In vitro anti-Candida Activity of Zataria multiflora Boiss.
ABSTRACT Zataria multiflora Boiss known as Avishan Shirazi (in Iran) is one of the valuable Iranian medicinal plants. The aim of study was to evaluate anti-Candida activity of Z. multiflora against different species of Candida in vitro. Anti-Candida activity of the aqueous, ethanolic and methanolic maceration extract of the aerial parts of Z. multiflora Boiss was studied in vitro. Anti-Candida activity against Candida species was done using serial dilutions of extracts in Sabouraud's dextrose agar. Minimal inhibitory concentration (MIC) of the methanolic and ethanolic extracts was 70.7 and 127 mg l(-1), respectively. Aqueous extract showed no remarkable activity against Candida species. We conclude that methanolic extract of the aerial parts of Z. multiflora Boiss has more anti-Candida effect at 70.7 mg l(-1) compared to ethanolic extract 127 mg l(-1). In addition, the isolates of Candida parapsilosis were more susceptible to methanolic extract than other tested species.
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ABSTRACT: Rosmarinic acid (RA) was obtained from Zataria multiflora tops' extract and its structure was confirmed by spectroscopic methods. Various in vitro cultures were established on Murashige and Skoog (MS) or Modified Tobacco (MT) medium containing growth hormones. The results indicated that cultures of Z. multiflora biosynthesize RA (55-355 mg/100 g dry wt.) and the highest accumulation were reached on MT media containing NAA 2 mg/l.Fitoterapia 07/2004; 75(3-4):315-21. · 2.23 Impact Factor
- Flavour and Fragrance Journal - FLAVOUR FRAG J. 01/2000; 15(2):119-122.
- Flavour and Fragrance Journal - FLAVOUR FRAG J. 01/2000; 15(6):373-376.
Advance Access Publication 12 December 2006eCAM 2007;4(3)351–353
In Vitro Anti-Candida Activity of Zataria multiflora Boiss
Ali Zarei Mahmoudabadi1, Muhammad Ali Dabbagh2and Zahra Fouladi3
1Department of Medical Mycoparasitology, School of Medicine, Jundishapour University of Medical Sciences,
2School of Pharmacy, Jundishapour University of Medical Sciences and3School of Nursing and Midwifery,
Jundishapour University of Medical Sciences, Ahwaz, Iran
Zataria multiflora Boiss known as Avishan Shirazi (in Iran) is one of the valuable Iranian medicinal
plants. The aim of study was to evaluate anti-Candida activity of Z. multiflora against different species
of Candida in vitro. Anti-Candida activity of the aqueous, ethanolic and methanolic maceration extract
of the aerial parts of Z. multiflora Boiss was studied in vitro. Anti-Candida activity against Candida
species was done using serial dilutions of extracts in Sabouraud’s dextrose agar. Minimal inhibitory
concentration (MIC) of the methanolic and ethanolic extracts was 70.7 and 127 mg l?1, respectively.
Aqueous extract showed no remarkable activity against Candida species. We conclude that methanolic
extract of the aerial parts of Z. multiflora Boiss has more anti-Candida effect at 70.7 mg l?1compared to
ethanolic extract 127 mg l?1. In addition, the isolates of Candida parapsilosis were more susceptible to
methanolic extract than other tested species.
Keywords: anti-Candida – Candida – herbal medicine – Zataria multiflora Boiss
Zataria multiflora Boiss (Lamiaceae) is a valuable medicinal
plant grown extensively in Iran, Pakistan and Afghanistan (1).
The chemical compositions of extracts have been extensively
characterized in Iran (2–5) and Pakistan (6). The extract
contains thymol, carvacrol (4,6), zatrinal, oleanolic acid,
betulic acid, rosmarinic acid (5) and monoterpenoids,
sesquiterpenoids, p-cymene, y-terpinene (3,4).
