International Journal of Cosmetic Science, 2006, 28, 125133

Evaluation of in vitro antimicrobial activity of Thai

basil oils and their micro-emulsion formulas against
Propionibacterium acnes
J. Viyoch*,, N. Pisutthanan, A. Faikreua*,, K. Nupangta*, K. Wangtorpol* and J. Ngokkuen*
*Department of Pharmaceutical Technology, Faculty of Pharmaceutical Sciences, Naresuan University, 65000
Phitsanulok, Thailand, Cosmetic and Natural Product Research Center, Health Sciences Research Institute, Naresuan
University, 65000 Phitsanulok, Thailand and Department of Pharmaceutical Chemistry and Pharmacognosy, Faculty
of Pharmaceutical Sciences, Naresuan University, 65000 Phitsanulok, Thailand

Received 2 June 2005, Accepted 15 December 2005

Keywords: micro-emulsions, Ocimum oils, Propionibacterium acnes, Thai basil oils

Synopsis
The aim of this study was to evaluate the efficacy
of Thai basil oils and their micro-emulsions, on in
vitro activity against Propionibacterium acnes. An
agar disc diffusion method was employed for
screening antimicrobial activity of the essential oils
of Ocimum basilicum L. (sweet basil), Ocimum sanctum L. (holy basil) and Ocimum americanum
L. (hoary basil) against P. acnes. Minimum inhibitory concentration (MIC) values of the basil oils
were determined using an agar dilution assay. The
obtained results indicated that the MIC values of
sweet basil and holy basil oils were 2.0% and
3.0% v/v, respectively, whereas hoary basil oil did
not show activity against P. acnes at the highest
concentration tested (5.0% v/v). Gas chromatography-mass spectrometry analysis revealed that
methyl chavicol (93.0%) was the major compound
in sweet basil oil, and eugenol (41.5%), c-caryophyllene (23.7%) and methyl eugenol (11.8%)
were major compounds in holy basil oil. Hoary
Correspondence: Jarupa Viyoch, PhD, Department of
Pharmaceutical Technology, Faculty of Pharmaceutical
Sciences, Naresuan University, 65000 Phitsanulok,
Thailand. Tel.: +66 55 261078; fax: +66 55 261057;
e-mail:jarupav@nu.ac.th, jarupav@hotmail.com
This paper was presented as a poster presentation at
Pharmaceutical Sciences World Congress (PSWC2004),
held during May 30June 3, 2004 at Kyoto International
Conference Hall, Japan.

basil oil contained high amounts of geraniol

(32.0%) and neral (27.2%) and small amounts of
methyl chavicol (0.8%). The Oil-in-water (o/w)
micro-emulsions of individual basil oils with concentrations corresponding to their MIC values
were formulated. The stable o/w micro-emulsion
system for basil oil consisted of 55.0% v/v water
phase, 10.0% v/v oil phase (2.0 or 3.0% v/v sweet
basil or 3.0% v/v holy basil oil plus 7.0% v/v isopropyl myristate), 29.2% v/v polysorbate 80 and
5.8% v/v 1,2-propylene glycol. Hydroxyethylcellulose at a concentration of 0.5% w/v was used as
thickening agent. According to the disc diffusion
assay, the formulations containing sweet basil oil
exhibited higher activity against P. acnes than
those containing holy basil oil, and the thickened
formulations tended to give a lower activity
against P. acnes than the non-thickened formulations. The prepared micro-emulsions were stable
after being tested by a heatcool cycling method
for five cycles. These findings indicate the possibility to use Thai sweet and holy basil oil in suitable
formulations for acne skin care.
Resume
Le but de cette etude est devaluer lefficacite in vitro
des huiles de basilic thailandaises et de leurs
microemulsions vis-a`-vis de Propionibacterium acnes.
On utilise une methode de diffusion sur disque en
agar pour etudier lefficacite antimicrobienne des

