Alternative Medicine Review Volume 14, Number 4 2009

Original
Research

Essential Oils in the Treatment

of Intestinal Dysbiosis: A
Preliminary in vitro Study
Jason A. Hawrelak, PhD, BNat(Hons); Trudi Cattley, BSci;
Stephen P. Myers, PhD, BMed, ND

detrimental impact on beneficial members of the GIT microflora.

Abstract
INTRODUCTION: Dysbiosis is associated with a number of
gastrointestinal and systemic disorders. There is a need for
selectively acting antimicrobial agents capable of inhibiting
the growth of potentially pathogenic microorganisms, or those
found to be out of balance, while not negatively impacting the
bulk gastrointestinal tract microflora. OBJECTIVE: The purpose
of this in vitro study is to examine the potential of a selection
of essential oils as agents to treat dysbiosis. MATERIALS AND
METHODS: Eight essential oils were examined using the agar
dilution method, including Carum carvi, Citrus aurantium var.
amara, Foeniculum vulgare dulce, Illicium verum, Lavandula
angustifolia, Mentha arvensis, Mentha x piperita, and
Trachyspermum copticum. Doubling dilutions of the essential
oils were tested against 12 species of intestinal bacteria, which
represent the major genera found in the human gastrointestinal
tract (GIT). RESULTS: Carum carvi, Lavandula angustifolia,
Trachyspermum copticum, and Citrus aurantium var. amara
essential oils displayed the greatest degree of selectivity,
inhibiting the growth of potential pathogens at concentrations
that had no effect on the beneficial bacteria examined.
CONCLUSION: The most promising essential oils for the treatment
of intestinal dysbiosis are Carum carvi, Lavandula angustifolia,
Trachyspermum copticum, and Citrus aurantium var. amara. The
herbs from which these oils are derived have long been used
in the treatment of gastrointestinal symptoms and the in vitro
results of this study suggest that their ingestion will have little
More research is needed, however, to investigate tolerability and
safety concerns, and verify the selective action of these agents.
(Altern Med Rev 2009;14(4):380-384)
Introduction
Intestinal dysbiosis has been defined as quali-
tative and quantitative changes in the gastrointestinal
flora, their metabolic activities, and/or their local dis-
tribution that produces harmful effects on the host.1
Dysbiosis has been associated with a number of condi-
tions, including atopic eczema,2,3 rheumatoid arthritis,4
inflammatory bowel disease,5,6 and irritable bowel syn-
drome (IBS).7-9
Evidence suggests a possible etiological role
for dysbiosis in IBS, including epidemiological studies
that have found a significantly increased risk of IBS fol-
lowing antibiotic use10 and bacterial gastroenteritis.11-13
Other evidence comes from colonic fermentation stud-
ies, which have found patients with IBS produce sig-
nificantly greater amounts of colonic hydrogen than
healthy controls14 and have altered fecal short-chain
fatty acid profiles.15
Jason A Hawrelak, PhD(SCU), BNat(Hons) – School of Health & Human
Sciences, Southern Cross University; Goulds Naturopathica
Correspondence address: 73 Liverpool St, Hobart TAS Australia 7000
Email: drjah13@yahoo.com
 
Trudi Cattley, BSci – Australian Centre for Complementary Medicine Education
and Research, a joint venture of the University of Queensland and Southern
Cross University
Stephen P. Myers, PhD(UON), BMed, ND – School of Health & Human Sciences,
Southern Cross University; NatMed-Research

