Growth Inhibition of Escherichia coli. by Essential Oils of Rosemary (Rosmarinus
officinalis) and Lavender (Lavandula augustifolia)
Scott Lilly and Jordan Meek
Department of Biological Sciences
Saddleback College
Mission Viejo, CA 92692
This study was undertaken to ascertain whether a statistical difference existed in
growth inhibition of Escherichia coli, E coli. between Rosemary (Rosmarinus
officinalis) and Lavender (Lavandula augustifolia) essential oils (Could be clearer,
are you testing for a statistical difference or for inhibition of growth?). We expected
statistical significance to be observed regarding growth inhibition between both
Rosemary and Lavender in addition to both groups showing significantly more
inhibition then observed in control. (Scientific documents are not suppose to be
written in first person, I would recommend you change “We” to “researchers” or
something like that, also I’m not sure exactly what you are trying to tell me, please
make this more clear) Two sample groups and one control group containing 10 agar
dishes each were plated with E coli. and three equidistantly plated chads immersed
in Lavender oil, Rosemary oil, and DI water respectively. Plates were allowed to
incubate for forty eight hours and growth inhibition was measured across the chads
in millimeters with a ruler. Average inhibition (mm) in the control group was
observed to be 0.14 ± 0.07 (± S.E.M.). Average inhibition (mm) in the Rosemary
group found to be 2.9 ± 0.41 (± S.E.M.) and in the Lavender group to be 2.5 ± 0.55
(± S.E.M.). Results demonstrated no significant difference in growth inhibition of E.
coli between the Lavender and Rosemary groups (p = 0.4205, ANOVA). As expected
however, significant differences in inhibition were observed in comparison of both
the lavender and control group (p = 0.000032, ANOVA), and the rosemary and
control group (p= 0.000302, ANOVA). Results indicate that both Rosemary and
Lavender essential oils significantly inhibit E coli. growth but do not vary
significantly from each other in terms of inhibition.
Introduction
Aromatic plants, such as Rosemary,
Lavender, Oregano, and Thyme have long been
valued for the medicinal and aromatic uses of
their essential oil extractions. In addition, a
number of studies undertaken in the last twenty
five years have looked at the antibacterial
properties these plants possess. The bird species,
blue tit (Cyanistes caerulas) has been found to
include pieces of aromatic plants amongst its
normal nest building material (Blondel et al.
2009). Upon examination of the effect of these
plants on the bacteria present on the blue tits, it
was found that the plants significantly altered the
structure of the observed bacterial communities
specifically in regards to reducing the density of
colonies among hatchlings (Blondel et al. 2009).
Variability in antibacterial activity has also been
noted in these plants due to a variance in
concentration of essential oils specifically in
regards to ice nucleation active bacteria
(Karamanoli et al., 2004). In regards to the
effects of these plants on specific bacterial
strains, the aromatic plant basil (Ocimum
basillicum) has been shown to be effective in
inhibiting the growth of E coli. (Lopez et al.,
2005).
Relatively few studies have directly
looked at the variance in effect that different
essential oil extracts from these aromatic plants
have on bacterial growth. The objective of this
study was to observe the amount of growth
inhibition of Escheridia coli. from the
application of Lavender and Rosemary essential
oils and whether the two differed significantly in
their inhibition ability. It was hypothesized that
both groups would significantly inhibit bacterial
growth and that they would significantly vary
between each other in the amount of inhibition
Although the rosemary group had the
largest average zone of inhibition, as seen in
figure 1, it was found to have no statistical
-(where is the rest of this sentence?)difference when compared to the lavender group
(p= 4.2 x 10-1 , ANOVA). However, when the
rosemary group was examined against the
control group there was a significant statistical
difference (p= 3.2 x 10-4, ANOVA). The results
were also similar when the lavender group was
compared to the control (p=3.0 x 10-4, ANOVA).
(Why not just say that both groups demonstrated
significant inhibition of growth in relation to the
control group? It seems almost like you are
reporting the results backwards)
Materials and Methods
Results
Before evaluating the data, the diameter
length of the Chad was subtracted from each
measured zone of inhibition in order to get an
accurate measurement. The average zone of
inhibition on E.coli between the three groups
was analyzed for comparison. The rosemary
group had the greatest average growth inhibition
at 2.91 mm ± .414 (± S.E.M) with the lavender
group close behind at 2.45mm ± .546 (± S.E.M).
As expected the control group had the smallest
average, .14 mm ± .070 (± S.E.M)
(Figure 1). (How did you measure each zone?
Did you include the chad itself or did you
subtract the diameter of the chad and just
report the degree of inhibition?)
