3 Biotech

DOI 10.1007/s13205-014-0266-1

ORIGINAL ARTICLE

Antibacterial, antioxidant, antifungal and anti-inflammatory

activities of crude extract from Nitraria schoberi fruits
Javad Sharifi-Rad Seyedeh Mahsan Hoseini-Alfatemi
Majid Sharifi-Rad Jaime A. Teixeira da Silva

Received: 3 September 2014 / Accepted: 31 October 2014

The Author(s) 2014. This article is published with open access at Springerlink.com

Abstract This study is the first comprehensive investigation of the antibacterial, antioxidant, antifungal and antiinflammatory activities of a crude extract from Nitraria
schoberi L. (Zygophyllaceae) fruits. The extract was tested
against four Gram-negative (Pseudomonas aeruginosa,
Enterobacter aerogenes, Klebsiella pneumoniae and Acinetobacter lwoffii) and one Gram-positive (Staphylococcus
aureus) bacteria using the agar disc diffusion and microdilution methods. P. aeruginosa was inhibited the most
(widest inhibition zone) while K. pneumonia showed the
largest MIC value. The antioxidant activity of fruits
(0.02 mg/mL vs. 0.04, 0.06 and 1.00 mg/mL for atocopherol, butylated hydroxyanisole and ascorbic acid,
respectively) was determined by the paired diene method.
The antifungal activity of N. schoberi fruits was tested
against different fungi, including Aspergillus niger and

J. Sharifi-Rad
Zabol Medicinal Plants Research Center,
Zabol University of Medical Sciences, Zabol, Iran
J. Sharifi-Rad (&)
Department of Pharmacognosy, Faculty of Pharmacy,
Zabol University of Medical Sciences, Zabol, Iran
e-mail: javad.sharifirad@gmail.com
S. M. Hoseini-Alfatemi (&)
Pediatric Infections Research Center, Mofid Children Hospital,
Shahid Beheshti University of Medical Sciences, Tehran, Iran
e-mail: m.hoseinialfatemi@gmail.com
M. Sharifi-Rad
Department of Range and Watershed Management, Faculty
of Natural Resources, University of Zabol, Zabol, Iran
J. A. Teixeira da Silva
Miki-cho Post Office Retired, Ikenobe 3011-2,
P.O. Box 7, Kagawa-ken 761-0799, Japan

Candida albicans, with 300 lg/mL of fruit extract being

the most effective concentration. The percentage of antiinflammatory activity assayed for N. schoberi fruit extract
at 100, 200 and 500 lg/mL was 36.12, 59.89 and 88.33 %,
respectively. N. schoberi fruits possess potent antibacterial,
antioxidant, antifungal and anti-inflammatory properties,
and may be used as an antibacterial and antifungal to treat
diseases and/or as a protective agent against disorders
associated with oxidative stress and inflammation.
Keywords Agar disc diffusion method  Antifungal
activity  Anti-inflammatory activity  Antioxidant activity 
Antimicrobial activity  Zygophyllaceae
Abbreviations
BHA
Butylated hydroxyanisole
DMSO Dimethyl sulfoxide
MIC
Minimal inhibitory concentration

Introduction
Plants play a significant role in supplying food for humans.
Medicinal and aromatic plants have played a critical role as
therapeutic agents for a long time and thus hold great
economic value (Balunas and Kinghorn 2005). There is a
rising tendency to use natural drugs and herbal therapies,
partly because of the destructive nature of and side effects
of chemical drugs, as well as environmental pollution
(Balunas and Kinghorn 2005; Rad et al. 2013a; Sharifi-Rad
et al. 2014). Many naturally occurring agents in plant
extracts show antimicrobial, antioxidant, antifungal, anticancer and anti-inflammatory potential in several animal

123

3 Biotech

models and bioassay systems, and are pertinent to human

disease (Patwardhan 2005; Rad et al. 2014a).
Nitraria schoberi L. (Zygophyllaceae) is a medical
plant. Nitraria plants, specifically the leaves, fruits and
seeds, are often used in folklore medicine as an antispasmodic, and contain several classes of secondary metabolites including sterols, fatty acids, alkaloids and flavonoids
derivatives (Suo and Wang 2010; Hadj et al. 2011). The
aims of this study were to investigate the antimicrobial
activity of a fruit extract against four Gram-negative and
one Gram-positive bacteria as well as its antifungal and
anti-inflammatory activities.