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Thai basil oils and skin care

huiles essentielles dOcimum basilicum L. (basilic

doux), dO. Sanctum L. (basilic sacre) et O. americanum L. (basilic velu) vis-a`-vis de P. acnes. On determine la concentration dinhibition minimum (CIM)
des huiles de basilic par le test de dilution sur agar.
Les resultats obtenus indiquent que les valeurs de
concentration dinhibition minimum du basilic
doux et du basilic sacre sont respectivement 2.0% et
3,0% en V/V, alors que le basilic velu ne presente
pas dactivite vis-a`-vis de P. acnes a` la concentration
testee la plus elevee (5,0% en V/V). Il ressort de
lanalyse de chromatographie en phase gazeuse et
de spectometrie de masse (CG-SM) que le methyl
chavicol (93%) est le constituant principal de lhuile
de basilic doux, et que leugenol (41,5%), le
gamma-caryophylle`ne (23,7%) et le methyl eugenol
(11,8%) sont les constituants principaux de lhuile
de basilic sacre. Lhuile de basilic velu contient des
quantites elevees de geraniol (32,0%) et de neral
(27,2%) et de petites quantites de methyl chavicol
(0,8%). On formule des microemulsions H/E de chaque huile de basilic aux concentrations correspondant aux valeurs de concentration dinhibition
minimum. Le syste`me de microemulsion H/E stable pour lhuile de basilic consiste en:
- 55% en V/V deau et 10% en V/V de phase huileuse (2.0 ou 3.0% en V/V dhuile de basilic
doux ou 3.0% en V/V dhuile de basilic sacre
plus 7.0% en V/V disopropyl myristate),
- 29.2% en V/V de polysorbate 80 et 5.8% en V/
V de 1,2-propyle`ne glycol.
On utilise lhydroxyethylcellulose a` la concentration de 0,5% en P/V comme agent epaississant.
Selon la methode de diffusion sur disque, les formulations contenant lhuile de basilic doux presentent
une efficacite plus elevee contre P. acnes que celles
contenant lhuile de basilic sacre; les formulations
epaissies ont tendance a` presenter une activite contre P. acnes inferieure aux formulations non epaissies. Suite a` des tests bases sur la methode des
cycles chaud/froid (5 cycles), les microemulsions
preparees se reve`lent stables. Ces resultats montrent quil est possible dutiliser les huiles de basilic
doux et sacre dorigine thalandaise dans des formulations adaptees pour traiter la peau acneique.
Introduction
Acne vulgaris is one of the most common skin diseases which primarily affects children and adolescents. The pathogenesis of acne is multifactorial,
with primary features being increased sebum

126

production during early puberty, proliferation of

Propionibacterium acnes, abnormal follicular keratinization and inflammation. Acne skin care preparations containing antibiotics, which are useful for
treatment of mild to moderate inflammatory acne,
partially exert their beneficial effects by decreasing
the follicular population of P. acnes [1, 2]. However,
the widespread use of antibiotics in dermatological
treatments has led to the development of drugresistant P. acnes strains [3, 4].
Many reports have indicated the antimicrobial
activities of essential oils isolated from several species of plant [5 8]. Most essential oils have antimicrobial activity which is considerably less potent
than that of synthetic antibiotics. However, they
have several different modes of action and therefore may be able to combat the resistant strains of
the micro-organism.
There is currently a paucity of evidence to demonstrate efficacy in acne treatment with regards to
lower side effects and the ability to combat resistant strains of micro-organisms for essential oils.
Existing data showing successful acne treatment
are limited to Melaleuca alternifolia (tea tree) oil [9
11]. However, other essential oils with an empirical reputation for antimicrobial effects may also
possess useful anti-acne property. Recently, significant efficacy has been reported in reducing the
number of acne lesions in people suffering from
acne after being treated with preparations containing Ocimum gratissinum (African basil) oil [12].
These findings raise the question if the basil oils of
different species and/or grown at other locations
may also exhibit activity against P. acnes and may
have potential for application in acne skin care. In
this study, therefore, we focused upon Thai basil
oils derived from the aerial part of Ocimum species
(Labiatae) including Ocimum basilicum L. (sweet
basil), Ocimum sanctum L. (holy basil) and Ocimum
americanum L. (hoary basil) which are popular
culinary herbs and have been used since ancient
time as traditional medicine for various topical
applications, such as a poultice or salve for insect
bites and ringworm. Although, a number of
reports have mentioned the antibacterial properties
of these basil oils [13 15], to our knowledge, the
activity of Thai basil oils against P. acnes has not
been reported previously.
To enhance the efficacy of the basil oils for acne
treatment, the design of an effective delivery vehicle
for the bactericide is important. Micro-emulsions
are preferred because they normally enhance trans-