3.5
3
Growth Inhibition (millimeters)
1L of agar medium (Criterion Dehydrated
Culture Media) was prepared and autoclaved for
forty five minutes. The agar was then distributed
evenly to 30 petri dishes and allowed to cool for
a period of forty minutes. Three, single hole
punch chads of filter paper were prepared for
each Petri dish for a total of 90, these were
subsequently autoclaved for forty five minutes.
20mL of Escherichia coli, E.coli. (precultured in
the microbiology lab a few days prior) was
distributed in 0.5mL increments to each dish in a
lawn spread using a sterile spreader and standard
procedure aseptic techniques, left over E. coli
was disposed of properly. The 30 dishes were
split into three groups: the control group, the
rosemary group, and the lavender group. Three
chads were placed on each dish using tweezers.
Control group chads were dipped in water,
whereas rosemary and lavender group chads
were dipped in essential oil extracts of rosemary
and lavender respectively. Oil extracts were
sterilized via boiling and standard procedure
Aseptic techniques were followed in transferring
the chads to dish. All groups were incubated for
48 hours at a temperature of 37° Celsius. Upon
removal from incubation, growth inhibition was
measured as the diameter of E. coli absence
across the center of the chads.
2.5
2
1.5
1
0.5
0
Lavender
Rosemary
Control
Figure 1 - Average growth inhibition (mm) of
E.coli of the Lavender group was 2.45 ±
.546 (± S.E.M). The rosemary group was 2.91
± .415 (± S.E.M) and the control group was 0.14
± .070 (± S.E.M)
Discussion
The results presented provide evidence
that refutes one aspect of the two part hypothesis
and supports the other. (This needs some
cleaning, I’d jump straight to saying which part
of the hypothesis was refuted and which was
supported rather than having that statement) The
average growth inhibition between the Rosemary
group and the control group was found to be
significant as was the growth inhibition between
the Lavender group and control group. This data
supports the hypothesis that these particular
aromatic plants significantly inhibit bacterial
growth and reflect similar results to the study of
E coli. inhibition by basil (Lopez et al., 2005).
No statistical difference was found in
growth inhibition between the Lavender and
Rosemary groups. This provides evidence for the
validity of the null hypothesis, that there is not a
significant difference in the E coli. growth
inhibition ability of these two plants. These
results also appear to run counter to the study
conducted by Karamanoli et al., 2004 which
found that variances in concentration of essential
oils caused significant differences in antibacterial
activity (Karamanoli et al., 2004). (Making both
formal citations in the same sentence doesn’t
seem necessary, it seems possible to cut the et al.,
2004 from the first one since you sight that in the
end of the sentence) The differing results could
potentially have been caused by human error in
this study. Namely, chads may not have been
uniformly coated in their respective oils due to a
density difference amongst the two sample
groups. (If one was more dense, when the
immersed chad was shaken to remove excess oil
prior to plating, less might have dripped off
causing the two groups not to have equal
representation). (Instead of adding a parentheses,
try to explain why in the sentence, it makes it
seem more clear) Also, while sterilizing the
essential oils, the Rosemary oil was found to
come to its boiling point faster then did the
Lavender oil which could have skewed the
results. (How could it have skewed the results,
please explain more clearly)
This experiment looked at the effects of
two of these aromatic plants on a single strain of
bacteria’s growth. Further experiments could
employ a wider range of essential oils and a
larger sample size to better ascertain if/how these
aromatic plants differ in their effect on E coli.
growth. In addition, a single essential oil could
be used and tested against a variety of bacterial
strains to see if the inhibitory effect is
widespread to limited to E coli.(You made no
mention as to why Rosemary or Lavender have
an effect on bacterial growth, What is the
chemical compound(s) responsible for their antibacterial properties?)
Literature Cited
Blondel, J., Mennerat, A., Mirleau, P., Perret, P.
“Aromatic plants in nests of the blue tit
Cyanistes caeruleus protect chicks from
bacteria.” Ocelologia (October 2009):
Vol. 161, Iss. 4; 849
Karamanoli, K., Vokou, D., Menkissoglu, U.,
Constantinidou, H.-I. “Bacterial
Colonization of Phyllosphere of
Mediterranean Aromatic Plants”.
Journal of Chemical Ecology (2004):
2035-2048
Lopez, P., Sanchez, C., Battle, R., Nerin, C.
“Solid- and vapor-phase antimicrobial
activities of six essential oils:
suscepitibility of selected foodborne
bacterial and fungal strains.” J Agric
Food Chem (August 2005);
53(17):6939-46
Picagglia, R., Maroti, M., Giovanelli, E., Deans,
S.G., Eaglesham, E. “Antibacterial and
antioxidant properties of Mediterranean
Aromatic Plants”. Industrial Crops and
Products (1993): Vol.2, Iss.1; 47-50
Svoboda, K., Hampson, J., “Bioactivity of
essential oils of selected temperate
aromatic plants : antibacterial,
antioxidant, anti-inflammatory, and
other related pharmacological
activities” Plant biology Department,
SAC Auchincruive, Ayr, Scotland, UK.,
KA6 5HW
Literature Cited
Blondel, J., Mennerat, A., Mirleau, P., Perret, P.