Microorganisms
The microorganisms used in this study were obtained from
the microbiological laboratory of MRI Hospital in Shiraz
(Iran). These bacteria were isolated from MRI patients that
provided written permission to do so. Four Gram-negative
(Pseudomonas aeruginosa, Enterobacter aerogenes, Klebsiella pneumoniae and Acinetobacter lwoffii) and one
Gram-positive (Staphylococcus aureus) bacteria were
inoculated onto nutrient agar slants at 37 C and maintained at -80 C.
Antibiotic discs and microdilution assay

Materials and methods

Preparation of Nitraria schoberi fruit extracts
Fresh fruits of N. schoberi plants (Fig. 1a, b) were collected
from Gonabad (coordinates: 34210 1000 N, 58410 0100 E),
Razavi Khorasan Province, Iran in June 2013. The plant was
taxonomically identified by a botanist at the herbarium of
Pharmacognosy, Department of the Faculty of Pharmacy
affiliated to Shahid Beheshti University of Medical Sciences of Iran. All fruits were thoroughly washed in distilled
water three times then dried in an oven at 75 C for 48 h.
The dried fruits were powdered by a mechanical grinder and
then 20 g of powdered was dissolved in 200 mL of 85 %
methanol using a shaking water bath (50 rpm) for 24 h at
room temperature. After filtering through Whatman No. 1
filter paper, the filtrate was concentrated with a rotary
evaporator (Laborota 4000, Heidolph, Germany) at 45 C
for 35 min to remove solvent from the extract. The solidlike material that precipitated (i.e., the extract) was stored at
4 C until further analysis.

Antimicrobial activity was based on the disc diffusion

method (Thitilertdecha et al. 2008) utilizing a separate cell
suspension of each microorganism. The concentration of
each cell suspension was equilibrated to a 0.5 McFarland
standard and 50 lL of each microorganisms suspension
was spread on a MuellerHinton agar plate. In addition,
50 lL of diluted fruit extract (1 g/5 mL of distilled water)
was pipetted onto sterile blank discs (6 mm in diameter),
which were allowed to dry in an open sterile Petri dish in a
biological laminar flow bench. Discs were placed on the
surface of inoculated plates and incubated at 37 C for
24 h. Diameters (mm) of the zones of bacterial inhibition
minus the discs diameter were recorded (Aureli et al.
1992). Cephalosporin discs (Padtan Teb Co., Iran) were
used as positive controls. Cephalosporin is an antibiotic
with antibacterial activity that inhibits cell wall synthesis
although some bacteria such as Klebsiella spp., Proteus
spp. and Pseudomonas sp. are resistance to it (Wright
2010).
The minimal inhibitory concentration (MIC) values of
the plant extract versus each investigated microbial strain

Fig. 1 Nitraria schoberi plant used for extract analyses in this study. a Wild population; b Close-up of plant

123

3 Biotech

were determined by the microdilution assay in 96 multiwell microtiter plates according to the standard procedure
of the Clinical and Laboratory Standards Institute (CLSI
2010). All assays were carried out in MuellerHinton Broth
(MHB). The plant extract were dissolved in 5 % dimethyl
sulfoxide (DMSO) to a final concentration of 10 mg/mL.
Each strain was assayed with samples that were serially
diluted in broth to obtain concentrations ranging from
512.0, 256, 128, 64, 32, 16, 8, 4, 2, 1, 0.5, 0.25, 0.12 and
0.06 lg/mL. Overnight broth cultures of each strain were
prepared and the final microorganism concentration in each
well was adapted to 106 CFU/mL. The optimal incubation
conditions were 37 C for 24 h. The antimicrobial activity
to the plant extracts was classified according to the MIC
values as follows: MIC [1,000 lg/mL, no antimicrobial
activity; 512 B MIC B 1,000 lg/mL, mild activity; 128
B MIC \ 512 lg/mL, moderate activity; 32 B MIC \
128 lg/mL, good activity; 10 B MIC \ 32 lg/mL, strong
activity; and MIC \10 lg/mL, very strong activity (CLSI
2010). The experiment was carried out in duplicate and the
results were expressed as average values.
Antioxidant activity of fruits
The antioxidant activity of fruits was determined by the
paired diene method (Lingnert et al. 1979). The antioxidant
activity measured is the capacity of the extract to inhibit the
peroxidation of linoleic acid in which the double bond is
altered to a paired diene. Each extract sample (0.0130 mg/
mL) in methanol (100 lL) was mixed with 3 mL of 10 mM
linoleic acid (Sigma Chemical Co., St. Louis, MO, USA) to
form an emulsion in 0.2 M sodium phosphate buffer (pH 6.6)
in test tubes and placed in the dark at 37 C to hasten oxidation. After incubation for 15 h, 7 mL of 65 % methanol in
deionized water was added, and the absorbance of the mixture
was measured at 234 nm against a blank in a Hitachi U-2001
spectrophotometer (Tokyo, Japan). Antioxidant activity was
quantified as follows: antioxidant activity (%) = [(DA234 of
control DA234 of sample)/DA234 of control] 9 100. Analyses were repeated three times. a-Tocopherol, butylated
hydroxyanisole (BHA) and ascorbic acid (Sigma) were used
as standard controls. Antioxidant activity was expressed as
the EC50 value (mg/mL), which is the effective concentration
at which the antioxidant activity was inhibited by 50 %,
gained by interpolation from linear regression analysis.
Antifungal activity
Antifungal activity of N. schoberi fruits extracts was
assayed against two fungi (Aspergillus niger ATCC 9142
and Candida albicans ATCC 10231). The fungi were
cultured at 37 C for 1424 h and the densities were
adjusted to 0.5 McFarland standards at A530 nm (108

