Efeitos da Malva sylvestris na colite ulcerativa experimental em ratos

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Original Article
Effects of Malva sylvestris and Its Isolated
Polysaccharide on Experimental Ulcerative
Colitis in Rats
Azadeh Hamedi, PhD1, Hossein Rezaei, PharmD1,
Negar Azarpira, MD1, Mehrnaz Jafarpour, PharmD1,
and Fatemeh Ahmadi, PhD1
Abstract
Malva sylvestris is an edible plant that is consumed as a herbal supplement for its antiulcer and colon cleansing properties in
traditional Persian medicine. This study was designed to evaluate its effects on ulcerative colitis, which is a chronic gastro-
intestinal inflammation. Colitis was induced by rectal instillation of acetic acid solution. Rats in different groups received
aqueous, n-hexane, or ethanolic fractions of the plant before induction of colitis. Isolated polysaccharide of plant was also
tested in 2 groups before and after induction of colitis. Macroscopic and microscopic evaluation of colitis showed that the
aqueous fraction was very effective in preventing the inflammation and efficacy was lower for ethanolic and n-hexane fractions.
Polysaccharide was effective in reducing signs of inflammation, especially as pretreatment. These beneficial effects provide
evidences that this plant can be suggested for patients with this disease to improve their health condition or to reduce adverse
effects of their medication.
Keywords
experimental colitis, inflammation, inflammatory bowel disease, Malva sylvestris, polysaccharide
Received February 15, 2015. Accepted for publication April 25, 2015.
Idiopathic inflammatory bowel diseases, which include Crohn’s
disease and ulcerative colitis, are chronic inflammation of the
gastrointestinal tract with a growing prevalence especially in
modern countries (150-200 cases per 100 000 individuals).1
Although the highest incidence of inflammatory bowel disease
has been reported in the populations of Northern Europe and
North America and the lowest in Asia, the prevalence is
increasing in developing countries as well.2,3 Treatment of
inflammatory bowel disease is still challenging because of poor
understanding of the etiology of the disease. Choosing treat-
ment regimens depends on the subtype and severity of the
disease. These regimens consist of aminosalicylates4 and glu-
cocorticoids in mild to moderate cases5 and immunosuppres-
sants such as azathioprine and cyclosporine A in severe
cases.6 Severe cases may also require surgery. Inflammatory
bowel disease may be associated with the risk of colorectal
malignancy and cancers and may decrease the quality of life
of patients. Poor treatment outcomes and high rate of side
effects of the current medications used for this disease has
encouraged researchers to examine alternative treatments for
inflammatory bowel disease. Nowadays, tremendous attention
is paid to medicinal plants, natural products, complementary
and alternative medicine, and functional foods to find beneficial
effects, drugs, or strategies for the treatment of inflammatory
bowel disease as monotherapy or adjuvant therapy.7-10
Malva sylvestris L (Malvaceae), also known as cheeses,
high mallow (in English), or panirak or khabbazi (in Persian),
is a biennial or perennial plant that grows wild in farms and on
waste ground.11,12 Malva sylvestris is an edible plant wherein
its boiled leaves are taken as wholesome vegetable in stews
or marinated in yogurt as a side dish. In Persian folk medicine,
the plant’s decoction and its cooked aerial parts are used for
treatment of gastrointestinal lesions, as tonic for gastrointest-
inal tract, or as a side dish especially for its colon cleansing
properties.13 Also, in some Persian traditional and pharmaceu-
tical manuscripts such as Qarabadin-e-azam (a lithograph
manuscript written by Hakim Azamkhan in 1853 AD), Qaraba-
din-e-kabir (Aghili Shirazi, 1772), Qarabadin-e-salehi (Heravi,
1765), and Tohfat ol Moemenin (Tonekaboni, 1670 AD),14
1 Shiraz University of Medical Sciences, Shiraz, Iran
Corresponding Author:
Fatemeh Ahmadi, PhD, Department of Pharmaceutics, School of Pharmacy,
Shiraz University of Medical Sciences, Shiraz 71345, Iran.
