Stauber

Plant Oils vs. Bacteria

By: Sarah Joy Stauber
February 1- March 1, 201 2
Mrs. McKenna
Period 4 & 5

(Time1103, 2011)
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Table of Contents
Title Page...........................................................................................................................................................1
Table of Contents.....................................................................................................................................2
Acknowledgements................................................................................................................................3
Purpose, Hypothesis & Rational...................................................................................................4
Review of Literature.................................................................................................................................5
Materials.............................................................................................................................................................11
Procedure......................................................................................................................................................1 2
Variables.........................................................................................................................................................14
Results...............................................................................................................................................................15
Experimental Error...................................................................................................................................17
Conclusion...................................................................................................................................................18
Reference List.............................................................................................................................................19

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I would like to thank Mrs. McKenna for her patience and helping me write this paper.
Also, I want to give thanks to my parents, and especially my mom for pressuring
me into doing Science Fair this year and for listening to me explain how poison ivy
and peppers work. And then there are all my friends whom have given me the
moral support to push me through this, especially Elizabeth for also helping me
write this. And thank you to everyone I have forgotten to mention.

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Purpose:
To find a natural way to kill E. coli (with plant oils)

Hypothesis:
If E. coli is negatively affected by plant oils, then it may become susceptible
to destruction.

Rational:
Some chemicals naturally found in plants have properties that are able to
kill certain kinds of cells. One such is capsaicin, found primarily in cayenne
peppers and reactant to any form of stored energy. There are also
isothiocyanates, which is the spicy chemical found in foods like horseradish and
brussel sprouts. The last is ginger, which is composed of series of volatile oils
such as resin, zingiberol and phellandrene. With this information, a natural
experiment could be composed with the potential ability to kill the dreaded strains
of E. coli.

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Review of Literature
E. coli, or Escherichia coli is a bacteria found in municipal waste treatment
facilities and on farms contaminated by excretion of farm animals. It is deadly
and can spread and infect whole populations through the contamination of food
(Escherichia coli, 2009). One of the kinds of plants in use is hot peppers, which
are composed of capsaicin (What makes peppers hot may also be cool for what
ails you, 2003). Another is ginger-root, which is composed of multiple different
types of volatile oils and pungent
phenol compounds (Davies & Ryan,
2000).
tested

The last plant oil that will be

is

horseradish

which

is

composed of isothiocyanates, or allyl

isothyates (What makes horseradish
hot?, n.d., [online]).

Possibilities for

these naturally occurring plant oils to

cure, if not prevent this potentially
deadly disease are quite likely. Since it
is common knowledge that different

Illustration 1: (E. coli infection and protection,

2010, [online])

plant oils affect different organisms

(substantially in their own manner), it can be assumed that the testing of

different oils on E. coli cells will also affect the different aspects of the cell
differently.
E. coli is a commonly known bacterial pathogen that can easily spread
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over whole continents in mammal populations. Most of the time, it starts out in
a water source which is then consumed by cattle and other mammals. Because
this intestinal bacteria is in-taken through the digestive system, it also inhabits
the gastrointestinal system, slowly decomposing the organs of its live host.
Subsequently, the parasite is responsible for 2% of the western world's cases of
diarrhea.

The only common human to human cases are found in day care-

facilities where there there is a daily handling of excrement without appropriate

sanitation.

A relatively new strain called Enterohemorrhagic Escherichia coli,

also known as E. coli 0157:H7, an organism discovered in 1970, with the first
outbreak in 1988, was responsible for a large outbreak this summer causing
severe diarrhea, kidney failure via hemolytic-uremic syndrome (HUS), hematuria
(bloody urine), and a rare case of pyuria (pus in urine). On top of this, there is
an average between 10,000 and 20,000 cases per year of E. coli. The result of
this, HUS, causes horrific symptoms including watery and bloody diarrhea. This
happens when the bowel movement is replaced into blood.

Soon after this

happens, the host will often have a rapid recovery. Among the 10% of E. coli
cases that lead to HUS, only about 5% of them are fatal, creating the primary
death by kidney failure for children (E. coli 0157:H7 infection, 2007).
The short, gram-negative rod-like E. coli have an antigenic structure (a
stimulator

of

the

production

of

antibodies)

of

carbohydrates

and

lipopolysaccharides. These structures will sometimes form chains that produce

the poisonous endotoxins (toxins produced inside the body, then released upon
death of organism) and exotoxins (toxic excrete into host). These toxins give E.

