Pathogenic fungi like Candida albicans, Candida tropicalis and Trichophyton mentagrophytes are commonly encountered strains associated with a wide range of conditions including scalp infection, oral thrush, skin infection, and vaginal thrush. Unlike bacterial pathogens, fungal pathogens are difficult to control. Fluconazole is a commonly administered antifungal drug. The medical fraternity has been reporting an alarming increase in the development of resistance observed amongst fungal strains to Fluconazole. The present study involves screening of essential oils for their antifungal activity against Fluconazole resistant fungi. Essential oils of Black pepper, Cardamom, Cumin, Boswellia and Patcholi were selected for the study. The results indicated that all the oils inhibited fungal strains in varying degrees of dilutions. Essential oil of Boswellia was found to be the most effective in antifungal activity against Candida tropicalis and essential oil of Cardamom against Trichophyton mentagrophytes. To assess the effect of combination of essential oils with Fluconazole, synergistic action was also studied. The results indicated that essential oil of Boswellia and Fluconazole in combination acted as the most powerful antifungal agent against Candida tropicalis even at 1:10 dilution and 100μg/ml respectively. These results lead us to believe that active components present in essential oils should be a focus area of future in vivo research, especially in conjunction with existing antifungal drugs. The molecular mechanisms, mode of action, stability, toxicity, and efficacy of the active components isolated from essential oils need to be further studied and evaluated

Kamat, Dilip

Kamat, Sheela

Rabadia, Alpa Gopal

English

Advanced Research Journals

International Journal of Phytomedicine 3 (2011) 506-510 
http://www.arjournals.org/index.php/ijpm/index 
 
Original Research Article 
           
Antifungal Activity of Essential Oils against Fluconazole Resistant Fungi 
Alpa Gopal Rabadia1*, Sheela D. Kamat1 and Dilip V. Kamat1  
 
*Corresponding author: 
Alpa Gopal Rabadia1* 
1 Department of Microbiology, 
SVKM’s Mithibai College of Arts, 
Chauhan Institute of Science, & 
Amrutben Jivanlal College of 
Commerce & Economics 
(affiliated to the University of 
Mumbai), Vile Parle (West), 
Mumbai 400056, India 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
A B S T R A C T  
Pathogenic fungi like Candida albicans, Candida tropicalis and Trichophyton 
mentagrophytes are commonly encountered strains associated with a wide 
range of conditions including scalp infection, oral thrush, skin infection, and 
vaginal thrush. Unlike bacterial pathogens, fungal pathogens are difficult to 
control. Fluconazole is a commonly administered antifungal drug. The medical 
fraternity has been reporting an alarming increase in the development of 
resistance observed amongst fungal strains to Fluconazole. The present study 
involves screening of essential oils for their antifungal activity against 
Fluconazole resistant fungi. 
Essential oils of Black pepper, Cardamom, Cumin, Boswellia and Patcholi were 
selected for the study. The results indicated that all the oils inhibited fungal 
strains in varying degrees of dilutions. Essential oil of Boswellia was found to 
be the most effective in antifungal activity against Candida tropicalis and 
essential oil of Cardamom against Trichophyton mentagrophytes. To assess 
the effect of combination of essential oils with Fluconazole, synergistic action 
was also studied. The results indicated that essential oil of Boswellia and 
Fluconazole in combination acted as the most powerful antifungal agent 
against Candida tropicalis even at 1:10 dilution and 100μg/ml respectively. 
These results lead us to believe that active components present in essential 
oils should be a focus area of future in vivo research, especially in conjunction 
with existing antifungal drugs. The molecular mechanisms, mode of action, 
stability, toxicity, and efficacy of the active components isolated from essential 
oils need to be further studied and evaluated. 
Keywords: Essential oils, antifungal drug, drug resistance, synergistic action. 
 
Introduction 
Over the last few years, the indiscriminate use of 
antibiotics has led to the problem of multi-drug 
resistance. This situation has forced researchers to 
search for new antimicrobial substances from 
medicinal plants. The curative properties of aromatic 
and medicinal plants have been recognized since 
ancient times [1]. In recent years, there has been a 
gradual revival of interest in the use of medicinal and 
aromatic plants because plant-derived drugs have 
been reported to be safe and without side-effects [2]. 
 
Down the ages, essential oils and other extracts of 
plants have evoked interest as sources of natural 
products. They have been screened for their potential 
uses as alternative remedies for the treatment of many 
infectious diseases. The World Health Organization 
(WHO) has noted that the majority of the world’s 
ISSN: 0975-0185 
 
  
This work is licensed under a Creative Commons Attribution 3.0 License.  
 Rabadia et al. International Journal of Phytomedicine 3 (2011) 506-510 
 
507 
 
population depends on traditional medicine for primary 
healthcare. The modification of antibiotic activity is 
done by compounds that enhance the activity of the 
antibiotic, and plants are a rich source of these 
compounds [3]. 
 