2006 International Journal of Cosmetic Science, 28, 125133

J. Viyoch et al.

Thai basil oils and skin care

dermal drug transport by altering the skin barrier

function. Other advantages of micro-emulsions are
their high physical stability, good production possibilities and low costs of preparation. Micro-emulsions are thermodynamically stable systems and are
formed spontaneously by mixing together a water
phase, an oil phase and a surfactant/cosurfactant
mixture. Generally, the physical characteristics of
micro-emulsions are clear or slightly opalescent and
low viscosity systems. Their low viscosity can be
increased by addition of a thickening agent to the
aqueous phase. This is most useful for treatment of
acne since it improves the skin attachment and consequently enhances the accumulation of antibacterial agent to the target area.
Therefore the aim of this study was to investigate the in vitro susceptibility of P. acnes against
the basil oils of O. basilicum L., O. sanctum L. and
O. americanum L. grown in Thailand. The chemical
compositions of the individual oils were investigated to clarify the correlation between their
chemical compositions and activity against
P. acnes. Subsequently, micro-emulsions containing the selected basil oils were formulated and
their antimicrobial activities against P. acnes were
evaluated in vitro.
Materials and methods
Test basil oils
The basil oils of O. basilicum L. (sweet basil),
O. sanctum L. (holy basil) and O. americanum L.
(hoary basil) were kindly supplied by Thai-China
Flavours & Fragrances Industry Co., Ltd, Phra
Nakhon Si Ayuthaya, Thailand and complied with
the International Standard ISO 9002. These oils
were isolated from dried aerial parts of plants
grown in Thailand by hydrodistillation using a
cleavage-type apparatus.
Determination of antimicrobial activity
Propionibacterium acnes (DMST No. 14916) was
obtained from Department of Medical Sciences,
Ministry of Public Health, Bangkok, Thailand. The
agar disc diffusion method was used for the determination of antimicrobial activity of the test basil
oils and their micro-emulsions against P. acnes.
Briefly, the bacteria were placed in brain heart
broth (Merck KGaA, Darmstadt, Germany) containing 1% glucose and suspended to a density of

0.5 McFarland standard. This bacterial suspension

was used as inoculum for the disc diffusion and
agar dilution assay. A 1-mL prepared inoculum
was added to 15-mL sterile molten brain heart
agar (Merck KGaA). The resulting agar mixture
was vortexed until completely dispersed, immediately poured into sterile Petri dishes and then
allowed to solidify (approximately 30 min).
A 6-mm sterile paper disc (Whatman International
Ltd, Kent, UK) impregnated with 10 lL of the test
oils or micro-emulsions using a sterile micropipette, was placed in the middle of the inoculated
plate. After standing for 30 min, the plate was
incubated at 37
C for 72 h under anaerobic conditions using the BBL anaerobic system: gas jar, gas
pack and disposable anaerobic indicator (Oxoid,
Basingstoke, UK). After incubation, the zone of
inhibition was measured and recorded. Each test
was performed in triplicate and repeated twice.
Determination of minimum inhibitory
concentration
The agar dilution assay was used to determine the
susceptibility of P. acnes to the test oils. An individual test oil was added to 15-mL aliquots of sterile
molten agar containing 0.5% v/v polysorbate 80
(Srichand United Dispensary Co., Ltd, Bangkok,
Thailand) at the appropriate volumes to produce
the required concentration range of the test oil
(0.55.0% v/v). The inoculum (500 lL) was then
added to the resulting agar mixture and subsequently vortexed at high speed for 15 s until completely dispersed. The agar mixture was
immediately poured into sterile Petri dishes and
left to solidify for 30 min. The plates were incubated at 37
C for 72 h under anaerobic conditions
as previously described. Following the incubation
period, the plates were observed and the presence
or absence of growth was recorded. Minimum
inhibitory concentration (MIC) was defined as the
lowest concentration of the test oil where absence
of growth was recorded. Each test was performed
in triplicate and repeated twice.
Determination of composition of Thai basil oils
The chemical composition of the individual oils was
determined using a Agilent Technologies 6890 N
GC system (Agilent, Waldbronn, Germany) fitted
with a DB5-MS (5% phenyl-methylpolysiloxane)
capillary column (30 m, 0.25 mm i.d., 0.25 lm