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coli a variety of ways to spread beyond the digestive system and
poison other parts of the host's body (Escherichia coli, 2009).
The most serious cases occur when E. coli attaches tightly to
intestine walls and then release their exotoxins (Time1103,
2011, [online]).

Illustration 2: (E.
coli bacteria and
Ginger-root is composed of multiple different chemicals, how to handle?,
2011, [online])
one of the major ones are the volatile oils. Originally, this plant
was used medicinally by Asian, Indian, and Arabic medicine up to 4 thousand
years ago. China was using it two millennium ago for to help with digestion,
nausea, arthritis, & heart problems along as a cure for colds, flu-like symptoms,
headaches

&

painful

menstrual

periods.

The

knotted,

beige

rhizome

(underground stem) is used today for a variety of purposes: nausea, motion

sickness, cancer chemotherapy (nausea), reduce pain of osteoarthritis & heart
disease (Ginger, 2011).
Some of the more notable of the over 200 different volatile oils are
gingerol, shogaol, & zigibain.

Gingerol & shogaol are what create the strong

fragrance of between 1%- 2.5% of the rhizome and create the warm sensation
from circulation increase.

Other chemicals are

antispasmodic (relax muscles), carminative (relax

stomach & stimulate peristalsis which is the motion of
food through the gut, reducing gas), diaphoretic

Illustration 3: (Health benefits

(sweating
of ginger, n.d.)

toxins

through

skin),

&

stimulative

(speeding physiological systems). No one is quite sure how ginger is able to do

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all these things, but they do know that it can do all of the above and more. For
example, the burning sensation related to eating raw ginger-root comes from
zigibain (an enzyme that acts much like capsaicin). This is the chemical which
improves digestion & kills parasites and their eggs, while stimulating good
bacteria to remove of the rest of the prokaryotic bacteria (Davies & Ryan, 2000).
Another plant oil used in this experiment is capsaicin, the chemical that
makes up the hot parts of chili peppers.
compound,
substance,

which

in

its

This chemical is a water-insoluble

pure

can be measured at 16

million scoville heat units.

The

burning sensation often associated

with

capsaicin

interaction with
converted

to

is

caused

by

ATP, which is then

heat

upon

contact

(What makes peppers hot may also

be cool for what ails you, 2003,
[online]).

Capsaicin is said to be

tasteless and odorless in itself, while

the scent associated with it comes
from the pepper it is derived from. It
was discovered in 1816, with the
initial

exploration

Bucholtz.

done

by

P.A.

The first attempt to put

Illustration 4: (Bourbonfeet, 2010, [online])

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the varying heat levels into a form of measurement done by Wilbur Scoville in
1912. He had a group of volunteer tasters to test the different peppers, and
upon experimentation, he would spray sugar water into their mouth until the
burning sensation had dissipated. Once this had happened, he would record the
amount of sprays; making the unit of measurement (What is Caspacin? What
are Scoville Heat Units?, 2006, [online]).
Capsaicin is produced as a crystal by glands at the junction of the
pepper's placenta and pod walls which can be broken down by fats and lipids
(What makes peppers hot may also be cool for what ails you, 2003, [online]).
Because of its unique process of decomposition and formation, it is a potent
chemical in the use of circulation increase, a digestion stimulant, and the relief
of arthritis pains (Pepper, 1983). It is also used to regulate blood flow, reducing
chances of a heart attack because of its high quantities of calcium and vitamin C
(Nearman, n.d.). With a rating of 855,000 SHU, some say that the Jaga Jokia
pepper, found at a mountain's base in India, is the world's hottest pepper. The
heat of capsaicin packed in this pepper can become addictive because of the
endorphins (feel good hormones) released when it is eaten (What is capsaicin?
What are scoville heat units?, 2006, [online]).
Horseradish is a common plant native to north America most commonly
known in Passover as the bitter herb. When horseradish is processed, vinegar
may be added for a milder flavor (What makes horseradish hot?, n.d.). Though
if the myosinase is heated or starved of water, it will not activate the hot
burning sensation often associated with cruciferous plants. But glucosinolates

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can still be broken down through digestion. The chemical isothiocyanate that is
derived from the whole, raw horseradish root has been recently connected with
cancer-prevention in a study done by Jane Higdon (2005). Isothiocyanates are in
most plants in the mustard family, which studies have shown have a cancerpreventative property (Higdon & Drake, 2003). A type of this chemical called
SFU is an inducer for the phase II enzymes which protect DNA from cancercausing carcinogens.