Aqueous and alcoholic extracts of Z. multiflora have been
therapeutically used for relieving nociceptive pain (7,8),
recurrent aphthous stomatitis (RAS) (9), and prevent growth
of oral streptococci (10), Plasmodium falciparum (11) and
Trichomonas vaginalis (12) as well as used as an insect
repellent (6). Fataneh (13) has investigated anti-fungal
properties of Z. multiflora extract in vitro. In view of its potent
antibacterial and anti-fungal activities, we hypothesized that
Z. multiflora extracts may possess anti-Candida effects. We
have tested the hypothesis in vitro by comparing the aqueous,
ethanolic and methanolic extracts of Z. multiflora against
14 isolates of Candida albicans, C. parapsilosis, C. tropicalis
and C. glabrata.
Materials and Methods
The plant was collected from Shiraz, Iran, and identified by
Agricultural Research Centre, Ahwaz. The plant was identified
in the Systematic Laboratory, Agricultural Sciences Centre,
Ahwaz, Iran, where voucher specimens were deposited
(ZM 1). Aliquots of 100 g of the dried powder of the plant
were soaked in ethanol (1400 ml), methanol (1400 ml) and
distilled water (2150 ml) for 24 h and then filtered with
cloths. The extracts were concentrated to dryness in a vacuo at
53–55?Cand yielded 11.39 g (aqueous extract), 15 g(ethanolic
extract) and 13.3 g (methanolic extract).
C. albicans (n ¼ 7), C. tropicalis (n ¼ 3), C. parapsilosis
(n ¼ 2) and C. glabrata (n ¼ 2). All Candida species
isolatesof Candida werestudiedincluding
For reprints and all correspondence: Ali Zarei Mahmoudabadi, Department
of Medical Mycoparasitology, School of Medicine, Jundishapour University
of Medical Sciences, Ahwaz, Iran. Tel: þ98611-3330074; Fax: þ98611-
3332036; E-mail: email@example.com
? 2006 The Author(s).
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/
by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
were isolated from infected patients in the department of
medical mycoparasitology, Jundishapour University of medi-
cal sciences, Ahwaz, Iran. All isolates were identified by
France), germ-tube test, production of chlamydoconidia on
Corn meal agar and growth at 45?C. Isolates were maintained
on Sabouraud’s dextrose agar (SDA) at 4?C. Organisms were
subcultured on SDA and incubated at 37?C for 24 h. Several
colonies of each Candida species were collected in 2 ml of
sterile PBS to prepare a suspension. The suspension was
adjusted to 70% transmittance (T) by a spectrophotometer at
530 nm. This should result in a suspension containing about
1 · 106cfu per ml.
A serial dilution of each extract was prepared in SDA plates.
Aqueous, ethanolic and methanolic extracts were diluted by
the same solvent. The same solvent, at an appropriate
concentration was also used as a negative control. A plate
was considered as positive control without extracts and
solvents. Aliquots of 20 ml of standardised suspension of
different species of Candida were inoculated in to each plate.
The plates were incubated at 30?C for 24–48 h. The lowest
extract concentration where there was no visible growth was
the minimal inhibitory concentration (MIC) when compared to
control. All experiments were repeated three times and mean
In the present study the anti-Candida activity of three extracts
of Z. multiflora (aqueous, ethanolic and methanolic) was
evaluated against 14 isolates of Candida. In the first stage,
aqueous, ethanolic and methanolic extracts of Z. multiflora
applied on one isolate of each Candida species. Aqueous
extract of Z. multiflora showed no activity against Candida
species. As a result this extract was removed from the next
experiments. The ethanolic and methanolic extracts showed
remarkable activities against Candida species. The MIC for
both extract was between 50 and 150 mg l?1. In the second
stage, ethanolic and methanolic extracts were used for the
detection of MIC.
Table 1 shows details of mean MICs of ethanolic extract
against 14 isolates of Candida. As shown the lowest MIC was
for 7 isolates of C. albicans (125 mg l?1). Others MICs
were respectively C. glabrata (126 mg l?1), C. parapsilosis
(125 mg l?1) and C. tropicalis (131 mg l?1). Totally, the MIC
of ethanolic extract for 14 isolates of Candida was 127 mg l?1.