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127

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Thai basil oils and skin care

film thickness) coupled to a mass spectrometer

5973 (Agilent) detector. The temperature program
started at 50
C with a gradient of 3
C min)1 to
150
C, held for 15 min and finally increased to
260
C. Injector and detector temperature was set at
260
C. Carrier gas was helium with a flow rate of
1.4 mL min)1. The quantification of each compound was determined as relative percentage of the
total area in the chromatogram. Peak identification
was based on comparison of mass spectra with library data (Wiley Mass Spectra library) of the gas
chromatography-mass spectrometry (GC-MS) system and by comparison of retention times with
those in the literature [16].
Preparation of the micro-emulsions
The micro-emulsions of the individual basil oils
were formulated, and the oils in concentrations
corresponding to their MIC values were incorporated into the formulations by mixing them with
isopropyl myristate (Srichand United Dispensary
Co., Ltd) (oil phase). The oil phase was added to the
mixture of the water phase consisting of polysorbate 80 (Srichand United Dispensary Co., Ltd) (nonionic surfactant), 1,2-propylene glycol (Hong Huat
Co., Ltd, Bangkok, Thailand) (cosolvent) and deionized water. The respective amounts of oil phase,
water phase, surfactant and cosolvent are shown
in Table I. Transparent micro-emulsion systems
were obtained by slight heating of the formulations. Hydroxyethylcellulose (HEC) (Fluka Chemie
AG, Buchs, Switzerland) in an amount of 0.5%w/v
was then added to the premixed micro-emulsions
to improve their viscosity. The final systems were
mixed with a magnetic stirrer until homogeneous
dispersions were obtained.

Formulations

Compositions

Base

Micro-emulsions
Sweet basil oil (% v/v)

Holy basil oil (% v/v)

Isopropyl myristate (% v/v) 10

Polysorbate 80 (% v/v)
29.2
1,2-Propylene glycol (% v/v) 5.8
Deionized water (% v/v)
55.0
BaseH
Micro-emulsions-gel like
Hydroxyethylcellulose (g)
0.5
Formulation Base (mL)
95.5
Formulation SW2 (mL)

Formulation SW3 (mL)

Formulation HL3 (mL)

SW2

SW3

HL3

2.0
3.0

3.0
8.0
7.0
7.0
29.2
29.2
29.2
5.8
5.8
5.8
55.0
55.0
55.0
SW2H SW3H HL3H
0.5

95.5

0.5

95.5

0.5

95.5

appearances of the micro-emulsions were consequently observed (stable opalescent system, opaque
or phase separation) after the end of each cycle.
Additionally, to evaluate the persistence of the
antimicrobial efficacy, the antimicrobial activity of
the micro-emulsions was determined against
P. acnes using the disc diffusion assay at the end of
the five heatcool cycles.
Statistics
Students t-test was performed on data sets. Statistical significance was set at P < 0.05.
Results and discussion
Antimicrobial activity of Thai basil oils against
P. acnes

Physical characterization and stability of the

micro-emulsions
The physical characteristics including colour, transparency and viscosity of the prepared micro-emulsions were determined. The colour and
transparency were observed by visualization. The
viscosity measurement was performed using a Digital Rheometer (Model DV-III; Brookfield, Stoughton,
MA, USA).
The stability of the prepared micro-emulsions
was determined by heatcool cycling between 45
and 4
C with a storage time at each temperature
of 24 h. Five cycles were performed and the

128

Table I The formulations of the micro-emulsion containing basil oil

As shown in Table II, all tested basil oils showed

activity against P. acnes. Propionibacterium acnes
showed highest susceptibility to sweet basil oil at a
concentration of 2.0% v/v. Holy basil oil inhibited
P. acnes at 3.0% v/v whereas hoary basil oil did
not inhibit P. acnes at the highest concentration
tested (5.0% v/v).
Chemical compositions of Thai basil oils
In the present study, the composition of the major
components of sweet basil, holy basil and hoary

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J. Viyoch et al.

Thai basil oils and skin care

Table II Antimicrobial activity of Thai basil oils against

Propionibacterium acnes

Essential oil

Ocimum basilicum L.
(sweet basil)
Ocimum sanctum L.
(holy basil)
Ocimum americanum L.
(hoary basil)

Zone of inhibition (mm)*,

mean SD

MIC
(% v/v)

35.3 1.5**

2.0

28.7 0.6

3.0

10.6 0.6

>5.0

MIC, minimum inhibitory concentration.

