Iranian Journal of Pharmaceutical Research (2013), 12 (4): 801-810 Copyright © 2013 by School of Pharmacy

Received: August 2012 Shaheed Beheshti University of Medical Sciences and Health Services
Accepted: August 2013

Original Article

Cytotoxic, Antioxidant and Antimicrobial Activities and Phenolic Contents

of Eleven Salvia Species from Iran

Omidreza Firuzi, Ramin Miri, Mojtaba Asadollahi , Saba Eslami and Amir Reza Jassbi*

Medicinal and Natural Products Chemistry Research Center, Shiraz University of Medical
Sciences, Shiraz, Iran.

Abstract

The plants of the genus Salvia synthesize several types of secondary metabolites with
antimicrobial, cytotoxic, and radical scavenging activities and are used in the folk medicine of
different countries. Eleven Salvia species including S. aegyptiaca, S. aethiopis, S. atropatana,
S. eremophila, S. hypoleuca, S. limbata, S. nemorosa, S. santolinifolia, S. sclarea, S. syriaca,
and S. xanthocheila were collected from different localities in Iran and screened for their
cytotoxic activity using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)
colorimetric assay. The antioxidant potential and total phenol contents of the plant extracts
were assessed by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay and Folin-
Ciocalteu reagent respectively and finally antimicrobial activity of the above extracts were
determined by using agar disc diffusion (ADD) and nutrient broth micro-dilution (NBMD)
bioassays. Cytotoxic activity of methanol, 80% methanol and dichloromethane extracts of
these plants were assessed on 3 human cancer cell lines. All of the extracts of S. eremophila
and S. santolinifolia were active at IC50 values of 10.5-75.2 µg extract/mL, while the methanol
and dichloromethane extracts of S. limbata, S. hypoleuca and S. aethiopis showed considerable
cytotoxic activity against the tested cell lines. Among the tested plants for their antioxidant
activity, S. nemorosa, S. atropatana, S. santolinifolia, and S. eremophila were the most active
radical scavengers with higher total phenol contents while, S. limbata, S. xanthocheila and S.
aegyptiaca were the weakest ones. The methanol extracts of S. santolinifolia, S. eremophila, S.
sclarea and S. limbata inhibited the growth of all tested bacterial strains.

Keywords: Salvia; Phenolics; DPPH; Radical-scavenging; Cytotoxic activity; Antibacterial

activity.

Introduction oil and non-volatile terpenoids especially di- and

triterpenoids (4, 5). These plants also synthesize
Sage plants (of genus Salvia) are known for polyphenols, including flavonoids and caffeic
their uses in the folk medicine and as additives acid derivatives (6, 7).
in food products in different countries including The diterpenoids such as abietane pigments
Iran and Turkey (1, 2). They are widely spread (5), pimarane (1) and labdane type diterpenoids,
in both countries and are rich in volatiles such as and triterpenoids together with volatile mono-
mono-and sesquiterpenoids (3) in their essential and sesquiterpenoids found in their essential oils
are responsible for different biological activities
* Corresponding author: including antimicrobial (1, 8), cytotoxic (9, 10),
E-mail: [email protected] enzyme inhibitory (11-13) and anti-leishmania
Firuzi O et al. / IJPR (2013), 12 (4): 801-810