colony-forming units (CFU)/mL). The antifungal assays

were carried out by the disc diffusion method (Bauer et al.
1996). 100 lL of the microbial suspensions (108 CFU/mL)
was spread on nutrient agar (Merck, Germany) plates
(100 mm 9 15 mm). Discs (6 mm diameter) were
impregnated with 10 lL of different concentrations of
extract (50, 100, 150, 200, 250 and 300 lg/mL) and placed
on the inoculated agar. All the inoculated plates were
incubated at 37 C for 24 h. Positive control discs comprised ketoconazole (50 mcg/disc) (Padtan Teb Co., Iran)
for fungi. In addition, two microliters of 5 % (v/v) DMSO
was used as the negative control. Antifungal activity was
determined by measuring the zone of inhibition (in mm).
MIC was determined using serial dilutions of the extract
(5120.05 lg/mL) using the microdilution test confirmed
by Clinical and Laboratory Standards Institute (Wayne
2006). The fungal strains were suspended in LuriaBertani
medium and the densities were regulated to 0.5 McFarland
standards at 570 nm (108 CFU/mL). Fungal suspensions
(100 lL) and extracts were added to microtiter plates and
incubated at 37 C for 24 h. The sterile control was medium without fungi while the growth control was medium
with fungi, but without extract. The growth in each well
was compared with that in the control well. MICs were
visually detected in comparison with growth of the control
well and delineated as the lowest concentration of the
extract that inhibited growth.
Anti-inflammatory activity
The anti-inflammatory activity of N. schoberi fruits was
tested by the protein denaturation method as described by
Padmanabhan and Jangle (2012). Briefly, the reaction
included 1 mL of various concentrations of N. schoberi
fruits extracts (100, 200 and 500 lg/mL in distilled water)
and 3 mL of phosphate-buffered saline (pH 6.5) which
were blended with 2 mL of egg albumin and incubated at
25 C for 15 min. A denaturation reaction was induced in a
65 C water bath for 12 min. After cooling, absorbance
was measured at 660 nm (A660) with a Shimadzu A160
spectrofluorometer (Shimadzu, Japan) using double distilled water as the blank. The percentage inhibition of
protein denaturation was appraised by the following formula (Padmanabhan and Jangle 2012):
% inhibition As  Ac =Ac   100
where As and Ac represent sample and control
absorbance, respectively.
In this assay, diclofenac sodium (Padtan Teb Co., Iran),
a powerful non-steroidal anti-inflammatory drug, was used
as the standard drug in the same concentrations as N.
schoberi fruit methanol extracts (100, 200 and 500 lg/mL
in distilled water).

123

3 Biotech

and 25 lg/mL, respectively. This result confirmed the disc

diffusion data since minimum MIC and maximum MIC
were related to P. aeruginosa and A. lwoffii, respectively
(Fig. 2).
Bacteria that are resistant to antibiotics may jeopardize
medicine. Some such antibiotic-resistant bacteria have
greater importance than others, including S. aureus, A.
lwoffii, E. aerogenes, K. pneumonia and P. aeruginosa
(Chopra 2007). S. aureus, a Gram-positive bacterium, is a
highly changeable pathogen in response to antibiotics with
considerable importance in human medicine (Alfatemi
et al. 2014). It is responsible for a wide range of hospital
and community-acquired infections globally, ranging from
skin infections and food poisoning to life-threatening situations such as toxic-shock syndrome, endocarditis,
pneumonia, bacteremia and osteomyelitis (Kim et al. 2006;
Pottinger 2013; Ray et al. 2013; Zecconi and Scali 2013).
A. lwoffii is widespread, and a comparatively harmless
organism with the ability to persist in a hospital environment for prolonged periods (Rathinavelu et al. 2003; Garn
et al. 2013). Enterobacter aerogenes causes bacterial
infections, and is usually acquired in a hospital or in hospital-type atmospheres. Enterobacter aerogenes usually
brings about opportunistic infections, implying that it will
usually only cause an illness in a person or host that has a
compromised immune system although studies are now
showing that it is causing increased alarm in community
infections even though it seldomly causes disease in a
person with a healthy immune system (Lavigne et al.
2013). Klebsiella pneumonia, responsible for nosocomial
pneumonia in immunoincompetent people, produces
approximately 10 % of all infections that are acquired in
hospitals including urinary tract infections, pneumonia,
surgical wounds, and biliary tract wounds (Tam 2013). The
most important Gram-negative bacterium is P. aeruginosa,
which has become a significant cause of infection, particularly in patients with compromised host defense

Statistical analysis
The extract was prepared in triplicate for antibacterial,
antioxidant, antifungal and anti-inflammatory activities
tests. Data were subjected to analysis of variance following
a completely random design to determine the least significant difference (LSD) at P \ 0.05 using SPSS v. 11.5
(IBM SPSS, New York, USA).