Essential oils (also called volatile oils) are natural, 
complex, aromatic, oily liquids obtained from plant 
materials (flowers, buds, seeds, leaves, twigs, bark, 
herbs, wood, fruits, and roots) and composed mainly of 
terpenes, in addition to some other non-terpene 
components. They can be obtained by expression, 
fermentation or extraction but the method of steam 
distillation is most commonly used for commercial 
production. They have been shown to possess 
antibacterial, antifungal, antiviral, insecticidal, and anti-
oxidant properties [4][5]. The hydrophobicity of 
essential oils enables them to partition in the lipids of 
the cell membrane and mitochondria, rendering them 
permeable and leading to leakage of cell contents [6].  
 
Concomitant with its widespread use, there have been 
increasing reports of Fluconazole resistance. While 
there is a perception that this problem is growing in 
frequency, its true reason is not known, nor is an 
optimum strategy for its prevention apparent [7]. 
Alternative therapies need to be developed to 
suppress the emergence of antifungal resistance. 
Conventional medicine applies the “silver bullet” 
method, where single target therapy is employed. The 
recent trend has been the “herbal shotgun” method like 
Ayurveda, where multitargeted approach of herbals 
and drugs is used. This can be achieved by the use of 
combinations of existing agents or the development of 
new, safer and effective agents primarily from plant 
sources which can exhibit synergy with drugs [8]. 
 
The aim of this study was to test five essential oils and 
to study the synergistic action between Fluconazole 
and essential oils for their antimicrobial activity against 
Fluconazole-resistant fungi. 
 
Materials and methods 
Test Organisms 
The fungal strains used in the study were Candida 
albicans, Candida tropicalis and Trichophyton 
mentagrophytes. They were obtained from 
Brahmakumari Hospital, Mumbai, India. The cultures 
were maintained on Sabouraud’s agar slants 
containing 50μg/ml of Fluconazole. Inocula were 
prepared by diluting overnight cultures in saline. Fresh 
subcultures were used for each experiment. 
 
Essential oils 
Essential oils of Black pepper, Cardamom, Cumin, 
Boswellia and Patcholi were used for the study. They 
were obtained from Konark Herbals and Healthcare, 
Mumbai, India. 
 
Antifungal Activity of Essential oils 
The antifungal activity of essential oils and the dilutions 
of essential oils prepared in propylene glycol were 
tested by the agar well diffusion method. The 
suspension of fungal strains in saline was swabbed on 
sterile Sabouraud’s agar plates. Wells were punched in 
the Sabouraud’s agar plate with the help of sterile cork 
borer of 9mm diameter. 100μl of different oils was then 
dispensed inside the wells with the help of a 
micropipette. The plates were then kept in the 
refrigerator for two hours and then incubated at 37ºC 
for 24 to 48 hours. The plate with T.mentagrophytes 
was incubated at room temperature for 3 days. After 
incubation, the plates were observed for the zone 
formation and the diameter (mm) of the zone was 
measured and tabulated (Tables 1 and 2). 
 
Synergistic Activity 
For synergistic activity between Fluconazole and 
essential oils, sterile strips of 5cmx1cm were used. 
The strip containing 100μg/ml of Fluconazole and the 
strip containing 100μl of 1:10 diluted essential oil were 
then placed perpendicular to each other (making a 
‘plus sign’) aseptically on a sterile Sabouraud’s agar 
plate swabbed with the fungal strain. This procedure 
was done for all the essential oils and Fluconazole 
against all the fungal strains. The plates were then 
incubated at 37ºC for 24 to 48 hours. The plates with 
T.mentagrophytes were incubated at room 
temperature for 3 days. After incubation the results 
were recorded and tabulated (Table 3) 
 
 
 Rabadia et al. International Journal of Phytomedicine 3 (2011) 506-510 
 
508 
 
Results & Discussion 
The antifungal activity of five essential oils and their 
dilutions against the three fungal cultures are shown in 
Tables 1 and 2. Cardamom, Black pepper, Cumin and 
Boswellia inhibited all the three fungal cultures even at 
1:10 dilution. Patcholi oil was ineffective as it did not 
inhibit any of the fungal cultures tested. 
 
Table 1: Antifungal activity of essential oils. 
 
 
 
Table 2:  
(1) Antifungal activity of dilutions of Cardamom oil. 
Name of the culture 
Zone of inhibition in mm 
1:2 1:4 1:6 1:8 1:10 
Candida albicans 21 19 17 15 13 
Candida tropicalis 15 14 13 12 10 
Trichophyton mentagrophytes 26 23 20 17 15 
 
(2) Antifungal activity of dilutions of Black pepper oil. 
 