“Aromatic plants in nests of the blue tit
Cyanistes caeruleus protect chicks from
bacteria.” Ocelologia (October 2009): Vol.
161, Iss. 4; 849
Karamanoli, K., Vokou, D., Menkissoglu, U.,
Constantinidou, H.-I. “Bacterial
Colonization of Phyllosphere of
Mediterranean Aromatic Plants”. Journal of
Chemical Ecology (2004): 2035-2048
Lopez, P., Sanchez, C., Battle, R., Nerin, C. “Solidand vapor-phase antimicrobial activities of
six essential oils: suscepitibility of selected
foodborne bacterial and fungal strains.” J
Agric Food Chem (August 2005);
53(17):6939-46
Picagglia, R., Maroti, M., Giovanelli, E., Deans, S.G.,
Eaglesham, E. “Antibacterial and
antioxidant properties of Mediterranean
Aromatic Plants”. Industrial Crops and
Products (1993): Vol.2, Iss.1; 47-50
Svoboda, K., Hampson, J., “Bioactivity of essential
oils of selected temperate aromatic plants :
antibacterial, antioxidant, anti-inflammatory,
and other related pharmacological activities”
Plant biology Department, SAC
Auchincruive, Ayr, Scotland, UK., KA6
5HW
Review Form
Department of Biological Sciences
Saddleback College, Mission Viejo, CA 92692
Author (s): Scott Lily, Jordan Meek
Title: Growth Inhibition of Escherichia coli. by Essential Oils of Rosemary (Rosmarinus
officinalis) and Lavender (Lavandula augustifolia)
Summary
Essentially, the researchers wanted to establish that aromatic plants, in this case Rosemary and
Lavender, have anti-bacterial qualities. There has been research regarding Cyanistes caerulas, or
the blue tit, where the use aromatic plants in their nest-building. After some research, it was
discovered that nests built with aromatic plants tend to have lower bacterial counts than those
built without them. The goal of these researchers was to isolate two of these aromatic plant from
other building materials found in these bird’s nests to see if they are capable of inhibiting
bacterial growth. The experiment consisted of thirty agar nutrient plates divided into three groups,
the control which consisted of water, and two experimental groups, one containing lavender and
the other containing rosemary. The E. Coli used in the experiment was pre-cultured and spread
across all the agar plates. Then for each plate, the researchers placed three paper chads
containing water, rosemary oil or lavender oil. From there, the researchers measured the diameter
of the zone of inhibition to gauge the degree of which the groups differed in one another. The
researchers found bacterial growth is inhibited by the presence of rosemary and lavender oils,
however they do not statistically differ between each other.
General Comments
The project itself was satisfactorily designed and executed. This project was a logical next step
given the observations of the blue tit’s nest-building materials, however it doesn’t explain why
these aromatic plants have anti-bacterial qualities. Given the potential this discovery offers, it
would be the most beneficial to analyze what chemical(s) are present in these plants that inhibit
bacterial growth and also, how these birds go about selecting materials to build their nests. Given
the resources available to the researchers it is understandable that such a project isn’t feasible
however, providing some kind of chemical data seems like it would be a key concern during the
project selection and research process. However, as previously stated the technical portions of
the experiment such as the number of chads, plates and groups insure that the data they did
receive from this experiment was reasonably accurate.
Technical Criticism
This paper was a final version
This paper was a rough draft
There were quite a few times where statements needed more clarity. For example, when the
researchers first illustrate the goal of the project it is unclear whether they are testing for proof of
inhibition or a significant difference which can be inhibitory or excitatory, later in the paper the
answer becomes apparent but clarity is better achieved as early as possible. Also, when the
researchers explained how they measured the diameter of the inhibitory zone, it was unclear
whether or not the chad itself was included in the measurement, it seem more accurate if the
diameter of the chad was excluded from the data because this would more accurately emphasize
the anti-bacterial characteristics of these aromatic plants. At one point during the explanation of
the results, half of a key sentence explaining the results is missing, however this seems like more
of a computer error than human error. Other than occasional unclear statements, there seemed to
be only minor format errors. Once, the paper drifted into first person for a sentence and also
there was a small citation error where the source was cited twice in the same sentence.
Recommendation
 This paper should be published as is
 This paper should be published with revision
 This paper should not be published