International Conference on Chemical, Environmental and Biological Sciences (CEBS-2015) March 18-19, 2015 Dubai (UAE)
In Vivo Antifungal Activity of Some Medical
Herbs and Propolis against Fungal pathogens
Associated with Ringworm
Shahin Nejat1*, Abdollah Ghasemi Pirbaluti2, Masoud Yazdani3 and Maryam Foroughi4

mentagrophytes, and microsporum gypseum (5). The use of
synthetic drugs accompany with side effects, so we decided to
study the effect of some herbal extract and propolis on
mentioned fungi.
Abstract-The present study investigated suppressor activity of the
alcoholic extract of some herbal drugs (Thymus daenesis, Satureja
bachtiarica, Althaea officinalis) and Propolis on some strains of
fungal dermatophytes (Trichophyton mentagrophytes, Trichophyton
verrucosum and Microsporum gypseum) which is an important agent
of human and animal’s pathogens. In this study, Amphotericin B was
used as positive control.
Results showed that, Propolis extract suppressed the growth of all
tested fungi with different degrees. In fact, a linear dose-dependent
dilution of the alcohol extract of Propolis against of all fungi was
obtained. Also among used herbal plants, a high dose of Satureja
bachtiarica suppressed the growth of Trichophyton mentagrophytes
and Trichophyton verrucosum. It can be concluded that instead of
chemical drugs, these natural substances can be used widely in
pharmaceutical and health care industries for control of fungal
diseases.
II. MATERIALS AND METHODS
PLANTS AND PROPOLIS MATERIALS:
A. Propolis was taken from bee hives in Chaharmahal &
Bakhtiari apiaries. Also, plants (Thymus daenensis, Satureja
bachtiarica and Althaea officinalis) were collected during
flowering stage from highlands of Chahrmal & Bakhtiari
province, Iran, and identified in Medicinal Plant Research
Center, Islamic Azad university of Shahrekord, The collected
plant materials dried in shade under room temperature
followed by grinding. Essential oil was obtained from 50g of
each plant with 600 ml distilled water by hydrodistillation
during 3 hours using a Clevenger-type.
PREPARATION OF ETHANOLIC EXTRACT:
Key Words: Antifungal, Medical Herbs, Propolis, Ring worm.
I. INTRODUCTION
I
N the recent decades, the interest in evaluating therapeutic
effects of plants and animal products has increased
dramatically [1], [2] as 80% of the world’s people rely on
complementary and alternative medicine for their health care
needs [3], [4]. Dermatophytosis is superficial fungal infection
caused by dermatophytes that invade the keratinized tissue of
humans and animals. Lesions from dermatophytosis exhibit an
inflammatory reaction induced to eliminate the invading fungi
by using the host's normal immune function [15]. Ringworm
occurs in all animal species in all countries but more
commonly where animals are accommodated in dense groups,
especially indoors. Animal susceptibility is determined largely
by immunological status, so that young animals are most
susceptible. The most common cause of dermatomycosis in
horses nad cattle, are trichophyton verrucosum, trichophyton
B. According previous reports ethanolic extract showed
more effectiveness in comparison with the plain or aqueous
extract. To prepare the ethanolic extract, 100 gram of fresh
plants and Propolis were added into 1000 ml ethanol (70%)
and kept for 24 hours. All extracts were kept at 4ºC before use.
The extracts were concentrated.
Different concentrations were made from all extracts. For
this purpose, the amount of 0.5 grams of concentrated extracts
of plants were weighed. By Sampler, amount of 1 cc (1000
micro liters) of DMSO (dimethyl sulfoxide), was added to it
and solve it well. A serial microdilution assay was used to
determine the minimum inhibitory concentration (MIC) values
for plant extracts. To apply it to measuring antifungal
activities, a slight modification was made to suit fungal growth
conditions (6). Serial dilutions were prepared as follows:
922 – 692 - 569 - :6/9 µG/ML
15 microliter of each dilution, was shed on disk sterile in
culture media containing the fungus. The laboratories were
kept for days at 10-7. Then the diameter of growth inhibition
were measured. Mediums were kept in laboratory for 7-10
days. Then diameter of growth inhibition zone were measured.
In this study, the antifungal drug, amphotericin B, was used as
a positive control. It should be noted that the fungal strains
were collected from animals referred to veterinary clinics, and
identified in the laboratory.
Shahin Nejat1* Department of Internal Medicine, Veterinary Medicine
Faculty, Shahrekord Branch, Islamic Azad University, Shahrekord, Iran
(corresponding author’s phone:00989132801364; Fax: 0098- 03833361045;
e-mail:[email protected]).
Abdollah Ghasemi Pirbaluti2 Department of Medicinal Plants Shahrekord
Branch, Islamic Azad University, Research Center of Medicinal Plants &
Ethno-veterinary,PO
Box
166,
Shahrekord,
Iran,
([email protected]).
Masoud Yazdani3, Graduated from Veterinary Medicine Faculty,
Shahrekord Branch, Islamic Azad University, Shahrekord, Iran (e-mail:
[email protected]).