Page 380

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Alternative Medicine Review Volume 14, Number 4 2009
There is also direct evidence that the gastrointes-
tinal tract (GIT) microflora of IBS patients differs from
that of healthy individuals. An older study found IBS pa-
tients have significantly fewer coliform bacteria, lactoba-
cilli, and bifidobacteria than controls.7 These findings are
supported by more recent studies that found lower fecal
concentrations of bifidobacteria in IBS patients, as well as
lower levels of lactobacilli in diarrhea-predominant IBS
patients.8
Whether this dysbiosis plays a role in the symp-
tomatology of IBS has not been conclusively proven.
However, the efficacy of probiotic agents in treating this
condition16-19 in combination with evidence outlined
above suggests a possible etiological role.
Thus, there is a need for selectively acting anti-
microbial agents capable of inhibiting the growth of po-
tentially pathogenic microorganisms, or those found to
be out of balance, while not negatively impacting the bulk
GIT microflora. In addition, since such agents may be
prescribed concurrent with probiotics, it is beneficial that
the antimicrobial agent not interfere with the growth of
the supplemented probiotic organisms (e.g., lactobacilli
and bifidobacteria).
Objective
The objective of this study is to examine the po-
tential of a selection of essential oils as agents to treat
intestinal dysbiosis. The essential oils investigated were
chosen from carminative herbs traditionally used in the
treatment of gastrointestinal disorders, including Carum
carvi (caraway), Citrus aurantium var. amara (bitter or-
ange), Foeniculum vulgare dulce (sweet fennel), Illicium
verum (star anise), Lavandula angustifolia (lavender),
Mentha arvensis ( Japanese peppermint), Mentha x piper-
ita (peppermint), and Trachyspermum copticum (ajowan).
Materials and Methods
Essential Oils
Pure essential oils were purchased from two
sources: New Directions (Sydney, NSW, Australia) and
Sydney Essential Oil Company (Sydney, NSW, Australia).
The essential oils purchased from New Directions includ-
ed Carum carvi, Foeniculum vulgare dulce, Illicium verum,
Mentha x piperita, and Trachyspermum copticum. Mentha
arvensis, Lavandula angustifolia, and Citrus ­aurantium var.
amara were sourced from Sydney Essential Oil Company.
Copyright © 2009 Thorne Research, Inc. All Rights Reserved. No Reprint Without Written Permission.
Essential Oils/Dysbiosis
Organisms and Growth Conditions
Microorganisms were obtained from the Aus-
tralian Collection of Microorganisms, University of
Queensland, with the exception of Bifidobacterium bifi-
dum and Bifidobacterium longum, which were obtained
from the CSIRO Starter Culture Collection. Organ-
isms were as follows: Bacteroides fragilis ACM 4768,
Candida albicans ACM 4574, Clostridium difficile ACM
5047, Clostridium perfringens ACM 5116, Enterococcus
faecalis ACM 4769, Escherichia coli ACM 1083, Eu-
bacterium limosum ACM 383, Lactobacillus acidophilus
ACM 547, Lactobacillus plantarum ACM 96, Bifido-
bacterium bifidum CSCC 1903, Bifidobacterium longum
CSCC 5188, and Peptostreptococcus anaerobius ACM
5059. These organisms represent the major genera of
microorganisms found in the human GIT.20
Organisms were maintained on Reinforced
Clostridial Agar (Oxoid), Wilkens-Chalgren Anaerobe
Agar (Oxoid), Mueller Hinton Agar (Oxoid), or De-
Man Rogosa Sharpe Agar (Oxoid). Inoculum was pre-
pared by suspending colonies from 24-72 hour cultures
in sterile saline. Using a CrystalSpec Nephelometer™
(Becton Dickinson & Company, Maryland, USA) sus-
pensions were standardized to a 0.5 McFarland stan-
dard, giving ~108 colony forming units (CFU) per mL
for the bacteria and 107 CFU per mL for Candida albi-
cans. Aerobic bacteria were diluted 1:10 in saline prior
to inoculation.
Minimum Inhibitory Concentration
(MIC) Determination
MICs were determined by agar dilution us-
ing Mueller Hinton Agar for anaerobic organisms,
Wilkens-Chalgren Anaerobe Agar for all anaerobes
except the two Lactobacillus species, which were grown
on DeMan Rogosa Sharpe Agar. A series of twofold
dilutions of each essential oil (from 2.0-0.004 percent
volume per volume [v/v]) was prepared and placed
in sterile Petri dishes. Each dilution was placed into
three Petri dishes and one of three agars was added to
each plate and mixed thoroughly. Tween-20 (Sigma)
was incorporated into the agar at a concentration of
0.5 percent (v/v) to enhance solubility. Clindamycin,
­neomycin, ­ampicillin, and ketoconazole (in doubling
dilutions from 64 to 0.05 μg/mL) were used as positive
controls, while dimethylsulfoxide (DMSO), Tween-20,
Page 381
 Alternative Medicine Review Volume 14, Number 4 2009