Results and discussion

Antibacterial activity
The fruits of N. schoberi exhibit varying degrees of inhibition against all bacteria tested (Table 1). Inhibition zones
of A. lwoffii, E. aerogenes, K. pneumonia, S. aureus and P.
aeruginosa were 7, 18, 14, 16 and 25 mm, respectively,
indicating that N. schoberi fruit extract has a maximum
effect on P. aeruginosa and a minimum effect on A. lwoffii.
P. aeruginosa and A. lwoffii were resistant to cephalosporin
(control drug), but sensitive to N. schoberi fruit extract
(Table 1). The MIC of A. lwoffii, E. aerogenes, K. pneumonia, S. aureus and P. aeruginosa was 138, 40, 180, 95
Table 1 Mean diameters of inhibition zone with Nitraria schoberi
methanol fruit extract and two positive controls
Microorganism

Inhibition zone diameter (mm)

Fruit extract

Acinetobacter lwoffii

7 0.05

Cephalosporin
0 0.00

Enterobacter aerogenes

18 0.17

5 0.57

Klebsiella pneumoniae

14 0.08

0 0.00

Staphylococcus aureus

16 1

4 0.21

Pseudomonas aeruginosa

25 0.57

0 0.00

Values are mean of SE of three replicates

450
400

350
300

MIC (g)

Fig. 2 MIC values of Nitraria

schoberi methanolic fruit
extract against five tested
bacteria. Values are mean of
SE of three replicates. Means
with different letters within a
column are significantly
different (P \ 0.05; LSD)

250
200

150

100

50

0
Acinetobacter
lwoffii

Enterobacter
aerogenes

Klebsiella
pneumoniae

Microorganism

123

Staphylococcus
aureus

Pseudomonas
aeruginosa

3 Biotech

mechanisms (Tepe et al. 2004; Jeyaraj et al. 2013). Pseudomonas aeruginosa is the main and usual pathogen isolated from patients who have been hospitalized for longer
than 1 week (Ahmed et al. 2013). It is a frequent cause of
nosocomial infections such as pneumonia, urinary tract
infections, and bacteremia. Pseudomonal infections are
complicated and can be life threatening. Gram-positive
bacteria such as S. aureus are primarily responsible for
post-operative wound infections, toxic-shock syndrome,
endocarditis, osteomyelitis and food poisoning (OMalley
et al. 2009).
Studies on the discovery of natural antibacterial
sources from plants are increasing. Ahmad and Beg
(2001) studied the ethanolic extracts of 45 Indian
medicinal plants customarily used for their antimicrobial activity against specific drug-resistant bacteria and
a yeast C. albicans of clinical origin. 40 of these plant
extracts showed different levels of antimicrobial
activity against one or more of the assayed bacteria,
anticandidal activity was observed in 24 plant extracts
while broad-spectrum antimicrobial activity was discovered in 11 plants (Lawsonia inermis, Eucalyptus sp.,
Holarrhena antidysenterica, Hemidesmus indicus,
Casuarina equistifolia, Terminalia belerica, T. chebula, Emblica officinalis, Camelia sinensis, Syzgium
aromaticum, and Punica granatum). No similarity was
observed between the vulnerability of test strains with
plant extracts and antibiotic resistance of the microbial
strains (Salmonella paratyphi, Shigella dysenteriae, S.
aureus, Bacillus subtilis, Escherichia coli, C. albicans).
Qualitative phytochemical examinations, thin layer
chromatography (TLC) and a TLC-bioautography of
specific active extracts exhibited the presence of
ordinary phytocompounds in the plant extracts,
including phenols, tannins and flavonoids as the major
active components.
Mabona et al. (2013) investigated the antimicrobial
characteristics of southern African medicinal plants against
dermatologically relevant pathogens. Plants exhibiting
noteworthy broad-spectrum activities (MIC values
B1.00 mg/mL) against the assayed pathogens comprised
extracts from Diospyros mespiliformis, Aristea ecklonii,
Chenopodium ambrosioides, Eucalyptus camaldulensis,
Elephantorrhiza elephantina, Gunnera perpensa, Harpephyllum caffrum, Melianthus comosus, Hypericum perforatum, Warburgia salutaris and Terminalia sericea. The
organic extract of E. elephantina, a plant reportedly used to
treat common acne (Propionibacterium acnes), demonstrated
considerable
antimicrobial
activity
(MIC = 0.05 mg/mL) against P. acnes. Similarly, D.
mespiliformis, traditionally used to treat ringworm, also
displayed antimicrobial activity against Trichophyton
mentagrophytes (MIC = 0.10 mg/mL) and Microsporum

