Research Proposal

Lauren Lagana

Lumbriculus variegatus and its response to exposure to Calcium Magnesium Acetate

Introduction/Background

We propose to perform this experiment on the basis of the lack of knowledge of the
effects of Calcium Magnesium Acetate on aquatic invertebrates, such as Lumbriculus
variegatus. We therefore propose to proceed with the experiment detailed below to increase the
known information about the effects of CMA and at what concentrations it is most toxic to
aquatic invertebrates.

Calcium magnesium acetate is essentially rock salt. It is used as a de-icing agent during
the winter months (Anonymous).1 So far as has been researched (McFarland and O'Reilly) this
substance isn’t hazardous to the environment due to the small amounts in which the substance
would be introduced to the environment.4 However, we believe that this research had unrealistic
results or didn't properly test multiple concentrations.

We will address the past and similar research that has been completed, our hypotheses,
and the intended timeline for the tests. Also, we will discuss the limitations and significance of
this research. The abstracts below are the foundation for this experiment.

Literature Review

‘Behavior, Growth, and Reproduction of Lumbriculus Variegatus (Oligochaetae) in

Different Sediment Types’

A.M. Sardo, A.M. V.M. Soares, and A. Gerhardt

The purpose of this study is to determine “the best sediment types for use of L.
variegatus in sediment toxicity tests, based on grain-size”; and whether or not it is more
beneficial to use artificial cloning or natural cloning to increase the number of test organisms.
The results showed that “locomotory ability was highest in fine [sediment]”. The second part of
this experiment concluded that “artificial cloning is not a recommended method to obtain
additional test organisms”.

The information gathered by this experiment is useful to our own laboratory experiment
because we are testing the locomotory abilities of L. variegatus, and the artificial cloning and its
effects on the clones. Therefore, the research concluded by Sardo, Soares, and Gerhardt is
highly valuable to our own experiment. We anticipate this article will also be useful to future
experiment of a similar nature.2

“Evaluation of calcium magnesium acetate and road salt for contact hypersensitivity
potential and dermal irritancy in humans.”

Cushman, JR, Duff, VA, Butfau, GH, Aust, LB, Caldwell, N, Lazer, W

Calcium magnesium acetate, a de-icing agent, is a common chemical to which “workers

may be dermally exposed”. Human exposure to CMA has shown to have “moderate acute
irritation was observed [only at 1 skin site exposed to] 30% road salt”. The results of this
experiment predict that CMA will not “cause significant dermal effects [to] exposed workers”.
 It is anticipated that the comparison between hyper sensitivity of humans versus
Lumbriculus variegatus is a large margin. However, we believe the similarity of these two
experiments will allow for more accurate hypothesis on the effects of CMA on the worms. Thus
this article has become quite useful to our particular group’s experiment.3

“Environmental Impact and Toxicological Characteristics of Calcium Magnesium

Acetate”
McFarland, BL, O’Reilly, KT
The purpose of this study was to determine the impact of CMA on both the
environment and animals. It has shown that CMA has “little or no toxic effects on the aquatic
species that have been tested, including vertebrates and invertebrates” for short-term tests. This
experiment concluded the same results for roadside vegetation and road worker’s health. The
resulting conclusion is that CMA would have an insignificant impact, both environmentally and
toxically.

The results of this study are significantly different from our own experiments. We
have concluded that CMA does indeed have significant negative effects on the invertebrates,
Lumbriculus variegatus, and in extremely small doses as well. We therefore conclude that the
above experiment was helpful in the sense that it has shown how important it is to factor enough
time into the experiment and in multiple varieties of concentration of solutions.4

Hypothesis

We hypothesize that:

Pulsation Rate Test: If the solution type is related to pulsation rate in L. variegatus, then the
CMA solutions will speed up the pulsation rate of L. variegatus, and lead to eventual death.

Locomotion Test: If L. variegatus are exposed to the CMA solution, then the worms will become
slower than the control group, as liquid is drawn out of the body.