*Diameter of zone of inhibition including disc diameter of 6 mm.
**P < 0.01, comparing between inhibition zone of sweet basil
and holy basil oils.

basil oils as determined by GC-MS are shown in

Tables IIIV. The dominant material in sweet basil
oil was methyl chavicol (93.0%) followed by a-bergamotene (2.2%) and 1,8-cineole (1.0%). Eugenol
(41.5%) and cis-caryophyllene (c) (23.7%) were
the first and second highest percentage compound
in holy basil oil respectively. Methyl eugenol
(11.8%) was also present in this oil. For hoary
basil oil, the first and second highest percentages
of compounds present were geraniol (32.0%) and
neral (27.2%) respectively. Methyl chavicol was
not found in holy basil oil, whereas it was present
at less than 1% present in hoary basil oil.
The chemical compositions obtained from the
Thai basil oils coincide with previous reports
[17 19]. However, some reports have indicated
that linalool and eugenol are major compounds of
holy basil oil [20, 21] and linalool [13, 22] or thymol is the major compound of hoary basil oil [14].
The observed differences are probably due to differences in environment, genetic factors and nutritional status of the plants.

Table III Chemical composition of Ocimum basilicum L.

(sweet basil) oil
Peak
no.

Retention
time (min)

Compounds

Relative
area (%)

1
2
3
4
5

11.28
16.33
18.99
29.19
37.19

1,8-Cineole
Camphor
Methyl chavicol
a-Bergamotene
Epizonarene

1.0
0.8
93.0
2.2
0.8

Table IV Chemical composition of Ocimum sanctum L.

(holy basil) oil
Peak
no.

Retention
time (min)

1
2
3
4
5
6

26.04
27.38
28.08
28.46
29.81
30.76

7
8
9

31.15
31.52
31.91

Compounds

Eugenol
b-Elemene
Methyl eugenol
c-Caryophyllene
a-Humulene
Naphthalene,
1,2,3,4,4a,5,6,
7-octahydro-4a,
8-dimethyl-2-isopropenyl
b-Selinene
a-Selinene
Valencene/ecemophilene

Relative
area (%)

41.5
5.5
11.8
23.7
1.4
1.0

4.1
4.6
2.6

Table V Chemical composition of Ocimum americanum L.

(hoary basil) oil

Peak no.

Retention
time (min)

Compounds

Relative
area (%)

1
2
3
4
5
6
7
8
9
10
11
12
13

10.99
11.15
12.12
14.47
18.29
18.54
18.90
20.93
22.29
28.42
29.19
29.81
33.52

p-Cymene
Limonene
(E)-b-ocimene
a-Terpinolene
Verbenol
a-Terpineol
Methyl chavicol
Neral
Geraniol
trans-Caryophyllene
a-Bergamotene
a-Humulene
(Z)-a-bisabolene

1.8
1.0
1.0
4.5
1.4
0.7
0.8
27.2
32.0
6.2
2.7
1.4
7.7

The antimicrobial activity of sweet basil oil, with

methyl chavicol as main component, against various Gram-positive and Gram-negative bacteria, yeasts and moulds has been reported in the past [23].
In addition, the low activity of hoary basil oil
against P. acnes is most likely due to the low concentration of methyl chavicol as reported in this study.
These findings suggest that the antimicrobial activity of sweet basil oil against P. acnes may result from
the high presence of methyl chavicol. However, the
activity against P. acnes of holy basil oil was not
very different from that of sweet basil oil, although
we could not detect methyl chavicol in holy basil
oil. It is likely that the activity against P. acnes is not
limited to high amount of methyl chavicol but also

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Thai basil oils and skin care