canis (MIC = 0.50 mg/mL). The aqueous root extracts of

P. prunelloides combined with E. elephantina in a 1:1 (v/v)
ratio showed synergistic interactions against S. aureus,
gentamycinmethicillin resistant S. aureus, Staphylococcus
epidermidis and C. albicans. Fractionation of A. ecklonii
resulted in the separation of plumbagin, exhibiting
remarkable antimicrobial activity in which the MIC ranged
from 2.00 to 16.00 lg/mL. Rad et al. (2013b) evaluated the
antibacterial activity of stem and flower essential oils from
Sinapis arvensis on five pathogenic multidrug-resistant
bacteria strains by the agar disc diffusion method. The
Gram-positive and Gram-negative bacteria that were most
susceptible to the stem and flower essential oils of S. arvensis were S. aureus (NCTC7428) and P. aeruginosa
(MTCC 2453), respectively.
Previous studies have shown that antibacterial activity is
caused by various classes of secondary metabolites,
including fatty acids, sterols, alkaloids and flavonoids
derivatives, flavonoids and phenolic compounds (Tulyaganov and Allaberdiev 2003; Suo and Wang 2010; Hadj
et al. 2011). In earlier studies, it was determined that N.
sibirica fruits contained phenolic compounds (Senejoux
et al. 2012).
Antioxidant activity
The results for antioxidant activity of the extract tested are
summarized in Table 2. The efficiency of antioxidant
activity is inversely related with the extracts EC50 values.
The antioxidant activity, i.e., EC50 values, was 0.04, 0.06,
1.00 and 0.02 mg/mL for a-tocopherol, butylated
hydroxyanisole (BHA), ascorbic acid and N. schoberi
fruits, respectively (Table 2). The fruit extract was not
significantly different to a-tocopherol and BHA
(P \ 0.05), but only significantly different to ascorbic acid
(P \ 0.05).
Rad et al. (2014b) investigated the free radical-scavenging and antioxidant activities of different parts of N.
schoberi plants collected from Zabol, Sistan and Baluchestan Province, Iran. They showed that the maximum
free radical and antioxidant activities were associated with

Table 2 EC50 values (mg/mL) of the Nitraria schoberi methanolic

fruit extract in two assays
Antioxidant activity
Fruit extract

0.02 0.00 b

a-Tocopherol

0.04 0.00 b

BHA

0.06 0.02 b

Ascorbic acid

1.00 0.00 a

Values are mean of SE of three replicates. Means with different

letters within a column are significantly different (P \ 0.05; LSD)

123

3 Biotech

the methanolic extracts rather than with other extracts

(aqueous and chloroform). These activities were maximum
in fruits, and lower in leaves and roots. The methanolic,
aqueous and chloroform extracts showed 86, 70 and 57 %
scavenging activity, respectively. In this study, we investigated the fruits of N. schoberi collected from Gonabad,
Razavi Khorasan Province, Iran, and also observed antioxidant activity, indicating that this activity remains constant, independent of the geographic location. N. schoberi
may be considered a broad and effective source of healthy
antioxidants and bioactive phytochemicals.
Choi et al. (2002) evaluated the root bark and leaves of
several Korean medicinal plants to appraise their free
radical-scavenging capacity and antioxidant activity using
common assays with dichloromethane, ethanol or methanol
extracts. Flavonoids, including catechin, morin, naringenin,
quercetin and rutin, were included and used as standards.
Among the plant extracts, the root bark of Morus alba L.
and the leaves of Saururus chinensis showed stronger
values than other plant extracts.
Bouayed et al. (2007) conducted a comparative study
between the leaves of Lavandula officinalis L., the roots
of Verbena officinalis L., the flowers of Calylophus
lavandulifolius, the leaves of Melissa officinalis L. and
the flowers of Althea kurdica plants from the same
geographic origin, the Hamadan region in the west of
Iran and growing in the same natural conditions. The
antioxidant activities differed significantly between all
plant parts while some plants were rich in natural antioxidants, especially the leaves of L. officinalis and M.
officinalis.
Nivas et al. (2010) investigated the methanolic extract of
young and mature leaves of nine coastal plants from the
West coast of Maharashtra, India to appraise their free
radical-scavenging activity. The highest antioxidant activity was recorded in young leaves of Hibiscus tiliaceus L.
(76 %), followed by Syzigium corymbosa (71 %), Calophyllum inophyllum L. (68 %) and Colubrina asiatica (L.)
Brongn. (55 %). The leaves of all these species were rich in
flavonoids (6.0316.63 mg/g of dry weight) and total
polyphenols (12.1226.23 mg/g of dry weight). These
compounds mainly contributed to the antioxidant potential
of these plants.
The above evidence from the literature suggests that
some plants have potent antioxidant activity and free radical-scavenging properties. Plant phenolic compounds have
powerful antioxidant activity and may assist in the protection of cells against oxidative damage caused by free
radicals, which, with other reactive oxygen species, are a
causative factor of different diseases such as asthma,
mongolism, arthritis, carcinoma, dementia and Parkinsons
disease (Perry et al. 2000).

123

Table 3 Antifungal activity of Nitraria schoberi fruit extracts

against two fungal strains
Fruit extract (lg/mL)
50

Aspergillus niger Candida albicans

2.1 0.0

1.0 0.0

100

5.6 0.0

2.5 0.2

150

9.2 0.1

6.3 0.0

200

14.5 0.0

13.8 0.0

250

15.7 0.0

15.1 0.0

300

18.2 0.7

17.9 0.0

DMSO (negative control)

Ketoconazole (positive control)

1.0 0.0
14.7 0.0

0.5 0.0
21.1 0.0

284.5 0.3

342 0.0

MIC

Data are expressed as mean SE of inhibition zone diameter (mm)

for different concentration of extract, controls and minimum inhibitory concentration (MIC) (lg/mL)
DMSO Dimethyl sulfoxide