properties (14). Some of the species of Salvia the shade and were used for solvent extraction.
including S. hypoleuca (15) and S. syriaca (16)
are unique sources of the rare sesterterpenoids in Solvent extraction of the plants
the terrestrial plants. The interesting biological The aerial part of each plant was separately
activities resulting from their chemical diversities extracted with dichloromethane, methanol and
prompted different researchers worldwide to 80% methanol for the cytotoxic and antibacterial
examine these plants for their constituents and bioassays. Extracts were prepared as follows;
biological activities. 3 g of the dry plant was macerated in 60 mL
As a starting point for selection of target plants of the solvents for 24 h. The extraction was
for the isolation of bioactive natural products, repeated twice and the resulting extracts were
we subjected eleven sage species collected added to each other. The extract was then filtered
from different localities in Iran to antioxidant, and concentrated in a rotary evaporator under
antimicrobial and cytotoxic bioassays against reduced pressure for the removal of solvents.
human cancer cell lines. They are S. aegyptiaca, The resulting concentrated extracts were kept
S. aethiopis, S. atropatana, S. eremophila, at -20 °C until their use for antimicrobial and
S. hypoleuca, S. limbata, S. nemorosa, S. cytotoxic tests. Shortly before each experiment,
santolinifolia, S. sclarea, S. syriaca, and S. the syrup was dissolved in the appropriate
xanthocheila. solvent (DMSO) and used in the bioassay. The
extracts that were used for antioxidant and
Experimental total phenols measurements were prepared in a
different way. Twenty-five mg of dried powdered
Reagents plant material was extracted in 1.5 mL 80%
Quercetin was obtained from Acros Organics methanol for 48 h and an aliquot of the extract
(Geel, Belgium). Fetal bovine serum (FBS), RPMI without further concentration was subjected to
1640, trypsin and phosphate buffered saline (PBS) the above-mentioned assays.
were purchased from Biosera (Ringmer, UK).
Dimethylsulfoxide (DMSO), Folin-Ciocalteu Cell lines and culture
reagent, nutrient broth, hexane, methanol and The following human cancer cell lines were
sodium carbonate were purchased from Merck purchased from the National Cell Bank of Iran,
(Darmstadt, Germany). 3-(4,5-dimethylthiazol- Pasteur Institute, Tehran, Iran; HL60 (human
2-yl)-2,5-diphenyltetrazolium bromide (MTT), acute promyelocytic leukemia cells), K562
2,2-diphenyl-1-picrylhydrazyl (DPPH), (human chronic myelogenous leukemia cells)
chloramphenicol, and hydrochloric acid and MCF-7 (human breast adenocarcinoma
32% were obtained from Sigma-Aldrich cells).
(St. Louis, MO, USA). Doxorubicin and The cells were cultured in sterile T25 flasks
penicillin/streptomycin were purchased from in RPMI 1640 medium supplemented with fetal
EBEWE Pharma (Unterach, Netherlands) and bovine serum (20% v/v for HL60 and 10% v/v
Invitrogen (San Diego, CA, USA), respectively. for K562 and MCF-7 cells), penicillin (100
p-iodonitrotetrazolium violet (INT) was obtained units/mL) and streptomycin (100 µg/mL). HL60
from Fluka. and K562 cell lines were grown in suspension,
while MCF-7 cells were grown in mono layer
Plant material cultures in humidified air constituting 5% CO2
Plants studied in this report were collected at 37 °C.
in June and July 2008 from different areas of
Iran (Table 1) and identified at the Medicinal Cytotoxicity assay
and Natural Products Chemistry Research The inhibitory effect of plant extracts on cell
Center (MNCRC), Shiraz, Iran by Dr. Mojtaba growth was assessed by 3-(4,5-dimethylthiazol-
Asadollahi. The voucher specimens were 2-yl)-2,5-diphenyltetrazolium bromide (MTT)
deposited at MNCRC herbarium. Aerial parts of assay. This colorimetric assay is based on the
the plants were air-dried at room temperature in conversion of the yellow tetrazolium bromide

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(MTT) to the purple formazan by the action of Microsoft Excel and Curve Expert statistical
mitochondrial enzyme succinate dehydrogenase programs and expressed as μg plant material
in viable cells. The powdered extracts were extracted with the solvent/ 1 mL 10-4 M DPPH
dissolved in DMSO, and then diluted in growth (µg PM /ml DPPH).
medium at least 200 times. Cells were seeded in
96-well plates at the density of 50,000 cells/mL Determination of the total phenol content in
in 100 µL medium and incubated for 24 h. Then, the plant extracts
50 µL of medium was replaced with fresh medium The total phenol contents of the plant extracts
containing 3 different concentrations of the were determined by the Folin-Ciocalteu method
extracts. After 72 h of incubation, the medium of as described previously with some modifications
each well was replaced by RPMI without phenol (19). Briefly, 3.16 mL water was added to a 40 µL
red containing 0.5 mg/mL MTT and incubated at solution of the plant extract (80% methanol) and
37 °C for 4 h. DMSO was used to dissolve the 200 µL Folin-Ciocalteu reagent, and the mixture
formed formazan crystals. The potency of cell was shaken until it became homogenous. To this
growth inhibition for each extract was expressed solution was added 600 µL of a 0.25% sodium
as IC50 value, defined as the concentration that carbonate after 8.5 min incubation at room
caused a 50% of maximum inhibition of cell temperature. The above solution was further
viability. The absorbance of different wells was incubated at RT for 2 h and its absorbance
measured at 570 nm, with background correction was measured at 765 nm against the blank.
at 655 nm using a microplate reader. Inhibition The concentrations of the total phenolics were
percentages were plotted against different measured against a series of gallic acid standard
concentrations of the extracts and cisplatin. The solutions and expressed as mg equivalent of
IC50s were calculated by best fit equations using gallic acid in 1g dry plant material (mg EG/g
Curve Expert statistical program. PM) (19).