One of the aspects of this study tested levels of

isothiocyanates in subjects bodies. The studies came back saying that people
(on average) who had cancer also had low, if any, levels of this isothiocyanates.
Some of the best sources for the chemical occur in
horseradish, brussel sprouts, mustard greens, kale and
broccoli (Higdon & Drake, 2008). Less commonly, it has been
used medicinally for fevers, bronchitis, relieving sinuses and
other antibiotic purposes (Foster & Duke, 1990).
Illustration 5:
(Molecule of the
In February of 1995, there was a report of a woman
week- allyl iscyanate,
2007)
spilled 1.5 quarts of highly concentrated isothiocyanate for
making a potent horseradish sauce. When the police arrived, they found that
they had to call a hazardous waste team

to clean up the mess.

A lead

firefighter said that horseradish can be very toxic if it were to come in contact
with a person's skin or eyes (New horseradish variety: Hazardous materials-unit
hot, 1995).

So maybe isothiocyanate will also act like an antibiotic with the

given E. coli, forcing it to die.

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Materials:

4 petri dishes (with agar solution on bottom dish)

16 sterile disks

1 mL of ginger

1 mL of horseradish

1 mL ofTabasco sauce

water

forceps

glass bowl

alcohol

latex gloves (or disposable)

5 pipettes

glass L

Sterno fuel

matches

1 work area

sink

pencil/ pen

paper

1 permanent marker

1 pair of safety goggles

sand

1 metric ruler

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Procedure:

1. Put on gloves and safety goggles

2. Using sterile technique, remove inner cryovial from plastic vial
3. Add 0.5 mL liquid media with a sterile pipette to the vial
4. Allow the E. coli to sit for 20 seconds to allow it to rehydrate
5. Pipette E. coli culture up and down a few times ensure it is well mixed
6. Dip a cotton swab into the E. coli culture
7. Swiping back and forth, spread the culture sample in a tube of agar
8. Replace cap on cryovial
9. Put cryovial in tube rack and test tube of agar (on its side) in incubator
10. After at least three days, it is ready for use
11. Clear work area
12. Clean work surface with disinfecting spray
13. Bring all the materials to the newly clean work area
14. Pour alcohol into glass bowl & open lid to jar of sand (ready to put fire out)
15. Draw dividing lines on the bottom petri dish (with permanent marker), into
quarters, and label cayenne, horseradish, control & ginger, then number each
quadrant 1, 2, 3, or 4
16. Take off the lid of the Sterno fuel and light it (with matches)
17. Put on gloves
18. Open the lid of one of the petri dishes with agar broth
19. Mix the tube of E. coli and put it on the test tube rack

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20. Suck up E. coli with dropper and drop 3 drops into petri dish (side with agar)
21. Dip glass L into alcohol and light with fire (from Sterno fuel) for 5 sec. to
sanitize
22. Allow glass L to cool for 20 seconds
23. Scrape glass L back and forth, then side to side and then around in the petri
dish (to spread E. coli)
24. Follow steps 18-23 of the next 3 petri dishes
25. Crush about one mL of horseradish in mortar and pestle with 3 mL of water (in
pipette)
26. Place solution (with new pipette or sterilized forceps) into the center of the spot
plate and mix with one (1) or two (2) drops of water
27. Soak 4 sterile disks into the solution
28. Place one sterile disk with forceps (after sanitation like that of the glass tube) into
each quadrant (of 1 petri dish)
29. Put the cover on the petri dish.
30. Wash mortar and pestle in sink under warm water
31. Follow steps 25-30 with 1 mL of ginger, & water (control)
32. Put a couple drops of Tabasco sauce into the spot plate
33. Follow steps 27-29 with the Tabasco sauce
34. Place all petri dishes in incubator
35. Measure the diameters of E. coli dye-off (with metric ruler) after 24 hours and
again after 6 days of being in an incubator at 37 C
36. After examination of results, sanitize with bleach and properly dispose
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Variables:

Independent Variable: Plant oils (ginger, horseradish & cayenne)