Physical characterization and stability of the

micro-emulsions
As the oils of sweet basil and holy basil showed good
activity against P. acnes, they were selected for their
incorporation in micro-emulsions. The compositions
of the micro-emulsions are listed in Table I. The prepared micro-emulsions were transparent, of oil-inwater (o/w) type and of low viscosity. An increase
in the viscosity of the emulsions can be obtained by
adding a thickening agent that is soluble in the
external aqueous phase. This results in the change
in the viscosity of the emulsions from a liquid to a
gel-like consistency and usually provides more stable emulsions [28, 29]. In this study, the thickening
agent, HEC (high viscosity) in an amount of
0.5%w/v was dispersed into the external phase of
the micro-emulsion to increase its viscosity. The
rheology of the thickened and non-thickened microemulsions was measured using a Brookfield Digital
Rheometer. As shown in Fig. 1, non-thickened
micro-emulsions including formulations of Base,
SW2, SW3 and HL3 were low viscosity Newtonian
fluids with viscosities of about 3001000 cP.
As shown in Fig. 2, when the micro-emulsions were
thickened with 0.5% HEC w/v, the obtained microemulsions remained transparent with increased
viscosity and gel-like behaviour.

130

SW2

Viscosity (cP)

to other essential oil components. It is known that

eugenol, methyl eugenol and caryophyllene have
antimicrobial activities [24, 25]. In addition, the
activity of plant extracts with high contents of eugenol against P. acnes has been reported in the past
[26, 27]. Therefore, the major antimicrobial compound of holy basil oil against P. acnes may be eugenol. Additionally, a synergistic effect between all
components should be taken into consideration as
well. It is also interesting to note the low susceptibility of P. acnes to hoary basil oil containing high
amounts of geraniol and neral exhibited in this
study. These findings are not in agreement with the
previous study that demonstrated high susceptibility of P. acnes to lemon myrtle oil containing high
amount of geraniol (51.4%) and neral (40.9%) [7].
The presence of some compounds possibly inhibiting
the antimicrobial activity of geraniol and neral in
the essential oil of hoary basil may result in its low
activity. The results of the present study, therefore,
indicate the need for further investigations to
understand the antimicrobial effects of basil oils
against P. acnes.

2500

SW2H

2000

SW3
SW3H

1500

HL3

1000

HL3H
Base

500

BaseH

0
0

10

20
30
40
Shear rate (rpm)

50

Figure 1 Flow curves of the o/w micro-emulsions with

and without the thickening agent.

Variability of the physical stability of the formulations was visually obtained, as shown in
Table VI. The formulations of Base, BaseH, SW3,
SW3H, HL3 and HL3H were transparent o/w
micro-emulsions and did not have a tendency to
coalesce after five cycles of heatcool cycling. The
appearance of the prepared micro-emulsions including SW3H at before and after stability test is
shown in Fig. 2. Unexpectedly, the formulation of
SW2 was separated to two phases during storage
at 45
C after three cycles whereas the formulation
of SW2H exhibited opaque character after five
cycles. It is likely that the increase in the essential
oil concentration plays a positive effect on the stability of the formulation system by a possible
increase of the phase inversion temperature (PIT)
[30 33] of these systems, resulting in more stable
systems also at lower temperatures.
As mentioned above, a direct relationship exists
between the viscosity and the stability of an emulsion. Increase in the viscosity provides more stable
emulsion by reducing the flocculation rate of the
emulsion [28, 29, 34]. Therefore, the SW2H for-

SW3H :
at after stability test

SW3H :
at before stability

Figure 2 Physical appearance of some prepared microemulsions before and after stability testing for five cycles.

2006 International Journal of Cosmetic Science, 28, 125133

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Thai basil oils and skin care

Table VI Appearance of the prepared micro-emulsions

before and after stability testing for five cycles
Appearance

Formulations

Before

After

Base
BaseH
SW2

Transparent liquid
Transparent gel
Transparent liquid

SW2H

Transparent gel

SW3
SW3H
HL3
HL3H

Transparent
Transparent
Transparent
Transparent

Transparent liquid
Transparent gel
Phase separation
(after three cycles)
Opaque gel
(after five cycles)
Transparent liquid
Transparent gel
Transparent liquid
Transparent gel

liquid
gel
liquid
gel

mulation with HEC exhibited higher stability than

SW2 formulation without HEC.
Since the formulations of SW3, SW3H, HL3 and
HL3H exhibited good physical stability, these formulations were selected for determining their antimicrobial efficacy against P. acnes after the stability
tests have been performed. The in vitro antimicrobial activity of the different formulations against
P. acnes is shown in Table VII. Base and BaseH formulations showed no inhibitory effect. The inhibition zones of the pure basil oils were larger when
compared to those of the micro-emulsion because of
their higher concentration. The inhibition zones of
the formulations containing 3.0% v/v sweet basil
oil (SW3 and SW3H) showed an about four to five
times bigger diameter of the disc, whereas the inhiTable VII Antimicrobial activity of the prepared microemulsions against Propionibacterium acnes before and after
stability testing for five cycles
Zone of inhibition (mm)*,
mean SD