Determination of the free radical Antibacterial agar disc diffusion method

scavenging activity of the plant extracts by To examine the antibacterial activity of the
spectrophotometric methods plant extracts, three Gram-negative bacteria
The free radical scavenging activity of the (Escherichia coli: PTCC1330, Klebsiella
plant extracts was measured by the method of pneumoniae: PTCC1053 and Salmonella
Blois (17) with some modifications (18) and typhi: PTCC1609) and three Gram-positive
compared to that for quercetin as a standard bacteria (Staphylococcus aureus: PTCC1112,
radical scavenger. Briefly, 25 mg of dried Staphylococcus epidermidis: PTCC1114,
powdered plant was extracted in 1.5 mL 80% Bacillus subtilis: PTCC1023) were chosen and
methanol for 48 h. 25-100 μL of this extract tested in agar disc diffusion (ADD) bioassays.
were adjusted to 200 μL by methanol to obtain The minimum inhibitory concentrations (MIC)
different concentrations of the plant᾽s extract of the active extracts were determined using
and then added to 3800 μL 10-4 M DPPH nutrient broth micro-dilution (NBMD).
solutions in methanol. After 30 min shaking of Bacteria were grown in nutrient broth
the solutions in the dark, the absorptions of the (Merck) overnight at 37 ºC and before seeding
DPPH solutions were measured at 517 nm. The the agar plates, their optical density were
percentage of the reduced DPPH was calculated measured at 600 nm and adjusted to 0.1. An
by the following equation: aliquot, containing 5 mg of the crude extract
Percentage of DPPH reduction = ((A0 – A1) (dichloromethane, methanol and 80% methanol)
/ A0) x 100), where A0 is the absorbance of the were applied onto paper disc of 6 mm diameter.
control, and A1 is the absorbance in the presence The dried papers were placed on agar seeded
of sample. The IC50s were calculated by linear with 1 mL of the above bacteria suspension in
regression equations of the DPPH inhibition a Petri dish. The Petri dishes were placed for
percentage from different concentrations of the 5 h at 4 °C that the metabolites could diffuse
extracts and the standard antioxidants, using in the medium. The plates were incubated at

