance from the current. Other species have flattened bodies so
they can hide beneath rocks. Plants in these fast-moving waters include moss and algae. The animals in the
slower moving waters are more adapted to warmer and slower moving water. Plankton begin to appear and
aquatic plants with roots grow on the shore.
Most streams depend on nearby terrestrial ecosystems for an energy source. Aquatic insects eat bacteria,
which have softened leaves and wood. From here, the food chain continues to fish, larger fish, then to
predators such as bears, and eagles. Humans eat these fish, too.
Pond and lake habitats, or lentic habitats, include a shallow water zone, an upper open water zone, a deep
water zone and a bottom zone. Often, the shallow water zone contains emergent vegetation, such as
cattails, submerged vegetation, and floating vegetation like duckweed. Species such as plankton and fish
live in the open water zone. Temperature and the amount of oxygen affect life in the deep-water zone.
Pollutants also affect life in all of these zones.
Daphnia Pulex
Daphnia pulex, commonly known as the water flea, are in the class Brachiopod. These fresh water
crustaceans are found in ditches and ponds all over North America except for high arctic areas and the
tropics. They are also expected to be found in South America. There is little variation in the physical
characteristics of the daphnia throughout its range.
Daphnia are mostly pond dwellers where the oxygen content is higher that that of lakes, but some have
been found in lakes. These creatures are dominant in nature during periods of low turbidity, but tests have
shown that high turbidity has little effect on them. In mixed populations of daphnia pulex and magna, pulex
usually out-competes the magna and by the end of the summer is the only inhabitant. Populations of
daphnia are usually low in the winter and early spring, but when water temperatures rise between 6 and
12 degrees Celsius, their population increases. These population increases lead to approximately 200
to 500 individual daphnia per liter. During the summer, populations in ponds decline during the summer
months, then increase in the fall. The life span of the daphnia increases as the temperature of the water
decreases due to a lower metabolic activity.

Daphnia pulex are approximately 3.5 millimeters in length and move with darting leaps through the water.
They are reddish in color and encased in a transparent, bivalve shell. Daphnia have five to seven teeth in
the middle pecten. Feathered, branched antennae on the head are used for swimming. Daphnia eat algae,
bacteria and plankton.
There are four distinct periods in the approximately 50-day life cycle of the daphnia pulex. These periods
are; egg, juvenile, adolescence and adult. During each of these periods, the daphnia goes through several
instar stages. Periods of instar occur between molting of the exoskeleton or bivalve shell and are
completes when a molt occurs. Growth occurs immediately after each molt while the new carapace is still
flexible.
The organism growth rate is greatest during the juvenile stage. The adolescent period is very short and
has only one instar, when the first clutch of eggs are deposited in the brood chamber. The adult pulex has
18 ? 25 instars that last about two to seven days depending on environmental conditions. During each adult
instar, four events occur in just a few minutes: the birth of young, molting, growth, and the release of new
eggs into the brood chamber. Daphnia pulex commonly produces six to ten young during each instar.
The Environmental Protection Agency uses daphnia pulex as a test organism of choice for the toxicity test.
Comparative tests have shown that daphnia are more sensitive than trout. They are also used because they
do not have a protective mucus secretion that protects fish against pollutants, which better shows the
lethality of the pollutants. There are also fewer culturing problems with the daphnia pulex over daphnia
magna.
Biochemical Oxygen Demand
The biochemical oxygen demand, BOD, test is used to determine how much oxygen bacteria need to
breakdown a pollutant or waste. The test usually runs for five days at a constant temperature of 20
degrees Celsius. Just before starting the test, the amount of dissolved oxygen in the test sample is
measured. At the end of the test, the oxygen concentration is measured again. To find the oxygen
demand, you subtract the amount of dissolved oxygen on day five from the amount of dissolved oxygen on
day one. For example, if the amount of dissolved oxygen, DO, is 3mg/l on day one and the DO on day five is
2mg/l, then the oxygen demand is 1mg/l. Next, you multiply this by the dilution ratio. So, the biochemical
oxygen demand in the example test is 100mg/l, at a 100 total dilution.
BOD is important because if there isnt very much dissolved oxygen in the water, the bacteria or daphnia
have difficulty in surviving. The higher the BOD level, the less oxygen there is available for the animals in
the solution. A high BOD level usually causes the higher life forms in the water to suffocate and die.
Bioassay Tests
Bioassays, or toxicity tests, use biological organisms to assess the effect of chemical effluents on the
organisms and their receiving waters. The objective of this test is to estimate the "no effect" or "safe"
concentrations of a pollutant on aquatic life. Most often, test observations are limited to growth,
reproduction, and death. This test approach is different that conventional testing which are based only on
a chemical specific basis, but toxicity tests use a whole effluent approach. The whole effluent approach is
used to determine the combined affect of the pollutant.
Bioassays are used to:
* Determine if environmental conditions are suitable for aquatic life;
* Establish acceptable pollutant concentrations in receiving waters;
* Compare the effects of wastewater to other toxic chemicals;
* Determine the amount of treatment needed for wastewater; and
* Evaluate compliance with government permits.