Original Research

and plain agars were used as negative con-

anaerobius trols. Plates were dried at room tempera-

2.2
4.5

4.5
4.5

2.2

1.1
1.1

0.275
Peptostreptococcus
ture prior to inoculation.
Plates were inoculated with 1-2
µL spots containing approximately 105

4.5
>4.5

>4.5
>4.5

2.2

1.1
1.1

2.2
Lactobacillus plantarum
CFU for the anaerobic bacteria and 104
CFU for the aerobic bacteria and C. albi-
4.5
>4.5

4.5
4.5

2.2

0.55
0.55

2.2
cans using a multipoint replicator (Mast
Lactobacillus acidophilus

Laboratories Ltd, Liverpool, UK). Aerobic

Table 1. The Effects of Selected Essential Oils on Common Members of Human Gastrointestinal Tract Microflora

organisms were incubated aerobically for

4.5
>4.5

>4.5
>4.5

2.2

1.1
1.1

0.55
Eubacterium limosum
20-24 hours at 35°C; anaerobic organisms
were incubated anaerobically for 48 hours
at 35°C. Minimum inhibitory concentra-
4.5
>4.5

>4.5
>4.5

2.2

0.55
2.2

0.55
Escherichia coli
tions were determined after the incubation

Results were determined as minimum inhibitory concentrations (% v/v); – = bacterial growth failure in this experiment
periods. The MIC was defined as the low-
1.1
>4.5

>4.5
4.5

>4.5

4.5
0.55

0.275
est concentration of essential oil that com-
Enterococcus faecalis

pletely inhibited the growth of the organ-

ism in question.21 The presence of a single
0.275
1.1

0.55
0.275

1.1

0.55
0.275

0.13
Clostridium perfringens
colony or a thin haze within the area of the
inoculated spot was disregarded.
0.275

0.275
0.55

2.2

0.275

0.13
_

Clostridium difficile
Results
Minimum Inhibitory
0.55
4.5

2.2
1.1

0.55

0.55
0.55

0.13

Candida albicans

Concentrations
The MIC assay results of the nine
2.2
4.5

0.275
0.55

1.1

0.55
0.275

0.275

Bifidobacterium longum
essential oils are presented in Table 1. All
essential oils tested displayed significant
antimicrobial activity. The most potent es-
2.2
>4.5

1.1
2.2

2.2

0.55
0.55

0.275

Bifidobacterium bifidum
sential oil was Trachyspermum copticum,
which inhibited the growth of all microor-
0.55
1.1

1.1
2.2

0.55

0.55
0.55

0.13

ganisms at a concentration of <2.2 percent.

Bacteroides fragilis

The most selectively acting oils were Ca-

rum carvi, Lavandula angustifolia, and Tra-
Japanese peppermint

chyspermum copticum, which inhibited the

growth of potentially pathogenic organ-
Common Name

isms such as Bacteroides fragilis, Candida

Sweet fennel
Bitter orange

Peppermint
Star anise

Lavender

albicans, and Clostridium spp., at concen-

Caraway

Ajowan

trations that had no impact on either spe-

cies of lactobacilli or bifidobacteria or the
majority of other colonic organisms. Citrus
Citrus aurantium var. amara

Trachyspermum copticum
Foeniculum vulgare dulce

aurantium var. amara displayed weaker an-

Lavandula angustifolia

timicrobial effects, but was also selective in

Mentha x piperita
Mentha arvensis
Botanical Name

activity. The other oils were not selective in

Illicium verum
Carum carvi

their activity. None of the negative controls

(DMSO, Tween-20, and plain agar) had
any impact on microbial growth.