(3) Antifungal activity of dilutions of Cumin oil. 
Name of the culture 
Zone of inhibition in mm 
1:2 1:4 1:6 1:8 1:10 
Candida albicans 18 16 14 12 10 
Candida tropicalis 15 13 11 9 6 
Trichophyton 
mentagrophytes 
20 17 15 13 10 
 
 Zone of inhibition in mm 
Fungal cultures Cardamom oil Black pepper oil Cumin oil Patcholi oil Boswellia oil 
Candida albicans 24 16 20 No inhibition 28 
Candida tropicalis 18 15 17 No inhibition 26 
Trichophyton 
mentagrophytes 
28 36 22 No inhibition 25 
Name of the culture Zone of inhibition in mm 
1:2 1:4 1:6 1:8 1:10 
Candida albicans 14 13 12 11 9 
Candida tropicalis 12 11 10 9 8 
Trichophyton 
mentagrophytes 
34 31 28 26 23 
 Rabadia et al. International Journal of Phytomedicine 3 (2011) 506-510 
 
509 
 
(4) Antifungal activity of dilutions of Boswellia oil. 
Name of the culture Zone of inhibition in mm 
1:2 1:4 1:6 1:8 1:10 
Candida albicans 24 23 21 20.5 19 
Candida tropicalis 24.5 22 19 17 15 
Trichophyton 
mentagrophytes 
23 19 15 14 12 
 
 
The results of synergy (Table 3) indicated that the 
combination of Fluconazole and Cardamom oil exhibited 
good antifungal activity against C.albicans. However, the 
combination of Fluconazole and Boswellia oil exhibited the 
most powerful antifungal activity against C.tropicalis even at 
1:10 dilution. No synergistic action was observed for 
Fluconazole and any of the essential oils in combination 
against T.mentagrophytes. 
 
Table 3: Results of Synergistic action. 
Name of the 
culture 
Agent(s) used 
Zone of 
Inhibition (mm) 
Candida 
albicans 
Fluconazole 7 
Cardamom oil 5 
Fluconazole + 
Cardamom oil 
14 
Candida 
tropicalis 
Fluconazole 3 
Boswellia oil 4 
Fluconazole + 
Boswellia oil 
12 
 
Essential oils have been known to possess 
antimicrobial activity by their action through the 
disruption of the cell membrane [9]. It is important to 
investigate those plants that have been used in 
traditional medicine as potential sources of novel 
antimicrobial compounds [10].  
In general, the comparison of results involving in vitro 
studies of plant extracts is problematic. First, the 
composition of plant oils and extracts is known to vary 
according to local climatic and environmental 
conditions. Secondly, the method used to assess 
antimicrobial activity and the choice of test 
organism(s), varies between publications [11][12]. For 
the abovementioned reasons, in vivo studies may be 
required to confirm the validity of the results obtained. 
Conclusion 
In order to select a compound that could act in 
synergism with a drug, it is necessary to understand 
the complete molecular mechanism of the drug action 
in the presence and absence of the natural 
compound. The problems that still need to be 
addressed are stability, selectivity and bioavailability 
of these natural products, and any adverse herb-drug 
interaction [8]. 
 
The applications of the above studies of these 
compounds in the in vivo systems or in clinical studies 
require pharmacokinetic and pharmacodynamic 
parameters. The interaction of drugs leading to either 
synergism or antagonism is not indicative of their 
pharmacodynamic efficacy [13]. In summary, this 
study confirms that many essential oils possess in 
vitro antifungal activity. However, if essential oils are 
to be used for medicinal purpose, issues of safety and 
toxicity will need to be addressed. 
 
Authors’ contributions 
AR conceptualized the research, and conducted the 
experiment and analysis. SD and DK helped in study 
design, and assisted AR with the drafting of the 
manuscript. All authors have read and approve of the 
final manuscript. 
 
Acknowledgements 
We acknowledge the help provided by Brahmakumari 
Hospital, Mumbai for providing the fungal cultures and 
the help provided by Konark Herbals and Healthcare, 
Mumbai for providing the samples of essential oils. 
This study was conducted within the institutional 
framework of the Department of Microbiology, Mithibai 
 Rabadia et al. International Journal of Phytomedicine 3 (2011) 506-510 
 
510 
 
College, and was funded by the general pool of funds 
available to doctoral students of the Department.  
 
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Antifungal activity of essential oils against Fluconazole resistant fungi

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Antifungal activity of essential oils against Fluconazole resistant fungi

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