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37 °C for 18 h. The antibacterial activity was with IC50 values of 10.5-75.2 µg extract/mL. The
determined by measuring the diameters of the second most active plant was S. santolinifolia,
clean inhibitory zone (IZ) around each paper as all of its extracts (with the exception of
disc. Chloramphenicol was used as the positive 80% methanol extract on MCF-7 cells) were
control (20). The most active crude extracts effective on the tested cell lines with IC50 values
were found to be those that were extracted with of 47.0-147.1 µg extract/mL. This indicates that
methanol, therefore the methanol extracts of the there must be some compounds with different
plants (data are not shown) were chosen for MIC polarities in all of the extracts of these two plants
determination. that were responsible for the cytotoxic activity.
The methanol and dichloromethane extracts of
Antibacterial minimum inhibitory S. aegyptiaca, S. aethiopis, S. hypoleuca and
concentration (MIC) using nutrient broth micro- S. limbata also showed significant cytotoxic
dilution (NBMD) activity against the tested cell lines (Table 2).
NBMD was performed using serial two-fold However, cisplatin the anticancer positive
dilution of the plant extracts added to bacterial control was several folds more active with IC50
suspension in nutrient broth as previously values of 0.8 ± 0.1, 2.1±0.2 and 6.9±1.8 µg/mL
described (21). The plant extracts or positive for HL60, K562 and MCF-7 respectively, than
control was dissolved in DMSO in different the entire of the extracts significantly (Table 2;
concentrations and was added (5 µL) to 95 µL of p ≤ 0.001) except for dichloromethane extract
fresh media and 100 µL of bacterial suspension of S. eremophila p = 0.57. This shows that
(OD=0.1 at 600 nm) in a 96-well microplate. the activity of the extract and the drug are not
The microplates were incubated at 37 ºC for 24 different from each other.
h in a shaking incubator and then 10 µL of 0.5% Recently, some of the above studied plant
INT solution in water was added to each well species were subjected to cytotoxic bioassays
and incubated for further 30 min at the above by different authors. For instance the methanol
condition. The MIC was considered as the lowest extracts of S. eremophila and S. santolinifolia
concentration of the extract or antibacterial have been examined on different human cancer
standard which discolored the purple color of cell lines, however, the cytotoxic doses have been
the INT solution. reported above 200 µg extract/mL (9). In contrast,
the triterpenoids and abietane diterpenoids
Statistical analysis isolated from the aerial parts of S. eremophila
Values shown in tables 2, 3 and 4 are the average have shown similar cytotoxic activities on 5
of 3-5 measurements ± SE. Correlation coefficient human cancer cell lines, which is comparable
(R) between the two variables in table 3 were with our results (10). In another experiment, six
calculated by MS Excel software. The IC50s were flavonoids with reported cytotoxic activity and
calculated using Curve Expert statistical program. rosmarinic acid were isolated from S. limbata
One-way analyses of variance (ANOVA) post (22), which may be the cause of the relatively
hoc multiple comparison (Tukey) tests was used high cytotoxicity of the extracts of this plant on
for the determination of signification between cancer cells examined in this paper.
different measurements using SPSS software and
expressed as probability factor, p-value. P ≤ 0.05 Antioxidant activity and total phenolic
was considered to be significant. content
Since 80% methanol is recommended as
Results and Discussion extracting solvent for phenolic compounds (23),
we extracted the plants with the above solvent
Cytotoxic activity system and measured their antioxidant activity
Cytotoxicity of the plant extracts was using DPPH radical scavenging and the total
measured on 3 human cancer cell lines (Table phenolic content by Folin-Ciocalteu assay
2). The most active plant was S. eremophila, (Table 3). Among the tested plants, S. nemorosa
as all of its extracts were active on 3 cell lines (IC50 138.43 ± 4.6 µg PM /mL DPPH p ≤ 0.001

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Table 1. Location and herbarium specimens of the plants species.

Plant Name Location Herbarium Number Date
Darab towards Rostagh, Fars
N 28º, 35’ a June 2008
Salvia aegyptiaca L. PC-87-90
E 54º, 47’
1260 m
Arasbaran Forest- East Azarbaijan
N 38º, 53’ August 2008
Salvia aethiopis L. PC-87-91
E 46º, 50’
1800 m
Cheleghah, Sepidan, Fars
N 30º, 17’ July 2008
Salvia atropatana Bunge. PC-88-19
E 51º, 56’
2370 m
Darab, Fars
N 28º, 41’ June 2008
Salvia eremophila Boiss. PC-87-92
E 54º, 37’
1170 m
Kandovan-Chalus road- Mazandaran
N 36º, 10’ July 2008
Salvia hypoleuca Benth. PC-88-18
E 51º, 18’
2500 m
Shahin dej, west Azarbayjan
N 36º, 39’ August 2008
Salvia limbata C. A. Mey. PC-87-93
E 46º, 32’
1500 m
Marzanabad, Chalus, Mazandaran
N 36º, 27’ July 2008
Salvia nemorosa L. PC-88-20
E 51º, 18’
480 m
Darab, Fars
N 28º, 41’ June 2008
Salvia santolinifolia Boiss. PC-87-98
E 54º, 37’
1170 m
Sepidan (Ardakan) towards Komehr, Fars
N 30º, 24’ July 2008
Salvia sclarea L. PC-87-99
E 51º, 54’
2600 m
Shiraz-Sepidan road, after shool village, Fars
N 29º, 58’ June 2008
Salvia syriaca L. PC-87-100
E 52º, 10’
2090 m
Kandovan-Chalus road- Mazandaran
Salvia xanthocheila Boiss. ex N 36º, 10’
PC-87-101 July 2008
Benth. E 51º, 18’
2500 m
a) The approximate collection coordinates of the plants