Page 382

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Alternative Medicine Review Volume 14, Number 4 2009
Discussion
The antimicrobial properties of eight essential
oils were evaluated against common members of the hu-
man gastrointestinal tract microflora. The essential oils
were chosen based on the traditional uses of the herbs
from which the essential oils are derived. For example,
Mentha x piperita,22 Carum carvi,23 Foeniculum vulgare
dulce,24 Mentha arvensis,25 Illicium verum,26 and Lavan-
dula angustifolia27 have long been utilized as carmina-
tives in Western herbal medicine. Citrus aurantium var.
amara has a long history of use in traditional Chinese
medicine for gastrointestinal antispasmodic and car-
minative activities,28 and Trachyspermum copticum has
been used in Ayurvedic medicine to relieve colic, flatu-
lence, diarrhea, and dyspepsia.29
The most selectively acting oils were Carum
carvi, Lavandula angustifolia, and Trachyspermum copti-
cum, which at one concentration inhibited the growth
of a number of potentially pathogenic microorganisms
(Candida albicans, Clostridium spp., Bacteroides fragilis),
while having no impact on the four species of beneficial
microbes examined. Citrus aurantium var. amara essen-
tial oil was also selective in its activity. At concentrations
that inhibited the growth of Bacteroides fragilis and Clos-
tridium perfringens, no other species of bacteria or fungi
was affected. Hence, these oils appear to have the most
potential in the treatment of dysbiosis, where their use
could help balance the GIT microflora.
IBS patients have been found to have lower
fecal counts of lactobacilli, bifidobacteria, and coliform
bacteria.7-9 The results of this study suggest that Carum
carvi, Lavandula angustifolia, Trachyspermum copticum,
and Citrus aurantium var. amara essential oils could be
used in the treatment of IBS without negative ramifica-
tions on already disordered GIT microflora.
Other extracts were equally effective in killing
both beneficial and potentially pathogenic members of
the GIT flora, including Mentha x piperita, Foeniculum
vulgare dulce, Mentha arvensis, and Illicium verum essen-
tial oils. Foeniculum vulgare dulce and Illicium verum es-
sential oils were, however, less active toward lactobacilli
than bifidobacteria or the potentially pathogenic organ-
isms. Nonetheless, in concentrations that inhibited
the growth of potentially pathogenic microbes, some
­beneficial bacteria were also inhibited.
Copyright © 2009 Thorne Research, Inc. All Rights Reserved. No Reprint Without Written Permission.
Essential Oils/Dysbiosis
Of these agents, only Mentha x piperita is com-
monly prescribed, due to its demonstrated efficacy in
IBS.30-32 The study results suggest that the ingestion
of M. x piperita essential oil may inhibit the growth of
some common members of the GIT microflora. Until
more research is conducted ascertaining the in vivo ef-
fects of M. x piperita essential oil on GIT microflora,
it is prudent to prescribe a probiotic agent (containing
both bifidobacteria and lactobacilli) concurrently with
M. x piperita essential oil.
Generalization of these results to in vivo situ-
ations is limited, however, by the nature of the study
design. It is unknown what impact the processes of
digestion and absorption will have on an essential oil’s
antimicrobial activity. Thus, the results of this in vitro
experiment need to be interpreted cautiously and seen
as solely preliminary. In vivo studies are needed to verify
the selectivity of action displayed by these essential oils,
as well as to address tolerability and safety concerns.
Future in vitro studies should take into account
other common members of the GIT flora, such as Ru-
minococcus spp., Streptococcus spp., Peptococcus spp.,
Actinomyces spp., and Fusobacterium spp.,20 as well as
gas-producing microbes like methanogens and sulfate-
reducing bacteria.33 The effects of carminatives on these
latter two groups of bacteria would be particularly in-
teresting. However, the results would still be prelimi-
nary and would not provide definitive evidence of in
vivo effectiveness. Definitive answers await randomized,
double-blind, placebo-controlled human trials utilizing
the “gold standard” of microflora assessment techniques
– 16S ribosomal RNA sequencing – to accurately de-
lineate changes in the GIT microflora after ingestion of
these essential oils.34
Conclusion
The most promising essential oils for the treat-
ment of intestinal dysbiosis appear to be Carum carvi,
Lavandula angustifolia, Trachyspermum copticum, and
Citrus aurantium var. amara. The herbs from which these
oils are derived have long been used in the treatment
of gastrointestinal symptoms and these in vitro results
suggest that their ingestion will have little detrimental
impact on beneficial GIT microflora. More research is
needed to investigate tolerability, safety ­­concerns, and
verification of selectivity.
Page 383