Dependant Variable: E. coli Reaction
Control: Petri dish with water disks
Constants:
-Experimenter-- throughout the entire experiment, I was the only person
putting together the experiment and measuring diameter
-Glass L-- the same piece was used to spread the E. coli around in the
petri dish, with similar motions
-Alcohol-- the substance spread on utensils and burned for sanitation was
the same through the experiment so everything was burned similarly
-Sterno burner-- the same flame was used for sanitation of the utensils
because it would produce the same heat throughout sanitation
-Incubator-- the same incubator stayed at the same temperature
throughout the six days between data analysis
-E. coli-- samples from the petri dishes were from the same test tube of the
bacteria, leaving the same type for experimentation
-Forceps-- were used for dipping the sterilized disks into the substances,
and placing them onto petri dish. The same ones were used each time to
avoid contamination.
-Incubator-- was set to the same temperature for the whole 6 days that the
E. coli was in there, and it was the same one
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Results:
Quadrant
1
2
3
4
1
2
3
4

Diameter of
Cayenne/ Tabasco
7mm
7mm
7mm
7mm
8mm
9mm
9mm
8mm

E. coli Dye-off
Ginger
Horseradish
7mm
0mm
7mm
0mm
7mm
0mm
7mm
0mm
7mm
0mm
32mm
0mm
15mm
0mm
36mm
0mm

Control
0mm
0mm
0mm
0mm
0mm
0mm
0mm
0mm

Average Diameter of E. coli Dye-off

Day Tabasco (Cayenne)
Ginger
Horseradish
Control
1 7mm
7mm
0mm
0mm
6 8.5mm
22.5mm
0mm
0mm

Average Diameter of E. coli Dye-off

Diameter of dye-off (mm)

25

20

15

Tabasco (Cayenne)
Ginger
Horseradish
Control

10

0
1

Time (Days)

Data Analysis:
With the data found, it is safe to say that both the Tabasco sauce and ginger were
effective in their purpose to kill E. coli. On the other hand, the horseradish and water (control)
had little to no effect on the growth of E. coli.

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Illustration 6: Horseradish result

Illustration 7: Tabasco result

Illustration 8: Ginger result

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Experimental Error:

In performing the experiment testing the effect of various plant oils on E. coli
specimens. Some of the possible errors were: glass L didn't cool for exactly 20
seconds, containment of contaminated samples, differing sanitation times (under fire),
and inaccurate methods of measurement. A prominent contamination problem that
arose was in the horseradish petri dish. The horseradish sample obtained had likely
been sitting in a bottle (unrefrigerated) for some time. Before it was used in the
experiment, it had a distinct fermented scent to it, making fungus contamination a
plausible explanation for lack of results/ unique pattern.
There were still precautions to prevent other errors. One such is when there was
a point of pause (20 secs) between burning alcohol on glass L and spreading E. coli.
Others were the multiple samplings to provide extra data, disinfecting the work surface
and utensils, wearing gloves (to avoid contamination) and using the same constants.
The quadrants proved an effective way to have multiple samples (avoiding inaccurate
and incorrect data).

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Conclusion:

There is a natural way to kill E. coli (with plant oils). If E. coli is negatively
affected by plant oils, then it may be susceptible to destruction.
After preparing the E. coli for use, the sample was used multiple times as a
testing basis for the independent variables. The plant oils were soaked into disks and
each placed into one (1) quadrant of the petri dishes. The results found show that
ginger and Tabasco were effective in destroying the E. coli samples, while the control
and horseradish showed none to little result.
The data proves that if E. coli and plant oils react, the bacteria may be
susceptible to destruction. Since only two of the three oils tested had any direct
reaction on E. coli, it is safe to say that some plant oils can kill E. coli, but not all. It
appears as if there are some natural ways to slow down, if not stop an E. coli bacterial
infection. These conclusions may lead one to find a drug containing these plant
chemicals for people infected with this bacteria.

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References
Bourbonfeet. (2010, December 3). Ghost pepper sauce and the quest for the
perfect chip [Web log post]. Retrieved from Vegan Debauchery:
http://vegandebauchery.blogspot.com/2010_12_01_archive.html
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Medical Center website: http://www.umm.edu/altmed/articles/ginger000246.htm

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House Heaven website: http://www.gingerbread-house-heaven.com/healthbenefits-of-ginger.html
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