with all of the extracts except for S. atropatana, 25.70 ± 0.04 mg EG/g PM; p ≤ 0.0001 with all of
S. aethiopis, S. eremophila, S. hypoleuca, S. the extracts), S. santolinifolia (IC50 117.34 ± 4.07
santolinifolia, and S. sclarea; 30.36 ± 1.08 mg µg PM /mL DPPH, p ≤ 0.04 with all of the extracts
EG/g PM; p ≤ 0.0001 with all of the extracts), S. except for S. atropatana, S. eremophila and S.
atropatana (IC50 89.47 ± 5.97 µg PM /mL DPPH nemorosa; 20.21 ± 0.87 mg EG/g PM p ≤ 0.0001
p ≤ 0.001 with all of the extracts except for S. with all of the extracts except for S. eremophila,
eremophila, S. nemorosa and S. santolinifolia; S. hypoleuca and S. syriaca), and S. eremophila

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Table 2. Cytotoxic activity of different extracts of Salvia species on human cancer cell lines.
IC50 (µg/mL) b
Plant a Extraction solvent
HL60 Cells K562 cells MCF-7 Cells
Dichloromethane 99.7 ± 3.9 97.0 ± 6.2 116.1 ± 5.0
Salvia aegyptiaca Methanol NA c
NA NA
Methanol 80% NA NA NA

Dichloromethane 44.6 ± 8.2 41.3 ± 4.5 44.4 ± 5.0

Salvia aethiopis Methanol NA 50.1 ± 4.2 79.4 ± 12.3
Methanol 80% NA NA NA

Dichloromethane 10.5 ± 0.6 15.8 ± 2.9 e 45.6 ± 1.8

Salvia eremophila Methanol 11.9 ± 1.8 15.6 ± 2.9 47.7 ± 1.9
Methanol 80% 24.7 ± 1.9 42.7 ± 2.5 75.2 ± 6.6

Dichloromethane 53.3 ± 2.5 48.6 ± 2.7 83.0 ± 14.2

Salvia hypoleuca Methanol 95.7 ± 6.9 93.7 ± 7.7 105.7 ± 2.1
Methanol 80% NA NA 99.4 ± 10.0

Dichloromethane 51.0 ± 1.5 45.9 ± 0.7 64.3 ± 14.1

Salvia limbata Methanol NA 111.3 ± 11.4 148.9 ± 11.8
Methanol 80% NA 110.6 ± 16.3 NA

Dichloromethane NA NA NA
Salvia nemorosa Methanol NA NA NA
Methanol 80% NA 87.0 ± 6.7 NA

Dichloromethane 48.2 ± 1.4 78.7 ± 15.0 147.1 ± 68.1

Salvia santolinifolia Methanol 47.0 ± 3.5 49.4 ± 1.6 108.8 ± 9.2
Methanol 80% 49.2 ± 1.7 54.6 ± 2.2 NA

Dichloromethane NA NA NA
Salvia syriaca Methanol NA NA NA
Methanol 80% NA NA 75.4 ± 30.3

Dichloromethane NA NA NA
Salvia xanthocheila Methanol 166.9 ± 49.4 72.8 ± 9.2 127.8 ± 12.5
Methanol 80% NA NA NA

Cisplatin d 0.8 ± 0.1 2.1 ± 0.2 6.9 ± 1.8

a)
Salvia atropatana and Salvia sclarea were also tested, but none of their extracts were active on any of the cell lines. b) Values are
presented as mean ± SE of 4-5 experiments. c) NA: Not active; IC50 more than 200 µg/mL. d) Cisplatin was tested as a reference cytotoxic
compound and found to be more active than all extracts with p ≤ 0.001 except for the dichloromethane extract of S. eremophila.