As shown both C. albicans and C. parapsilosis are more
susceptible than other species.
Table 1 shows the details of mean MICs of methanolic extract
against tested Candida. As shown isolates of C. parapsilosis
(64 mg l?1) are more susceptible to methanolic extract of
Z. multiflora, followed by C. glabrata (66 mg l?1), C. albicans
(76 mg l?1) and C. tropicalis (76 mg l?1). Totally, the MIC of
methanolic extract for tested Candida was 70 mg l?1. In the
present study methanolic extract showed more activities than
ethanolic extract against 14 isolates of Candida.
Herbal and alternative medicines are popular in the general
population worldwide. A great number of modern drugs are
still derived from herbs (14). Iranian scientist, Avicenna
(980–1037) and Razi (846–930) published several books
on herbal medicine a few centuries ago and are still in use in
different libraries in Europe (15). Z. multiflora grows wild in
Table 1. Minimal inhibitory concentration (MIC) of ethanolic (E) and methanolic (M) extracts of Zataria multiflora Boiss (Lamiaceae) required for inhibition
of Candida species from 14 isolates from infected patients
Minimal inhibitory concentration (mg l?1)
C. albicans (7) C. tropicalis (3) C. glabrata (2) C. parapsilosis (2)
E extractM extract E extractM extract E extract M extract E extractM extract
14693 139 9312363 12563
13073 126 66 130 70125 66
110 76 12970
876530 394 229253133 250129Total
125.175.7131.376.3126.566.5 12564.5 Mean
10.89.5 5.611.93.5 3.50.01.5SD
Values in parenthesis refer to the number of isolates.
C. albicans (7): CA1, CA2, CA3, CA4, CA6, CA7, CA11; C. tropicalis (3): CT2, CT3, CT4; C. glabrata (2): CG 1, CG3; C. parapsilosis (2): CP1, CP2.
Anti-Candida activity of Zataria multiflora
central and southern Iran. Z. multiflora is used in traditional
herbal medicine for antiseptic, analgesic, and carminative
properties (2,7,16). Z. multiflora was also used for treatment of
‘Women disease’ in Iranian folklore (17). The leaf powder of
Z. multiflora is used as nutritional flavoring in Iran. It is
important to investigate scientifically those plants, which have
been used in traditional medicines as potential sources of novel
The presence of thymol, rosmarinic acid, and carvacrol in
the different parts of the plant was observed (6). The present
results indicate that methanolic extracts of the aerial part of
Z. multiflora have marked activity against isolates of Candida.
Probably, the anti-Candida activity of methanolic extract of
Z. multiflora is due to both rosmarinic acid and thymol that
extracted only into methanol (2). Probably, the anti-Candida
activity of methanolic extract of Z. multiflora is due to above
compounds. Z. multiflora is used in traditional herbal medicine
for women disease (Candidiasis vagina). Fataneh (13) have
shown that Z. multiflora has anti-fungal activity. They tested
several isolates of dermatophytes and saprophytic fungi
against Z. multiflora extract. Amanlou et al. (18) have shown
that Z. multiflora has antierythema in denture stomatitis
compared to miconazole gel, however, Z. multiflora gel did
not reduce the colony count of the denture surface as
efficiently as miconazole gel. The ethanolic extracts of aerial
parts of Z. multiflora showed antinociceptive activity (19,20).
Phytochemical screening supported the presence of flavonoids
in Z. multiflora. Some flavonoids exert antinociceptive activity
in mice (19). Ramazani et al. (8) reported six fractions of the
extracts of aerial parts of Z. multiflora that have antinocicep-
of potentially useful anti-Candida and is worthy for future
clinical study. In addition, measures must to be undertaken to
preserve the traditional knowledge about medicinal plants.
This study was supported by a grant from Jundishapour
University of Medical Sciences, Ahwaz, Iran.
1. Ali MS, Saleem M, Ali Z, Ahmad VU. Chemistry of Zataria multiflora
(Lamiaceae). Phytochemistry 2000;55:933–6.