While bioassay test results are useful for protecting human and animal safety, there is some controversy in
using the test because of the difficulty in getting repeatable data under the same conditions.
The Environmental Protection Agency recommends two different types of bioassay tests. They are static
non-renewal test and static-renewal test. In either of these tests 80% of the original adult daphnia in the
control must survive for the test to be acceptable. Of this 80%, 60% must produce 3 broods in 6-8 days
with 15 or more young. Some factors that can produce a less than precise test are the age, condition and
sensitivity of the test organism, temperature control, feeding, and oxygen content. The oxygen content
must be 4mg/L or higher. All of the organisms in the test must be counted within two hours or there could
be another brood hatching, which would increase the count. Some states use species native to the
receiving waters instead of the EPA recommended organism, which may be as or more sensitive than the
EPA recommended organism.
Some terms that are used by the EPA to define the results of the bioassay are LC50, NOEC, LOEC, EC, LC,
and IC.
* LC50 stands for lethal concentration 50%, which is the concentration at which half of the test organisms
died.
* NOEC, or no observed effect concentration is the highest concentration of pollutant at which there is no
observable effect on the test organism.
* LOEC stands for lowest observed effect concentration, this shows the lowest concentration that causes
an effect on the organisms.
* EC, or effective concentration is the pollutant concentration that causes an observable adverse effect on
the test organism.
* LC, lethal concentration, is closely related to LC50, in which a pollutant concentration causes death at a
given percent.
* IC stands for inhibition concentration that causes a reduction in the young per female or growth in the
test population.
Milk
Milk is made up of globules of butterfat that are suspended in a solution made up of milk sugar or lactose,
proteins, salts of calcium, chlorine, phosphorus potassium, sodium, and sulfur. Milk lacks dietary iron and
isnt an adequate source of vitamin C. Eighty to ninety percent of whole milk is water. When milk is fresh,
it has a slightly sweet taste and nice odor. Because the globules of butterfat have a lower specific gravity
than that of the milk solution, they will rise to the top of the container to form cream. Cow milk is 3.68%
fat, 4.94% sugar, 2.88% casein (protein), 0.51% albumin, 0.72% ash and 87.27% water.
About half of the milk that is produced in the United Stated is fresh milk. The other half goes info
evaporated and condensed milk, butter, dried milk powder, malted milk, cheese, yogurt, ice cream, and
lactose. The distribution of raw fresh milk is no longer possible because of the urbanization of America.
Today, milk undergoes pasteurization and is then chilled before it is packaged and delivered to insure its
safety. In the United States, commercial milk is cows milk, but in other countries commercial milk includes
goat, llama, reindeer, or buffalo milk.
Milk plant waste, which includes water, milk, and its by-products, has a BOD value of 1,000 mg/l. Whole
milk has a BOD of 102,500 mg/l, skim has 73,000 mg/l, and whey has a BOD of 32,000 mg/l. These are
extremely high biochemical oxygen demand values and milk as a pollutant is quite lethal to aquatic
environments.
Antifreeze
Antifreeze is a chemical that is added to a liquid to lower its freezing point. The freezing point is the
temperature, at which a liquid congeals into its solid state. For example, the freezing point of water is 32

