How Does Your Mouthwash Measure Up: Effects ... - Franklin College

How Does Your Mouthwash Measure Up: Effects of Specific Mouthwashes on
Staphylococcus epidermis and Bacillus subtilus
Linsey Page
Mouthwash Lab Report
4/23/02
Abstract:
This experiment was done in order to find out which mouthwash killed the bacteria Staphylococcus epidermis
and Bacillus subtilus the best. The mouthwashes used were Scope (clean mint baking soda), Listermint with fluoride,
Cepacol, Rembrandt, and Therasol. Water was used as a control for the experiment. It allowed students to practice
using T -values to determine significance of mouthwash effectiveness. The experiment also determined the active
ingredient in the better mouthwash. My own hypothesis was that Scope would work the best on both bacteria. My
hypothesis was proved wrong by Bacillus subtilis when Therasol eliminated the most bacteria. As for Staphylococcus
epidermis Therasol worked just as well as Scope and Cepacol in killing the bacteria. However, there was significance at
the 95% level between Scope and Cepacol. The charts, graphs, and the report below provide more information.

Introduction:
The battle for better breath is taking place all over the world. With each toothpaste and mouthwash claiming to
be the best, how can one possibly determine which one to use? This experiment may perhaps put an end to this
particular problem. Five fairly popular mouthwashes were used in this experiment: Scope, Listermint, Cepacol,
Rembrandt, and Therasol. The effectiveness of each was tested on two different bacteria: Staphylococcus epidermis
and Bacillus subtilus.
Staphylococcus epidermis is a bacterium that is found in the skin, hair follicles, and mucus membranes. It is
found in these areas so that when breaks in the protective skin occur it may enter. This normally takes place after
surgical procedures such as shunt and catheter insertions. This provides an optimum environment for colonization and
is known to cause diseases such as endocarditis and urinary tract infections (Talaro and Talaro).
Bacillus subtilus is a non-pathogenic, rod-shaped, gram-positive bacterium that is capable of producing
endospores during harsh conditions. It is often times used to produce insect toxins, peptide antibiotics, and anti- fungals
used in agricultural production (MICRON). It is a very well known bacterium; scientists have successfully sequenced
strain number 168 (EBI). Although neither of these bacteria is usually found in the mouth, they are easy to grow and
maintain in the laboratory environment.
My own hypothesis for this experiment is that Scope will work the best in killing or inhibiting both of the
bacteria used. Basis for this suggestion is the popularity among consumers and it's constant claim that it works best
Materials and Methods
The Microbiology Lab: Comparison of the Antibacterial Activity of Several Mouthwashes provides a list of
materials needed to perform this experiment (Browder 2). It also provides the procedure one must follow in order to
complete the experiment (Browder 3). It does, however, leave out the careful instructions needed to prevent
contamination of the petri plates. There were rules that had to be followed in order to keep the plates that grow the
bacteria sterile. Handling the bottom of the pipettes that were used would cause unnecessary contamination. The petri
plates that were used had to be kept closed as much as possible. Opening them too much or leaving them open would
cause airborne contaminants to pollute the plate. The tops of the tubes that were used and the tweezers had to be
sterilized before each use. Simply running each of them through a flame kills all unwanted materials.

After reviewing these rules, one could continue with the experiment exactly how it was written in the handout.
Scope (Clean Mint Baking Soda), Listermint with fluoride, Cepacol, Rembrandt, and Therasol were the mouthwashes
used in our experiment. We were to measure the zone of inhibition produced by each individual mouthwash. This zone
is the area in which the bacteria were killed, producing a clear spot. In this experiment, the control group was water.
The water would have no effect on the bacteria. This would allow one to compare the effects of the mouthwashes with
one that proved obsolete. By knowing what the agar would look like with no effects it would be easier to notice those
that had reactions to the bacteria.
Results:
My partners in this experiment were Brandi Emerick and Danny Williams. The results of my experiment were
as follows. Therasol caused the largest zone of inhibition on Bacillus subtilis. This occurred at the 95% significance.
Staphylococcus epidermis had a slight variation. According to the calculated T-values, Therasol and Scope killed the
bacteria with no differentiation. Scope and Cepacol had differences to the 95% significance. However, there is slight
confusion in the fact that Therasol and Cepacol both kill the bacteria with no differentiation. You can see the results of
the entire experiment on Graph #1. You can compare the effects of Therasol, Scope, and Cepacol on Graph #2.
The calculated T-values, along with the standard deviations and means of the entire class data are also attached
(Tables 2-5). Tables 2 and 3 represent the data for Bacillus subtilis and Tables 4 and 5 represent Staphylococcus
epidermis. The T -values, as mentioned above, reveals the relative significance between two different mouthwashes.
Just compare the T -values with the Degrees of Freedom to determine at what percentile the mouthwashes are
significant.
Conclusion and Discussion:
The results given by this experiment showed that overall Therasol, Scope, and possibly Cepacol were the best
mouthwashes available to the public. After retaining this data, research was done to determine the active ingredients in
each. (See table 1 for ingredients) Therasol, which had the largest zone of inhibition for B.subtilis, has an ingredient
called glycerin. This preservative is also used in the fermentation of nutrients to antibiotic production and as a
diagnostic aid (Merck). Therasol also proved successful in the destruction of S.epidermis. However, Scope and Cepacol

were also significant factors in eliminating this bacterium. In Scope there is an ingredient called cetyl pyridinium
chloride. This ingredient is known as a pharmaceutical preservative, which is used as a topical anti-infective, topical
antiseptic and disinfectant (Merck). Cepacol has both glycerin, found in Therasol, and cetyl pyridinium chloride, found
in Scope (Browder). This is the reason why it also was effective on the staphylococcus bacterium.
The given results for Bacillus subtilis went against my original hypothesis that Scope would be the best in
eliminating bacteria. Although, for Staphylococcus epidermis my hypothesis was partially correct, just not for the right
reason. The hypothesis used should have been, that those mouthwashes containing numerous preservatives would best
rid the mouth of any unwanted bacteria. Preservatives keep things fresh and do not allow bacteria to form. This is why
a major ingredient in the food markets are preservatives. The bacteria that have already formed will be killed once the
preservatives have been introduced.
In conclusion, this experiment gave experience in sterile techniques, use of T - values to determine significance,
and the writing of a formal lab report. It had students start with a problem and come to a certain decision to as to why
things occurred as they did. Each student had to come up with the reasoning behind why one mouthwash worked better
than another did, and see similarities between those that seemed to work the same. Placing these results and opinions
into a formal report was also good practice for classes and possible career opportunities to come.
References:
Browder, Steve. Personal Communication
Budavari, Susan. Merck Index. 11th ed. Merck and Co. Inc: Rathway, NJ. 1989.
European Bioinformatics Institute. http://www.ebi.ac.uk/proteome/BACSU/
Microbes in Norwich: Bacillus subtilis. ed. Dr. Stephan Bornemann.
http:/ /www.micron.ac.uk/Organisms/Bacillus/Bsubtilis.html
Talaro, Arthur and Kathleen. Foundations in Microbiology. Ed. Colin H. Wheatley. WM. C. Brown Publishers:
Dubuque, IA. 1993.