This activity helps
students gain an appreciation for how scientists determine
the human health effects of hazardous substances. Students
also demonstrate how hazardous substances can affect the health
of test animals. Note: This activity involves the exposure
of worms to a hazardous substance. Some students may object
to this on ethical or moral grounds.
Toxicology is the
study of the effects of poisons on living organisms. Scientists conduct
a variety of studies to discover toxicological information about hazardous
substances. Two of the most common types of studies are (1) epidemiological
studies-matching disease and other adverse health effects in
humans with possible causes-and (2) animal toxicological studies.
The Federal government's
Superfund Program, administered by the U.S. Environmental Protection
Agency (EPA), helps protect people and the environment by cleaning up
hazardous waste sites. Well-designed, properly controlled epidemiological
studies conducted at or near hazardous waste sites can provide information
important in making cleanup decisions.
On their own, these studies
are not always conclusive. This is primarily because it is difficult
to determine the exact amount of the chemical or chemicals contaminating
the site to which human populations have actually been exposed (had
contact with). Many times health histories are incomplete, and potentially
exposed populations are too small for statistical analyses. In addition,
many uncontrolled variables-such as genetics, exercise, diet, or cigarette
use-may complicate detecting the effects of the hazardous substances.
When epidemiological studies
cannot be done, well-designed animal studies can provide a wealth of
information. This information can be used to predict potential effects
in humans over a range of exposure levels-from acute,
a single exposure to a hazardous material for a brief length of time,
to chronic, continuous or repeated exposure to a hazardous substance
over a long period of time.
For more information on health
effects caused by hazardous substances, see the Suggested
Reading list found at the end of the Haz-Ed materials. Other
Haz-Ed materials that are related to this topic include Fact
Flash 1: Hazardous Substances and Hazardous Waste; Fact
Flash 9: Common Contaminants; and Activity
6: Examining the Effects of Pollution on Ecosystems.
Assemble the following
- Small plastic cups
(5 per group)
- Three empty one-liter
plastic soft drink containers with caps
- Refrigerated tap water
- Automobile antifreeze
- Live, fresh-water
black (hair) worms, sold as fish food in pet stores (50 or more
worms per group)
- 16-ounce measuring
- 1-ounce measuring
cup (used to measures doses of cough syrup)
- Plastic wrap
- Tape and markers
- Copies of the Student
Handout, Black (Hair) Worm Experiment (1 per group)
- Copies of the Student
Worksheet, Black (Hair) Worm Experiment (5 per group)
hair worms are inexpensive, easy to see because of their dark
color, and quite active. They survive best in a small amount
of refrigerated water (they die if submerged) and should be
kept in the refrigerator until class time. If washed every day,
they can live 1 to 2 weeks in the refrigerator. If worms are
not available, you may substitute some other fresh water invertebrate,
which can be obtained at tropical fish or pet stores. Brine
shrimp, available at some pet stores, also may be substituted
for fresh water invertebrates, but you will need to add table
salt at a 5 percent solution in the water before adding the
- Fill each of the three
1-liter containers (they must be clean) with 16 ounces of water and
clearly mark the 16-ounce level on the side. Pour out the water and
shake the containers dry.
- Label the containers either
Low (6 percent), Medium (12 percent), or High (24 percent) ethylene
glycol. For the low-dose solution pour 1 ounce of antifreeze in the
container marked Low (6 percent) and fill up to the 16-ounce mark
with water. For the medium-dose solution pour 2 ounces of antifreeze
in the container marked Medium (12 percent) and fill up to the 16-ounce
mark with water. For the high-dose solution pour 4 ounces of antifreeze
in the container marked High (24 percent) and fill up to the 16-ounce
mark with water. Shake or stir well. (NOTE: Any substance can be
toxic in a high enough concentration. Handle the antifreeze carefully.)
- Contact your local sanitation
or health department to request information on the proper disposal
methods for antifreeze in your community. Can it be poured down the
drain? Is there a recycling center for this type of substance?
- Using the information
in the Background section, discuss how
scientists conduct studies to get information on toxic substances.
- Divide the class into
teams of two to four students each. Caution students to handle
the antifreeze carefully.
- Provide each team with
five clean plastic cups, tape, marker, one copy of the Student
Handout, and five copies of the Student
- Have the students label
the first container "low dose," the second "medium dose," and the
third "high dose." Have them label the fourth container "control pre-test"
and the fifth "control post-test."
- Provide each team with
some live worms. Have all groups pour just enough cold water into
the "control pre-test" container to barely coat the bottom. Too much
water will drown the worms.
- Have the students place
about 10 worms in the water and watch for any behavioral changes,
recording the results at the end of 4 minutes on the Student
Worksheet. Have them leave the worms in the container.
- Have teams pour just
enough water-antifreeze solution into each container to barely coat
the bottom, using the solutions you prepared in advance. Have half
of the teams start with the "low dose" container first and proceed
in order to the "high dose." Have the other teams start with the "high
dose" container first and proceed in order to the "low dose" container.
- Have the students conduct
each test one at a time, using different worms for each container
(about 10 per container). Remind teams to record their observations
for each test on the Student Worksheet.
- Have all groups end the
experiment by pouring just enough cold water to barely coat the bottom
of the "control post-test" container. (NOTE: Control observations
at the beginning and end of the experiment are intended to help rule
out effects not related to the antifreeze, such as water temperature.)
- After the experiments,
have each group describe the worm behavior they observed during each
test and discuss the answers to the questions on the Student Worksheet.
- Have students dispose
of the antifreeze solutions properly, according to the information
you received from your local sanitation or health department.