Original Research
References
1. Hawrelak JA, Myers SP. The causes of intestinal
dysbiosis: a review. Altern Med Rev 2004;9:180-197.
2. Bjorksten B, Sepp E, Julge K, et al. Allergy
development and the intestinal microflora during the
first year of life. J Allergy Clin Immunol 2001;108:516-
520.
3. Kalliomaki M, Kirjavainen P, Eerola E, et al. Distinct
patterns of neonatal gut microflora in infants in whom
atopy was and was not developing. J Allergy Clin
Immunol 2001;107:129-134.
4. Peltonen R, Nenonen M, Helve T, et al. Faecal
microbial flora and disease activity in rheumatoid
arthritis during a vegan diet. Br J Rheumatol
1997;36:64-68.
5. Onderdonk AB. Role of the intestinal microflora in
ulcerative colitis. In: Hentges DJ, ed. Human Intestinal
Microflora in Health and Disease. New York, NY:
Academic Press; 1983:481-493.
6. Linskens RK, Huijsdens XW, Savelkoul PH, et al.
The bacterial flora in inflammatory bowel disease:
current insights in pathogenesis and the influence of
antibiotics and probiotics. Scand J Gastroenterol Suppl
2001;234:29-40.
7. Balsari A, Ceccarelli A, Dubini F, et al. The fecal
microbial population in the irritable bowel syndrome.
Microbiologica 1982;5:185-194.
8. Si JM, Yu YC, Fan YJ, Chen SJ. Intestinal
microecology and quality of life in irritable
bowel syndrome patients. World J Gastroenterol
2004;10:1802-1805.
9. Malinen E, Rinttila T, Kajander K, et al. Analysis
of the fecal microbiota of irritable bowel syndrome
patients and healthy controls with real-time PCR. Am
J Gastroenterol 2005;100:373-382.
10. Mendall MA, Kumar D. Antibiotic use, childhood
affluence and irritable bowel syndrome (IBS). Eur J
Gastroenterol Hepatol 1998;10:59-62.
11. Gwee KA, Leong YL, Graham C, et al. The role of
psychological and biological factors in postinfective
gut dysfunction. Gut 1999;44:400-406.
12. Rodriguez LA, Ruigomez A. Increased risk of irritable
bowel syndrome after bacterial gastroenteritis: cohort
study. BMJ 1999;318:565-566.
13. Alun Jones V, Wilson AJ, Hunter JO, et al. The
aetiological role of antibiotic prophylaxis with
hysterectomy in irritable bowel syndrome. J Obstet
Gynaecol 1984;5:S22-S23.
14. King TS, Elia M, Hunter JO. Abnormal colonic
fermentation in irritable bowel syndrome. Lancet
1998;352:1187-1189.
15. Treem WR, Ahsan N, Kastoff G, Hyams JS. Fecal
short-chain fatty acids in patients with diarrhea-
predominant irritable bowel syndrome: in vitro studies
of carbohydrate fermentation. J Pediatr Gastroenterol
Nutr 1996;23:280-286.
16. Halpern GM, Prindiville T, Blankenburg M, et al.
Treatment of irritable bowel syndrome with Lacteol
Fort: a randomized, double-blind, cross-over trial. Am
J Gastroenterol 1996;91:1579-1585.
Page 384
Copyright © 2009 Thorne Research, Inc. All Rights Reserved. No Reprint Without Written Permission.