Antifungal activity
The antifungal activity of N. schoberi fruit extract against
two fungi strains is shown in Table 3. Inhibition zones of A.
niger in response to 50, 100, 150, 200, 250 and 300 lg/mL
of fruit extract were 2.1, 5.6, 9.2, 14.5, 15.7 and 18.2 mm,
respectively. Inhibition zones of C. albicans in response to
50, 100, 150, 200, 250 and 300 lg/mL of fruit extract were
1, 2.5, 6.3, 13.8, 15.1 and 17.9 mm, respectively. DMSO
(negative control) and ketoconazole (positive control)
showed inhibition zones of 1.0 mm and 14.7 mm, respectively, for A. niger or 0.5 mm and 21.1 mm, respectively,
for C. albicans. The MIC of A. niger and C. albicans was
284 lg/mL and 342 lg/mL, respectively.
In the past decade, the prevalence of resistance to antifungal agents has increased. Resistance to antifungal agents
has important implications for healthcare, morbidity and
mortality in human life. Moreover, a search for new, more
secure and potent agents to combat critical fungal infections is required. Plants, especially medicinal plants, are as
a good candidate for this aim. In recent years, several
studies have been reported on the antifungal activity of
phenols, flavonoids, coumarins, quinones, saponins, xanthones, alkaloids, lectins, polypeptides, terpenoids and
essential oils from natural sources (Arif et al. 2009). Previous studies showed the presence of secondary metabolites, including alkaloids and flavonoids derivatives, in the
Zygophyllaceae (Tulyaganov and Allaberdiev 2003; Hadj
et al. 2011).
Anti-inflammatory activity
The results of anti-inflammatory assay of N. schoberi fruit
extract are shown in Fig. 3: 36.12, 59.89 and 88.33 %,

3 Biotech
120

Inhibition of protein
denaturation (%)

Fig. 3 Anti-inflammatory
effect of Nitraria schoberi fruit
extract. Values are mean SE
of three replicates; means with
different letters within a column
are significantly different
(P \ 0.05; LSD)

a
b

100

80

e
60
40

20
0
Plant extract / 100 Plant extract / 200 Plant extract / 500

Diclofenac / 100

Diclofenac / 200

Diclofenac / 500

Treatments/concentrations (/mL)

respectively, for 100, 200 and 500 lg/mL of N. schoberi

fruit extract. The anti-inflammatory percentage of diclofenac (control) at 100, 200 and 500 lg/mL was 90.31, 97.33
and 109.13 %, respectively. The percentage of inhibition of
protein denaturation by N. schoberi fruit extracts was significantly different in all treatments (P \ 0.05).
Inflammation is clinically defined as a pathophysiological procedure described by redness, fever, edema, loss of
function and pain (Hyun et al. 2004). Flavonoids possess
several biological and pharmacological activities: antimicrobial, anti-inflammatory, immunomodulatory, antiviral,
anticancer and antithrombotic activities (Havsteen 1983).
Many studies verified that flavonoids possess anti-inflammatory activity in several inflammation animal models.
Flavonoids can regulate cellular activities of inflammationrelated cells such as macrophages, lymphocytes, neutrophils and mast cells. For example, some flavonoids inhibit
T cell proliferation while others inhibit the release of histamine from mast cells (Hyun et al. 2004).

Conclusion
Nitraria schoberi fruits show promising use as new pharmaceuticals with antibacterial, antioxidant, antifungal and
anti-inflammatory activities. Even so, organic compounds
and active agents in the fruits need to be identified for the
plant to be used as herbal drug while the toxicity of the
active components, serum-attainable levels, pharmacokinetic attributes, their side effects, and diffusion in various
sites around the body also need to be determined. Nitraria
schoberi fruits decreased and inhibited the growth of food
pathogens. Therefore, use of this plant might decrease food
poisoning and increase food shelf life.
Acknowledgments The authors are very grateful to Department of
Range and Watershed Management, Faculty of Natural Resources,
University of Zabol for supports in this study.

Conflict of interest The authors declare no financial or other conflicts of interest.

Open Access This article is distributed under the terms of the
Creative Commons Attribution License which permits any use, distribution, and reproduction in any medium, provided the original
author(s) and the source are credited.