degrees Fahrenheit and 0 degrees Celsius. When antifreeze is added to water, the water wont freeze at
this temperature, it will freeze at a colder point. Antifreeze prevents coolants in cars, industrial plants,
tractors, and refrigeration liquids from freezing.
Good antifreeze should be miscible in the coolant, chemically stable, have a high boiling point, good heattransfer properties, and have low electrical conductivity and viscosity. Viscosity is a property of a fluid
that prevents it from flowing when under an applied force. Low viscosity fluids flow easily while high
viscosity fluids resist flow. The most widely used materials in antifreeze today are ethyl and methyl
alcohol and ethylene glycol. Most antifreeze products also contain nitrate, phosphate or another
anticorrosive agent. There is no BOD value for antifreeze. The ingredients in antifreeze are toxic as a
poison.
Oil
Petroleum, or crude oil, is a liquid composed of organic chemicals. This burnable mineral is found in large
quantities under the surface of the earth. It is used as fuel and a raw material in the chemical industry.
Petroleum and its derivatives are used in the manufacture of foodstuffs, medicines, plastic ware,
fertilizers, paints, building materials, and cloth, and to generate electricity.
Oil is formed under the surface of the earth by the decomposition of marine organisms. The remains of
tiny sea organisms, land organisms that are carried to sea, and plants that grow on the bottom of the
ocean, are meshed in with the silt and sand on the bottom of the ocean. The process of making crude oil,
beginning with these organisms, began millions of years ago. As the sediments grow thicker, they sink into
the floor of the ocean under their own weight. Then, additional deposits pile up, increasing the pressure
on the layer below by thousands of times. The temperature rises by several hundred degrees, and the
mud and sand harden into shale and sandstone. Carbonate precipitates and skeletal shells then harden into
limestone and the remains of dead organisms are transformed into natural gas and crude oil.
There are three broad classes of crude oil. They are the paraffin types, the asphaltic types, and the
mixed base types. The paraffin types are made up of molecules where the hydrogen atoms are always two
more that twice the number of carbon atoms. So, if there are two carbon atoms, then there are six
hydrogen atoms. Molecules in the asphaltic types are naphthenes; they are composed of twice as many
hydrogen atoms as carbon atoms. The mixed-base group has both paraffin hydrocarbons and napthenes.
The bacteria common to the BOD test dont consume oil, so there is no BOD value for it. It is simply toxic
as a poison. Chemical compounds such as gasoline, and petroleum products can poison fish and other marine
life. It is proven that by-products from the breakdown of these products is harmful to fish and wildlife.
Lignin Sulfonate
Lignin sulfonate is a wood product from the chemical family of lignin. Lignin sulfonate is a thick, dark brown
liquid that smells a little like "bacon bits". It boils at 212 degrees Fahrenheit and freezes at 25 degrees
Fahrenheit. It is widely used to produce stable emulsions and dispersions, to reduce the viscosity of
slurries, and to provide adhesion in different binding and granulating applications. An emulsion is the
mixture of two liquids that do not normally mix. Dispersions are similar to emulsions except it applies to a
solid in a liquid. A slurry is a semi-fluid substance such as slush, thin mud, mortar or cement that is
prepared with a high percentage of water.
Lignin sulfonate is used in the cement, concrete, admixture, gypsum board and ceramic industries. It is also
used as dust abatement on roads during the summer. In Yakima County alone, 431 miles of road are
covered in the season from April to July. To cover these roads, 1,709,442 gallons of lignin sulfonate are
used, which breaks down to 3,966 gallons per mile. Yakima County Road Maintenance pays $1,190,351.00 a
year to effectively combat the dusty roads. Water will breakdown lignin sulfonate, so if it rains, the road
gets dusty again, and the lignin sulfonate has to be reapplied to the roads. The lignin sulfonate that washes

off the roads could potentially get into our water. This, along with the dirt it washed in with, could kill the
animals of the rivers or lakes.
Lignin Sulfonate is also used to float pears at the beginning of the pear packing operation. It is used to
enhance the density of the water, making the water heavier so the pears will float out of the bins easier.
At the concentration used for fruit packing, the BOD of the float tank wastewater is 20,000 mg/l, which is
extremely high. The BOD of straight lignin sulfonate is 160,000 mg/l.
Conclusion
This concludes my research report. I hope that you enjoyed it and learned many new, interesting things.
Please remember not to pollute our waters. We need to protect our water, and the life found in these
environments so future generations will benefit from our care.
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Results
The original purpose of this experiment was to determine the effects of pollutants on aquatic life.
The results of this experiment show that the different pollutants did indeed have a negative effect on the
daphnia pulex.
The most lethal pollutant was oil, followed in order by, lignin sulfonate, milk, and antifreeze. Oil was so
toxic that, with the equipment that I had, I couldnt get the concentration low enough. My lowest
concentration, 0.02mL of oil in 100mL of water, is equivalent to 200 liters of oil in one million liters of
water. In the acute test, antifreeze was more lethal than milk, but in the chronic test, it was less lethal.
When the results of acute test number one are compared to acute number two, for milk and antifreeze, the
concentrations of acute number one were less lethal than the lower concentrations of the second test. The
results of the lignin sulfonate chronic test show that even at low concentrations, it is lethal over a longer
period of time. These test results show that this type of test has a great degree of variability.

Conclusion
My hypothesis was that in all four pollutants, the highest number of deaths, during both the chronic and
acute tests, would occur in the highest concentrations. I thought that the most harmful pollutants would
be antifreeze and motor oil.
The results indicate that my hypothesis should be partially accepted and partially rejected. I was correct
that the highest concentrations of pollutants would be the most lethal. I was incorrect that the most
harmful pollutants would be antifreeze and motor oil. In fact, my results showed that motor oil and lignin

sulfonate were overall the most lethal pollutants. Milk and antifreeze were not as toxic as I believed they
would be. Antifreeze proved to be the least toxic pollutant in the chronic test.
Because of the results of this experiment, I wonder if concentrations equal to or less than 0.05% would be
lethal in lignin sulfonate and motor oil, and at what concentrations no chronic effects would be seen. I also
wonder what would happen if I let the chronic tests run for more than seven days. When comparing the
results of the first and second acute tests, the second test was more lethal that the first, even at lower
concentrations. I believe this confirms the reported variability of bioassay tests.
If I were to conduct this project again, I would try to get the daphnia pulex sooner so I could start my
experimentation sooner. I would also suggest that if someone else were to do this test to start with lower
concentrations of pollutants from the beginning.