- Have the students plan
and conduct an experiment to determine if there is a concentration
of antifreeze and water that does not appear to change the behavior
of the worms over a 24- or 48-hour period. The purpose is to determine
if there is a threshold for an acute (rapid) effect; in other words,
a level of exposure below which there is not likely to be an adverse
health effect in the short-term. The students' experimental plan should
at the very least include an appropriate control group, a sufficient
number of worms, observation procedures, and an explanation of the
experimental conditions, including procedures for rinsing the worms
once a day, cleanliness, covering containers to prevent evaporation,
and refrigerator temperatures.
- Have the students search
the library for information on worm biology. Focus their attention
on worm anatomy and physiology, function in ecosystems, and whether
the adverse effects of antifreeze on worms might be compared to the
potential effects of antifreeze on human health or ecosystem health.
Teacher's Answer Key -- Black
(Hair) Worm Experiment
||Were there obvious
behavioral differences between the control groups and the antifreeze-exposed
groups? If yes, describe.
There should be differences in mobility even with only 10 animals
per group. After about 10 minutes nearly all worms exposed to the
antifreeze solutions probably will be dead. The higher the concentration
of antifreeze the faster they die. Also, the smaller the worms (young
worms) the faster they die. In the unlikely event there are no differences
between treated and control groups, perhaps more worms per group
are needed, or the antifreeze concentration is too low to cause
an observable effect, or the worms are not susceptible to the adverse
effects of antifreeze. All of these possibilities could be tested
in another experiment if materials allow.
||Did the concentration
of antifreeze in the water influence the degree of behavioral change?
If yes, describe.
After an initial increase in activity, you should find that the
higher the dose, the more quickly the worms' mobility decrease.
The degree and severity of toxic effects are primarily a function
of dose (the amount of contact or exposure to the chemical). However,
many other factors including differences in susceptibility among
individuals within a species also influence the outcome. Because
humans manifest an unusual degree of individual variability, large
numbers of people must be similarly exposed to clearly demonstrate
that a chemical causes a specific toxic effect. Using animal toxicity
studies to determine the potential adverse effects of specific substances
has many advantages. Genetically similar individual animals can
be used in relatively large numbers and exposures can be controlled
over a range of dose levels. The results of animal toxicological
studies are used to predict potential effects in humans at dose
levels relevant to possible human exposures.
Was there a safe
level of exposure? In other words, was there an antifreeze solution
that did not appear to cause an effect over the 4-minute observation
period? How could you tell?
Ideally, at least one dose level in an experiment should have
an observable effect different from the others during the observation
period. That is, if all the doses cause the same reaction, you
have only learned that antifreeze has an effect at a concentration
equal to or greater than the lowest dose used. You have not determined
the minimum concentration that will cause an effect, or the maximum
concentration that has no effect.
If all three doses in your experiment caused the same reaction
during the 4-minute observation period, you may want to repeat
the experiment using a lower concentration. For example, you could
prepare a 3 percent solution by pouring 1/2 oz. of antifreeze
into one of the liter containers you used earlier, and filling
it up to the 16 oz. mark with water. Then repeat the experiment
using this solution and observe the reaction. Are the results
different after 4 minutes? How about after the total time that
elapsed during the other experiment?
Does the acute (rapid,
short-term) effect of antifreeze on the worms indicate anything
about what the long-term or chronic (lifetime) effects might be?
No. Long-term or chronic exposures to hazardous substances frequently
result in different effects from those observed after only a brief
exposure. This makes the effects of long-term exposure more difficult
to link to a specific cause. Reproductive organs may be affected;
mutations in cell structure, liver damage, and so forth may not
show up until the next generation.
the only possible measure of effect? Why or why not?
although behavior is an inexpensive indicator of a potentially fatal
adverse effect, it is not the only measure of effect. It is used
in this experiment because it does not require sophisticated equipment
Can you determine
from this experiment the cause of death of the worms?
No. Damage could have been done to vital organs such as the liver
and kidney, which would subsequently make the worms too sick to
move, or the antifreeze could have directly affected their neuromuscular
system, brain, sensory systems, and other organs, thereby slowing
adversely affect living organisms through a variety of mechanisms,
many of which are not yet known. Some chemicals alter DNA, damage
DNA repair mechanisms, or destroy cells by damaging their membranes,
interacting with cell receptors, depleting substances essential
to cell survival, or inhibiting production of vital enzymes. Some
potentially hazardous substances are not hazardous until the body
breaks them down (or metabolizes them) into substances that are
toxic. For example, carbon tetrachloride is broken down in the
liver to a highly reactive chemical that initiates a chain reaction
which destroys a crucial liver cell enzyme system (cytochrome
possible that while antifreeze affects the health of worms, it has
no effect on humans? Is it also possible that antifreeze has no
effect on worms but adversely affects human health? If yes, describe.
The answer to both is yes. However, epidemiological studies and
accidental poisonings verify that antifreeze causes serious and
often fatal effects when ingested by humans. The liver breaks down
ethylene glycol into aldehydes, glycolate, oxalate, and lactate
that may initially cause nausea, seizures, respiratory failure,
coma, and cardiovascular collapse. Survivors of the acute phase
ultimately exhibit kidney failure, severe acidosis (lowered blood
pH), and low blood calcium levels. The fatal kidney damage results
mainly from the formation of oxalate-calcium crystals that precipitate
in the kidney tubules. These changes may also occur in the liver,
heart, blood vessels, and brain. In addition, the aldehydes, glycolate,
and lactate acidify the blood to dangerous levels.