References
Ahmad I, Beg AZ (2001) Antimicrobial and phytochemical studies on
45 Indian medicinal plants against multi-drug resistant human
pathogens. J Ethnopharmacol 74(2):113123. doi:10.1016/
S0378-8741(00)00335-4
Ahmed Z, Khan SS, Khan M (2013) In vitro trials of some
antimicrobial combinations against Staphylococcus aureus and
Pseudomonas aeruginosa. Saudi J Biol Sci 20(1):7983. doi:10.
1016/j.sjbs.2012.10.005
Alfatemi SMH, Rad JS, Rad MS, Mohsenzadeh S, Teixeira da Silva
JA (2014) Chemical composition, antioxidant activity and
in vitro antibacterial activity of Achillea wilhelmsii C. Koch
essential oil on methicillin-susceptible and methicillin-resistant
Staphylococcus aureus spp. 3 Biotech. doi: 10.1007/s13205-0140197-x
Arif T, Bhosale JD, Kumar N, Mandal TK, Bendre RS, Lavekar GS,
Dabur R (2009) Natural products- antifungal agents derived from
plants. J Asian Nat Prod Res 11(7):621638. doi:10.1080/
10286020902942350
Aureli P, Constantini A, Zolea S (1992) Antimicrobial activity of
some plant essential oils against Listeria monocytogenes. J Food
Prot 55:344348
Balunas MJ, Kinghorn AD (2005) Drug discovery from medicinal
plants. Life Sci 78(5):431441. doi:10.1016/j.lfs.2005.09.012
Bauer AW, Kirby WMM, Sherris JC, Turck M (1996) Antibiotic
susceptibility testing by a standardized single disk method. Am J
Clin Pathol 45(4):493496
Bouayed J, Piri K, Rammal H, Dicko A, Desor F, Younos C,
Soulimani R (2007) Comparative evaluation of the antioxidant
potential of some Iranian medicinal plants. Food Chem
104(1):364368. doi:10.1016/j.foodchem.2006.11.069
Choi CW, Kim SC, Hwang SS, Choi BK, Ahn HJ, Lee MY, Park SH,
Kim SK (2002) Antioxidant activity and free radical scavenging
capacity between Korean medicinal plants and flavonoids by
assay-guided comparison. Plant Sci 163(6):11611168. doi:10.
1016/S0168-9452(02)00332-1

123

3 Biotech
Chopra I (2007) The increasing use of silver-based products as
antimicrobial agents; a useful development or a concern. J Antimicrob Chemother 59(4):587590. doi:10.1093/jac/dkm006
CLSI (2010) Performance standards for antimicrobial susceptibility
testing: 20th informational supplement. CLSI document M100S20. Clinical and Laboratory Standards Institute, Wayne
Garn H, Neves JF, Blumberg RS, Renz H (2013) Effect of barrier
microbes on organ-based inflammation. J Allergy Clin Immunol
131(6):14651478. doi:10.1016/j.jaci.2013.04.031
Hadj SJ, Chevalot I, Harscoat-Schiavo C, Paris C, Fick M, Humeau C
(2011) Biological activities of flavonoids from Nitraria retusa
(Forssk.) Asch. and their acylated derivatives. Food Chem
124(2):486494. doi:10.1016/j.foodchem.2010.06.059
Havsteen B (1983) Flavonoids, a class of natural products of high
pharmacological
potency.
Biochem
Pharmacol
32(7):11411148. doi:10.1016/0006-2952(83)90262-9
Hyun E, Bolla M, Steinhoff M, Wallace JL, Soldato PD, Vergnolle N
(2004) Anti-inflammatory effects of nitric oxide-releasing hydrocortisone NCX 1022, in a murine model of contact dermatitis. Br J
Pharmacol 143(5):618625. doi:10.1038/sj.bjp.0705854
Jeyaraj M, Varadan S, Anthony KJP, Murugan M, Raja A,
Gurunathan S (2013) Antimicrobial and anticoagulation activity
of silver nanoparticles synthesized from the culture supernatant
of Pseudomonas aeruginosa. J Ind and Eng Chem
19(4):12991303. doi:10.1016/j.jiec.2012.12.031
Kim JS, Song W, Kim HS, Cho HC, Lee KM, Choi MS, Kim EC
(2006) Association between the methicillin resistance of clinical
isolates of Staphylococcus aureus, their staphylococcal cassette
chromosome mec (SCCmec) subtype classification and their
toxin gene profiles. Diagn Microbiol Infect Dis 56(3):289295.
doi:10.1016/j.diagmicrobio.2006.05.003
Lavigne JP, Sotto A, Nicolas-Chanoine MH, Bouziges N, Page`s JM,
Davin-Regli A (2013) An adaptive response of Enterobacter
aerogenes to imipenem: regulation of porin balance in clinical
isolates. Int J Antimicrob Agents 41(2):130136. doi:10.1016/j.
ijantimicag.2012.10.010
Lingnert H, Vallentin K, Eriksson CE (1979) Measurement of
antioxidative effect in model system. J Food Process Pres
3(2):87103. doi:10.1111/j.1745-4549.1979.tb00574.x
Mabona U, Viljoen A, Shikanga E, Marston A, Van Vuuren S (2013)
Antimicrobial activity of southern African medicinal plants with
dermatological relevance: From an ethnopharmacological
screening approach, to combination studies and the isolation of
a bioactive compound. J Ethnopharmacol 148(1):4555. doi:10.
1016/j.jep.2013.03.056
Nivas D, Sonar BA, Shaikh SS, Patil UH, Gaikwad DK, Chavan NS,
Sabale AB, Chavan PD (2010) Screening of some coastal plant
resources for their antioxidant potential, total polyphenol and
flavonoid content. Pharmacognosy J 2(7):151156. doi:10.1016/
S0975-3575(10)80083-5
OMalley M, Fowler J, Ilyas AM (2009) Community-acquired
methicillin-resistant Staphylococcus aureus infections of the
hand: prevalence and timeliness of treatment. J Hand Surg
34(3):504508. doi:10.1016/j.jhsa.2008.11.021
Padmanabhan P, Jangle SN (2012) Evaluation of in vitro antiinflammatory activity of herbal preparation, a combination of
four herbal plants. Int J Basic Appl Med Sci 2(1):109116
Patwardhan B (2005) Ethnopharmacology and drug discovery. J Ethnopharmacol 100(12):5052. doi:10.1016/j.jep.2005.06.006
Perry G, Raina AK, Nunomura A, Wataya T, Sayre LM, Smith MA
(2000) How important is oxidative damage lessons from
Alzheimers disease. Free Radic Biol Med 28(5):831834.
doi:10.1016/S0891-5849(00)00158-1
Pottinger PS (2013) Methicillin-resistant Staphylococcus aureus
infections. Med Clin North Am 97(4):601619. doi:10.1016/j.
mcna.2013.02.005