Formulations

Before

Base
BaseH
SW3
SW3H
HL3
HL3H

6.7
6.0
29.0
26.3
14.7
12.7

After

0.6
0.0
1.0**
2.1**
1.2
1.2

6.3
6.3
27.0
23.3
13.0
11.0

bition zones of the formulations containing 3.0%

v/v holy basil oil (HL3 and HL3H) were about two
to three times greater than the original disc diameter. These obtained results coincide with our previous experiment indicating the significantly higher
susceptibility of P. acnes to sweet basil oil when compared to susceptibility of P. acnes to holy basil oil.
The increase in viscosity by adding the thickening
agent in the external phase tended to provide smaller diameters of the inhibition zones. In fact the active antimicrobial compounds of the Thai basil oils
investigated in this study have lipophilic character
and may, therefore, be able to penetrate into deeper
layers of the skin and into the sebaceous glands
where most of the micro-organsims of P. acnes
reside. The high content of the surfactants in the
micro-emulsions may even increase this effect by
additional interaction of the surfactants with the
skin lipids and the sebum of the skin. Although the
water phase of the micro-emulsion will evaporate
after the application of the formulation to the skin,
the favourable properties of the micro-emulsion
may initiate the effect of cotransporting the lipophilic components of the basil oils into the deeper
layers of the stratum corneum and to the base of
the infected sebaceous glands. For this reason, our
prepared micro-emulsions would be effective in acne
skin care. Furthermore, the persistence of the antimicrobial efficacy of the micro-emulsions at the end
of the five heatcool cycles was observed in this
study. As shown in Table VII, the inhibition zones
of the SW3, SW3H, HL3 and HL3H micro-emulsions at before stability test were not significantly
different from those at after stability test. An example of inhibition zone, which was about four times
bigger than the original disc diameter of SW3H formulation at after stability test is shown in Fig. 3.
These findings indicate the stability of basil oil as a
function of the kind of emulsified system in which it
is vehicled.
Conclusions

0.6
0.6
1.7**
1.2**
1.0
1.0

*Diameter of zone of inhibition including disc diameter of 6 mm.

**P < 0.01, comparing between inhibition zone of SW3 and HL3,
and SW3H and HL3H formula.

All tested Thai basil oils exhibited antimicrobial

activity against P. acnes. However, P. acnes exhibited highest susceptibility to sweet basil oil with
MIC value of 2.0% v/v followed by holy basil oil
with MIC value of 3.0% v/v, whereas hoary basil
oil did not show an inhibitory effect at the highest
concentration tested (5.0% v/v). Sweet basil oil
and holy basil oil were incorporated into o/w
micro-emulsions because these formulated micro-

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J. Viyoch et al.

Thai basil oils and skin care

non-treated

SW3H

BaseH

Figure 3 Inhibition zone of some prepared micro-emulsions after stability testing for five cycles.

emulsions were expected to provide advantages

such as good storage stability, antimicrobial efficacy and non-oiliness after application to the infected skin. The micro-emulsion of 3% v/v sweet
basil or holy basil oils exhibited stable physical
characteristics and stable efficacy in antimicrobial
activity after being tested by heatcool cycling.
Our study suggests that Thai basil oils, especially
methyl chavicol-rich sweet basil oil and eugenolrich holy basil oil may be a promising source for
acne treatment. In addition, our study indicates
that the o/w micro-emulsions are suitable carrier
systems for these oils for use in acne skin care.
Acknowledgements
This work was supported by a research grant from
the Faculty of Pharmaceutical Sciences, Naresuan
University, Phitsanulok, Thailand and the National
Research Council of Thailand (NRCT). We would
like to thank Thai-China Flavours & Fragrances
Industry Co., Ltd, Thailand for supplying the
essential oils and the Department of Medical
Sciences, Ministry of Public Health, Thailand for
supplying the P. acnes strain. We also thank Prof.
Hans E. Junginger for his valuable discussion.
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