Maryam Foroughi4 Graduated of Plant Pathology, Faculty of Agriculture,
Islamic Azad University, Shiraz, Iran, e.mail: [email protected].
http://dx.doi.org/10.15242/IICBE.C0315128
154
International Conference on Chemical, Environmental and Biological Sciences (CEBS-2015) March 18-19, 2015 Dubai (UAE)
STATISTICAL ANALYSIS
IV. DISCUSSION
All treatments have done triplicate and statistical analysis
were conducted using SPSS 17.0, t-student, Mann-Whitney U
tests and differences were considered significant when P <
0.05.
III. RESULTS
The mean diameters of fungal growth inhibited by different
concentrations of extracts are shown in Table 1, 2 and 3. There
published many studies suggesting that Propolis exerts a strong
anti-fungal activity and it merits further investigation as a
potentially useful agent for the treatment of dermatophytosis
[10].
Propolis is a complex mixture of several resinous substances
known as a safe natural bee product and has been used in folk
medicine. It has been shown that Propolis can be used to treat
Candida and Malassezia infections [11], [12]. Agüero et al
(2010) proved that, the most susceptible species to Propolis
were Microsporum gypseum, Trichophyton mentagrophytes,
and Trichophyton rubrum [13]. Also, Silici et al (2005)
showed that Candida albicans, C. glabrata, Trichosporon
spp., and Rhodotorula spp. were susceptible to low
concentrations of Propolis, the latter showing a higher
susceptibility [14].
After evaluating the antifungal effects of four different
concentrations of dilutions, the positive control and Propolis
produced significantly large inhibition zones for all
microorganisms. Detailed antifungal effects of extracts against
three pathogens are demonstrated in Tables 1, 2 and 3.
TABLE I
Antifungal Properties of Ethanolic Extracts at Four Different
Concentrations on Microsporum Gypseum
herbal Strains
Thymus
daenesis
Satureja
bachtiarica
Althaea
officinalis
Propolis
Amphotericin B
Diameter of inhibition zones ± Standard deviation (MM)
Conc. of
Conc. of
Conc. of
Conc. of
500
250
125
62.5
2±2a
2±2a
2±2a
2±2a
2±2a
2±2a
2±2a
2±2a
6</:;±5/97a
68/77±5/97a
2±2a
2±2a
2±2a
6:/:;±7/29b
66±6/:8b
68/:;±5/97c
66±6b
2±2a
68/77±2/9<c
68/77±7/65a
Values within each row followed by different letters are significantly
different at P<0.05.
TABLE II
Antifungal Properties of Ethanolic Extracts at Four Different
Concentrations on Trichophyton Mentagrophytes
herbal Strains
Thymus
daenesis
Satureja
bachtiarica
Althaea
officinalis
Propolis
Amphotericin
B
Diameter of inhibition zones ± Standard deviation (MM)
Conc. of
Conc. of
Conc. of
Conc. of
500
250
125
62.5
2±2a
2±2a
2±2a
2±2a
</77±2/9<a
<±5b
2±2a
2±2a
2±2a
2±2a
2±2a
2±2a
68/:;±2/9<a
5:/:;±2/9<a
6:/77±5/96b
55/:;±5/59b
68/77±6/2<a
57/77±5/59c
67±6a
55/:;±2/9<b
Values within each row followed by different letters are significantly different
at P<0.05.
TABLE III
Antifungal Properties of Ethanolic Extracts at Four Different
Concentrations on Trichophyton Verrucosum
herbal Strains
Diameter of inhibition zones ± Standard deviation (MM)
Conc. of
Conc. of
Conc. of
Conc. of
500
250
125
62.5
In this study, the maximum diameter of inhibition zone
related to the Propolis. Interestingly, in all three fungi,
Propolis has more inhibitory effect than the positive control.
Also, three plant extracts were evaluated. The genus Satureja
represents 15 species in Iran, 9 of them are endemic [7] and
Satureja bachtiarica growth on rocky mountain and it
fragrances is same thyme group plants [8]. Our result showed
that, at concentrations of 500 and 250, the ethanolic extract of
S. bachtiarica had a limited effect on Trichophyton
mentagrophytes and Trichophyton verrucosum growth. At any
concentration, Thymus daenensis and Althaea officinalis had
no effect in fungi growth. Mohammadi et al (2014) showed
that the essential oils of Thymus daenensis had the moderate
activity against some fungi (Fusarium oxysporum, Aspergillus
flavus and Alternaria alternate) in comparison with the other
plants' essential oils [9].
Conclusion
The results of this study indicate the antifungal activity of
ethanolic extract of Iranian Propolis. The strong antifungal
activity of Iranian Propolis may be due to high total phenolic
and flavonoid contents and can be used in treatment of
dermatomycosis in human and animals. There are numerous
questions yet to be answered concerning chemical
compositions and antibacterial properties of Iranian Propolis
and further research is required for clarification.
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