123

Rad JS, Alfatemi SH, Rad MS, Iriti M (2013a) In-vitro antioxidant
and antibacterial activities of Xanthium strumarium L. extracts
on methicillin-susceptible and methicillin-resistant Staphylococcus aureus. Ancient Sci Life 33:109113. doi:10.4103/02577941.139050
Rad JS, Alfatemi SMH, Rad MS, Sen DJ (2013b) Phytochemical and
antimicrobial evaluation of the essential oils and antioxidant
activity of aqueous extracts from flower and stem of Sinapis
arvensis L. Am J Adv Drug Deliv 1(1):001010
Rad JS, Alfatemi SMH, Rad MS (2014a) In vitro assessment of
antibacterial activity of Salicornia herbacea L. seed extracts
against multidrug resistant gram-positive and gram-negative
bacteria. Int. J Biosci 4(6):217222. doi:10.12692/ijb/4.6.217-222
Rad JS, Alfatemi SMH, Rad MS, Iriti M (2014b) Free radical
scavenging and antioxidant activities of different parts of
Nitraria schoberi L. TBAP 4(1):4451
Rathinavelu S, Zavros Y, Merchant JL (2003) Acinetobacter lwoffii
infection and gastritis. Microb Infect 5(7):651657. doi:10.1016/
S1286-4579(03)00099-6
Ray GT, Suaya JA, Baxter R (2013) Microbiology of skin and soft
tissue infections in the age of community-acquired methicillinresistant Staphylococcus aureus. Diagn Microbiol Infect Dis
76(1):2430. doi:10.1016/j.diagmicrobio.2013.02.020
Senejoux F, Girard C, Aisa HA, Bakri M, Kerram P, Berthelot A,
Bevalot F, Demougeot C (2012) Vasorelaxant and hypotensive
effects of a hydroalcoholic extract from the fruits of Nitraria
sibirica Pall. (Nitrariaceae). J Ethnopharmacol 141(2):629634.
doi:10.1016/j.jep.2011.08.012
Sharifi-Rad J, Hoseini-Alfatemi SM, Sharifi-Rad M, Setzer WN
(2014) Chemical composition, antifungal and antibacterial
activities of essential oil from Lallemantia royleana (Benth. in
Wall.) Benth. J Food Safety. doi:10.1111/jfs.12139
Suo Y, Wang L (2010) Extraction of Nitraria tangutorum seed lipid
using different extraction methods and analysis of its fatty acids
by HPLC fluorescence detection and on-line MS identification.
Eur J Lipid Sci Technol 112(3):390399
Tam PYI (2013) Approach to common bacterial infections: community-acquired pneumonia. Pediatr Clin North Am 60(2):437453.
doi:10.1016/j.pcl.2012.12.009
Tepe B, Donmez E, Unlu M, Candan F, Daferera D, Vardar-Unlu G,
Polissioud M, Sokmen A (2004) Antimicrobial and antioxidative
activities of the essential oils and methanol extracts of Salvia
cryptantha (Montbret et Aucher ex Benth.) and Salvia multicaulis (Vahl). Food Chem 84(4):519525. doi:10.1016/S03088146(03)00267-X
Thitilertdecha N, Teerawutgulrag A, Rakariyatham N (2008) Antioxidant and antibacterial activities of Nephelium lappaceum L.
extracts. LWTFood. Sci Technol 41(10):20292035. doi:10.
1016/j.lwt.2008.01.017
Tulyaganov TS, Allaberdiev FK (2003) Alkaloids from plants of the
Nitraria genus. Structure of sibiridine. Chem Nat Compd
39(3):292293. doi:10.1023/A:1025482821368
Wayne PA (2006) Clinical and Laboratory Standards Institute,
CLSIM7-A7. 7th edn. Clinical and Laboratory Standards
Institute (CLSI). Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; approved standard
Wright GD (2010) Antibiotic resistance in the environment: a link to
the clinic? Curr Opin Microbiol 13(5):589594. doi:10.1016/j.
mib.2010.08.005
Zecconi A, Scali F (2013) Staphylococcus aureus virulence factors in
evasion from innate immune defenses in human and animal
diseases. Immunol Lett 150(12):1222. doi:10.1016/j.imlet.
2013.01.004