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Table 3. Total phenolic content and DPPH radical scavenging The results of this study are consistent with
potential of the 80% methanol extracts of plants.
our previous findings on screening of some of
Plant name IC50 DPPH a Total phenol b
the Lamiaceae plants (25). Among the extracts of
S. xanthocheila 457.00± 41.62 12.49± 1.18
twenty-four plants of the family Lamiaceae, the
S. limbata 557.40± 12.73 12.95± 0.70 extracts of Salvia in general were the most active
S. aegyptiaca 330.4± 11.06 13.83± 0.16 radical scavengers among which S. eremophila and
S. aethiopis 237.37± 8.05 14.13± 0.90 S. santolinifolia were the most active ones (25).
S. sclarea 190.74± 5.7 14.83± 0.80 In another study, the essential oil and methanolic
S. syriaca 315.1± 5.71 18.08 ± 0.41 extract of S. eremophila were subjected to DPPH
S. eremophila 114.57± 11.5 18.86± 0.98 bioassay (26). Only the methanolic extract of
S. santolinifolia 117.34± 4.07 20.21± 0.87
the plant significantly reduced the reagent (26).
S. hypoleuca 197.23± 6.86 20.27± 0.50
Examination of antioxidant and total phenol
contents of six Salvia species has resulted in
S. atropatana 89.47± 5.97 25.70± 0.04
determination of S. xanthocheila and S. sclarea
S. nemorosa 138.43± 4.6 30.36±1.08
as the weakest radical scavengers in those plant
Quercetin 1.79±0.046 -
series (27). S. hypoleuca and four other sage plants
a) DPPH IC50 (μg plant extracted or μg quercetin/ 1 mL 10-4 M were assessed for their DPPH antioxidant activity
DPPH), b) Total phenol (mg eq. gallic acid in 1 g dried plant). and total flavonoid contents, but no favorable
correlation was detected between the tests results
(IC50 114.57 ± 11.5 µg PM /mL DPPH, p ≤ 0.03 in different plants (28). Fourteen Turkish sage
with all of the extracts except for S. atropatana, species were examined for their antioxidant
S. nemorosa and S. santolinifolia; 18.86 ± 0.98 activity using DPPH reagent, among which
mg EG/g PM, p ≤ 0.002 with all of the extracts different extracts of S. sclarea and S. syriaca
except for S. santolinifolia, S. hypoleuca and S. showed similar inhibition percentage; however,
syriaca) were the most active radical scavengers these two plant extracts had lowest antioxidant
with the highest total phenol contents. However, activity (29). On the other hand, these plants are
S. limbata, S. xanthocheila, S. aegyptiaca and S. considered as medium radical scavengers in our
aethiopis in increasing order of efficiency, had sage series (Table 3).
the lowest radical scavenging potential (DPPH
IC50S 557.40 ± 12.73 to 237.37 ± 8.05 µg PM Antimicrobial activity of the sage extracts
/mL DPPH) with lower total phenol contents Antimicrobial activity of the methanol
compared to the above mentioned plants (Table extracts of the plants were measured on 6
3). There was not a perfect correlation (R2 = different Gram-negative and Gram-positive
0.48; R = -0.69) between antioxidant and total bacteria (Table 4). S. eremophila, S. limbata, S.
phenol content data in our test (Table 3). This santolinifolia and S. sclarea were the most active
may be due to the presence of different types plants and inhibited the growth of all tested
of phenolic compounds in the plant extracts. microorganisms at MIC values between 0.31-5
However the IC50 of quercetin, the standard mg/mL on the tested microorganisms (Table 4).
natural antioxidant is 1.79 ± 0.046 µg/mL DPPH On the other hand, S. aegyptiaca and S. aethiopis
which is significantly different from all of the were only active at MIC 5 mg/mL against the
tested plants extracts reported here (p ≤ 0.0001). growth of S. typhi and therefore resulted the
Several structure-activity-relationship studies weakest plants in the antimicrobial bioassay.
reported the effect of the number and position The gram positive bacteria with MICs 0.0125
of phenolic hydroxyls in the radical scavenging for B. subtilis and Staph. aureus and 0.025 for
potential of the phenolic compounds (24); for Staph. epidermidis were more susceptible to
instance when one of the ortho- or para- free chloramphenicol than the gram negative ones
hydroxyls in a phenolic compound are protected with MICs 0.05 mg/mL media.
by glycosylation or methylation, then its radical Most of the studies performed on this genus
scavenging activity is dramatically decreased in Iran evaluated the antimicrobial activity of
(18). the essential oils (3). Both essential oil and