2. Mohagheghzadeh A, Shams-Ardakani M, Ghannadi A, Minaeian M.
Rosmarinic acid from Zataria multiflora tops and in vitro cultures.
3. Mohagheghzadeh A, Shams-Ardakani M, Ghannadi A. Linalol-rich
essential oil of Zataria multiflora Boiss (Lamiaceae). Flavour Fragrance
4. Mohagheghzadeh A, Shams-Ardakani M, Ghannadi A. Volatile consti-
tutes of callus and flower-bearing tops of Zataria multiflora Boiss
(Lamiaceae). Flavour Fragrance J 2000;15:373–6.
5. Javidnia K, Tabatabai M, Shafiee A. Volatile constituents and antimicro-
bial activity of Zataria multiflora, population Iran. Irn J Chem Chem Eng
6. Saleem M, Nazli R, Afza N, Sami A, Ali MS. Biological significance of
essential oil of Zataria multiflora Boiss. Nat Prod Res 2004;18:493–7.
7. Hosseinzadeh H, Ramezani M, Salmani GA. Antinociceptive, anti-
inflammatory and acute toxicity effects of Zataria multiflora Boiss
extracts in mice and rats. J Ethnopharmacol 2000;73:379–85.
8. Ramezani M, Hosseinzadeh H, Samizadeh S. Antinociceptive effects of
Zataria multiflora Boiss fractions in mice. J Ethnopharmacol 2004;91:
9. Jafari S, Amanlou M, Borhan-Mohabi K, Farsam H. Comparative study of
Zataria multiflora and Anthemis nobelis extracts with Myrthus communis
preparation in the treatment of recurrent aphthous stomatitis. Daru
10. Owlia P, Pirveicy H, Saderi H, Rezvani MB, Mansouri S. Evaluation of
the antimicrobial effects of extract of Zataria multiflora against oral
Streptococci. Iranian J Pharm Res 2004;2:74–5.
11. Ziegler HL, Franzyk H, Sairafianpour M, Tabatabai M, Tehrani MD,
Bagherzadeh K, et al. Erythrocyte membrane modifying agents and
the inhibition of Plasmodium falciparum growth: structure-activity
relationships for betulinic acid analogues. Bioorg Med Chem 2004;12:
12. Abdollahy F, Ziaei H, Shabankhani B, Azadbakht M. Effect of essential
oils of Artemisia aucheri Boiss. Zataria multiflora Boiss, and Myrtus
communis L. on Trichomonas vaginalis. Iranian J Pharm Res 2004;2:
(Suppl 2): 35.
13. Fataneh F. Anti-fungal activity of Zataria multiflora extract in vitro.
Thesis, School of Pharmacy, Esfahan University of Medical Sciences,
1991 (in Persian).
14. Cooper EL. CAM, eCAM, bioprospecting: the 21st century pyramid. Evid
Based Complement Alternat Med 2005;2:125–7.
15. Saad B, Azaizeh H, Said O. Tradition and perspectives of Arab herbal
medicine: a review. Evid Based Complement Alternat Med 2005;2:475–9.
16. Zargari A. Medicinal Plants, Vol. 4., Tehran: Tehran University Press,
1990 (in Persian).
17. Rojhan MS. Health and treatment with medicinal plant and pharmacog-
nosy. Tehran: Tanin Co., 2000 (in Persian).
18. Amanlou M, Beitollahi JM, Abdollahzadeh S, Tohidast-Ekrad Z.
Miconazole gel compared with Zataria multiflora Boiss gel in the
treatment of denture stomatitis. Phytother Res 2006;20:966–9.
19. Ramesh M, Rao YN, Rao AV, Prabhakar MC, Rao CS, Muralidhar N,
et al. Antinociceptive and anti-inflammatory activity of a flavonoid
isolated from Caralluma attenuata. J Ethnopharmacol 1998;62:63–6.
20. Jaffary F, Ghannadi A, Siahpoush A. Antinociceptive effects of
hydroalcoholic extract and essential oil of Zataria multiflora. Fitoterapia
Received May 15, 2006; accepted October 30, 2006