Alternative Medicine Review Volume 14, Number 4 2009
17. Nobaek S, Johansson ML, Molin G, et al. Alteration
of intestinal microflora is associated with reduction in
abdominal bloating and pain in patients with irritable
bowel syndrome. Am J Gastroenterol 2000;95:1231-
1238.
18. Niedzielin K, Kordecki H, Birkenfeld B. A controlled,
double-blind, randomized study on the efficacy
of Lactobacillus plantarum 299V in patients with
irritable bowel syndrome. Eur J Gastroenterol Hepatol
2001;13:1143-1147.
19. Saggioro A. Probiotics in the treatment of
irritable bowel syndrome. J Clin Gastroenterol
2004;38:S104-S106.
20. Conway PL. Microbial ecology of the human large
intestine. In: Gibson GR, Macfarlane GT, eds. Human
Colonic Bacteria: Role in Nutrition, Physiology, and
Pathology. Boca Raton, FL: CRC Press; 1995:1-24.
21. European Committee for Antimicrobial Susceptibility
Testing (EUCAST) of the European Society of
Clinical Microbiology and Infectious Diseases
(ESCMID). EUCAST Definitive Document E.DEF
3.1, June 2000: Determination of minimum inhibitory
concentrations (MICs) of antibacterial agents by agar
dilution. Clin Microbiol Infect 2000;6:509-515.
22. Mills S, Bone K. Principles and Practice of
Phytotherapy. Edinburgh, UK: Churchill Livingstone;
2000:507-514.
23. van Wyk BE, Wink M. Medicinal Plants of the World.
Pretoria, South Africia: Briza Publications; 2004:82.
24. van Wyk BE, Wink M. Medicinal Plants of the World.
Pretoria, South Africa: Briza Publications; 2004:145.
25. van Wyk BE, Wink M. Medicinal Plants of the World.
Pretoria, South Africa: Briza Publications; 2004:205.
26. van Wyk BE, Wink M. Medicinal Plants of the World.
Pretoria, South Africa: Briza Publications; 2004:180.
27. Bone K. A Clinical Guide to Blending Liquid Herbs.
Edinburgh, UK: Churchill Livingstone; 2003:304-
307.
28. Bensky D, Clavey S, Stoger E, Gamble A. Chinese
Herbal Medicine Materia Medica, 3rd ed. Seattle, WA:
Eastland Press; 2004:510-516.
29. Williamson EM. Major Herbs of Ayurveda.
Edinburgh, UK: Churchill Livingstone; 2002:306-
310.
30. Liu JH, Chen GH, Yeh HZ, et al. Enteric-coated
peppermint-oil capsules in the treatment of irritable
bowel syndrome: a prospective, randomized trial. J
Gastroenterol 1997;32:765-768.
31. Dew MJ, Evans BK, Rhodes J. Peppermint oil for the
irritable bowel syndrome: a multicentre trial. Br J Clin
Pract 1984;38:394,398.
32. Kline RM, Kline JJ, Di Palma J, Barbero GJ. Enteric-
coated, pH-dependent peppermint oil capsules for the
treatment of irritable bowel syndrome in children. J
Pediatr 2001;138:125-128.
33. Gibson GR. Dietary modulation of the human
gut microflora using prebiotics. Br J Nutr
1998;80:S209-S212.
34. Tannock GW. Molecular assessment of intestinal
microflora. Am J Clin Nutr 2001;73:410S-414S.