Drop Test: If the CMA solution produces a negative chemotactic response in L. variegatus, then
the worms will move quickly away to escape it.

Methods

The Experimental Design of the proposed study is for thirty worms to be used per
hypothesis we would like to test. There will be three tests and thirty worms per test. Of these
thirty worms, ten will be in each group. Each of the three groups will be exposed to a different
concentration of the solution. The control group will be placed in regular spring water and the
two treatment groups will be placed in solutions of CMA and spring water at specific
concentrations. All observations will be recorded.

The first test is the Pulsation Rate Test, in which the worms will soak for 10-15 minutes
in solutions of 1% and 5% CMA and spring water. After which their pulsation rates will be
measured for ten seconds. The resulting number will be multiplied by six for the pulsations per
minute.

The second test is the Locomotion Test, in which the worms will soak for 10-15 minutes
in solutions of 0.5% and 1% CMA and spring water. They will then be placed in a Petri dish
turned ‘racetrack’ with measurements in centimeters on the side. The worms will be gently
prodded every five seconds for thirty seconds; which is the whole duration the worms will be on
the ‘racetrack’. The number of centimeters the worms travel will be multiplied by two for distance
traveled per minute.

The third test is the Drop Test, in which each worm will be placed in a Petri dish and a
single droplet of our solutions will be placed on the worm’s tail. The concentrations of the CMA
solutions will be 0.5% and 1%. The worms will then be observed for a positive, moving towards
its tail, or negative, moving away from its tail, response to the solution. They will be observed for
thirty seconds.

Timeline

Week 1
• Observe the worms under a microscope
• Pulsation Test
Week 2
• Setting up the Locomotion Test
• Locomotion Test
Week 3
• Drop Test
Discussion

The limitations of this experiment are few resources found concerning prior research of
this nature and the fact that our results would not be easily comparable to humans in the sense
of toxicity (Cushman, Duff, Butfau, Aust, Caldwell, Lazer)3 . The significance of this experiment
will be the increase in knowledge about the effects of CMA on aquatic invertebrates. We will
also be able to credit or perhaps discredit past research with this study depending on our
results. Therefore our research is necessary to help determine the environmental safety of using
CMA instead of the usual rock salt or other chemicals to de-ice environments (Brown).5 Our
semi-repeat experiment (McFarland and O’Reilly)4 of a prior study should either reinforce or
disprove previously published information.

Working Bibliography
1
[Anonymous]. CMA – Questions & Answers [Internet]. National Sales & Supply

Available from: http://www.nsalessupply.com/CMA%20Question%20and%20Answer.htm Accessed

2008 July 11.
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2
Sardo, A.M., Soares, A.M. V.M., Gerhardt, A. Behavior, Growth, and Reproduction of
Lumbriculus Variegatus (Oligochaetae) in Different Sediment Types [abstract] [online database]
Available from: http://web.ebscohost.com/ehost/detail?vid=4&hid=109&sid=8279254c-d826-4a7e-
8bd7-b38aa6b679f6%40sessionmgr102 Accessed on 2008 July 11
3
Cushman, JR, Duff, VA, Butfau, GH, Aust, LB, Caldwell, N, Lazer, W Evaluation of calcium
magnesium acetate and road salt for contact hypersensitivity potential and dermal irritancy in
humans Available from: http://www.ncbi.nlm.nih.gov/pubmed/1831107 Accessed on 2008 July 11
44
McFarland, BL, O’Reilly, KT Environmental Impact and Toxicological Characteristics of
Calcium Magnesium Acetate Available from:
http://md1.csa.com/partners/viewrecord.php?requester=gs&collection=ENV&recid=9306831
Accessed on 2008 July 11
3

5
[Ellen Brown] Solutions for Slippery Sidewalks [Internet]. Thrifty Fun. 1997

Available from: http://www.thriftyfun.com/tf16691674.tip.html

Accessed 2008 July 11.

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