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Table 4. Antimicrobial potential (MICa) of different plant extracts by nutrient-broth micro-dilution bioassay.
Plant name E. coli a K. pneumoniae S. typhi B. subtilis Staph. epidermidis Staph. aureus
S. aegyptiaca NA b
NA 5 NA NA NA
S. aethiopis NA 10 5 NA NA NA
S. atropatana 2.5 NA 5 2.5 NA NA
S. eremophila 2.5 2.5 0.31 2.5 0.3 0.6
S. hypoleuca NA 2.5 5 NA 1.25 1.25
S. limbata 5 2.5 1.25 5 2.5 2.5
S. nemorosa NA NA 10 NA NA NA
S. santolinifolia 2.5 5 1.25 2.5 1.25 2.5
S. sclarea 5 2.5 0.31 5 0.31 1.25
S. syriaca NA NA 2.5 NA 2.5 1.25
S. xanthocheila 5 2.5 2.5 2.5 NA 1.25
Chloramphenicol 0.05 0.05 0.05 0.0125 0.025 0.0125

a) Minimum inhibitory concentration (MIC) of the plant extracts in the bacterial suspension in the nutrient broth media (mg/ mL). b)
NA= not active.

methanol extract of S. eremophila showed S. santolinifolia are the two most interesting
relatively strong antimicrobial activity against bioactive plants in this study and we selected
Gram-positive and Gram-negative bacteria them for further investigation of their active
including E. coli, B. subtilis, Staph. aurous constituents. The aqueous methanolic extracts of
and Staph. epidermis (26). These findings S. nemorosa and S. atropatana are suggested for
are similar to MIC values that we obtained determination of their antioxidant constituents
for the methanolic extract of this plant on the in the current paper. S. limbata, S. xanthocheila
same bacteria (Table 4). The antimicrobial and and S. sclarea are suggested to be analyzed for
antioxidant activity of methanolic extract of S. their antimicrobial constituents. Finally, since
aegyptiaca from Tunisia were evaluated and S. hypoleuca and S. syriaca are two plants with
found to be the most active one among other very rare sesterterpenes, they may also be good
Salvia species tested (30). candidates for evaluation of their terpenoids in
the cytotoxic bioassays.
Conclusion
Acknowledgements
The plants of the genus Salvia are rich
in antioxidant polyphenols (7, 31, 32) and We are grateful to the vice-chancellor for
abietane diterpenoids such as rosmarinic acid research, Shiraz University of Medical Sciences,
(7, 31, 33) and carnosol and carnosic acid Iran for financial support of this project (Grants
(34). The diterpenoids isolated from shoots numbers 88-4924 and 88-4678, 88-4923 and 88-
and roots of different Salvia species showed 4811).
considerable anticancer activity (35, 36) as
well as antimicrobial (1, 5, 8, 37) properties. References
Therefore we choose the above plants for
(1) Ulubelen A, Topcu G and Johansson CB.
screening their extracts for the above mentioned Norditerpenoids and diterpenoids from Salvia
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antimicrobial potential of various extracts of (1997) 60:1275-1280.
the sage plants indicate that we can use these (2) Demirci B, Tabanca N and Başer KHC. Enantiomeric
data to choose the appropriate extract of the distribution of some monoterpenes in the essential
oils of some Salvia species. Flav. Fragr. J. (2002)
plants for further purification and identification
17:54-58.
of their active ingredients. S. eremophila and (3) Jassbi AR, Asadollahi M, Masroor M, Schuman MC,

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 Cytotoxic, Antioxidant and Antimicrobial Activities of Phenolic Contents

Mehdizadeh Z, Soleimani M and Miri R. Chemical (19) Waterhouse AL. Determination of Total Phenolics.
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