Email: ahmadi_f@sums.ac.ir
Journal of Evidence-Based
Complementary & Alternative Medicine
1-9
ª The Author(s) 2015
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DOI: 10.1177/2156587215589184
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several preparations containing Malva sylvestris leaves, flow-
ers, and fruits were explained as oral dosage forms, functional
food, and rectal enema for treatment of gastrointestinal and
rectal ulcers, pain, and/or inflammation.15 Recently, different
extracts of common mallow was reported to exhibit wound
healing, antioxidant, anti-inflammatory,12,16-19 and immuno-
modulatory effects.20-22 Also, different polyphenols, polysac-
charides, fatty acids, carotenoids, tocopherol, and ascorbic
acid in different amounts have been isolated from the edible
parts of the plant.21,23
Although beneficial gastrointestinal effects of Malva sylves-
tris have been approved in Persian traditional medicine, heal-
ing properties of this plant on colitis and colon inflammation
have not been investigated yet. With the aim of evaluating the
anti-inflammatory effect of this plant against induced colitis in
rats, aqueous, n-hexane, and ethanolic fractions and its isolated
polysaccharide (as possible active constituent) of the plant
were prepared and their efficacy on microscopic and macro-
scopic signs of colitis damage were studied and compared to
the control groups receiving prednisolone.
Materials and Methods
Materials
Prednisolone was obtained from Iran Hormone Pharmaceutical Com-
pany (Iran). All other solvents and chemicals were of analytical grade
and purchased from Merck (Darmstadt, Germany) or Sigma Aldrich
(Sigma, St Louis, MO).
Malva sylvestris Fractions
Malva sylvestris was collected from farms around Marvdasht (Fars,
Iran) in the flowering season (May 2012). The aerial parts were
shadow dried, milled, and preserved in well-covered container at
4�C. Authentication was performed by Miss Sedigheh Khademyan,
Taxonomist, Department of Pharmacognosy, Shiraz School of Phar-
macy, and its herbarium was preserved with the code PM-380 in the
department for further references.
A total of 200 g of powdered aerial parts was sequentially extracted
with n-hexane, ethanol, and water (3 � 1 L and 48 hours for each) at
room temperature. The fractions were concentrated by a rotary eva-
porator at 40�C and dried applying a speed vacuum dryer or a freeze
dryer and kept at �20�C.
Polysaccharide Isolation and Characterization
Aqueous fraction of Malva sylvestris was mixed with 4 volumes
of absolute ethanol, stirred vigorously, and left overnight at 4�C.
The precipitated polysaccharide was collected after centrifugation
(16 000 g, 20 minutes at 4�C). The sediment was freeze dried and
applied to an ion exchange column (equilibrated Sephadex A-25) and
eluted with gradient solutions of 0.02 to 2 M NaCl. Same fractions were
pooled and applied to a size exclusion column (Sepharose CL-6B). The
isolated polysaccharide was lyophilized and stored at �20�C.
To determine the monosaccharide content, 4 mg of the polysac-
charide was first hydrolyzed with trifluoroacetic acid (2 M) at
110�C for 2 hours and then tetramethylsilane derivatizated with
N,O-bis-(trimethylsilyl)acetamide. The monosaccharide content was
analyzed using a gas chromatography–mass spectrometry instrument
(7890A, Agilent). The gas chromatograph was equipped with a
HP-5MS capillary column (5% phenyl methyl siloxane, 30 m �
250 mm � 0.25 mm). The carrier gas was helium with a flow rate of
1.0 mL/min. The mass spectrometer (5975C MSD, Agilent) operated
in EI mode at 70 eV. The mass range was 30 to 600 m/z. Identification
of components was based on a comparison of the retention index and
mass spectra with Wiley (7 nL) andderivatized standards. The injec-
tion temperature was 150�C. Oven temperature was held at 150�C for
4.5 minutes, and then increased to 280�C (6�C/min). The temperature
was held at this temperature for 5 minutes. The protein content was
evaluated by Bradford assay and the uronic acids content was deter-
mined spectrophotometrically against standard galacturonic acid.
Molecular weight of the polysaccharide was determined as compared
with standard dextrans. A mixture of standard dextrans (667, 233, 80,
and 1 KDa) was applied on the same size-exclusion column and chro-
matographed with a same condition for the sample. The average mole-
cular weight of the isolated polysaccharide was calculated with
Nishitani formula: MW ¼
P
MiWi/
P
Wi, where MW is the average
molecular weight of the sample, Wi is the carbohydrate content in the
sample fraction, and Mi is the molecular weight of standard dextrans in
the corresponding fraction.24,25
Determination of Polyphenol Content of Fractions
A total of 50 mL of n-hexane was added to 2 g of dried fractions. Poly-
phenols were extracted with 60% methanol (3 � 100 mL). The col-
lected methanolic phase was washed with 20 mL of n-hexane and
dried with a rotary evaporator at 40�C.26,27 Polyphenols were dis-
solved by adding adequate amount of distilled water. The solution was
moved to a separation funnel and extracted with petroleum ether
(60:80). The water phase was saturated with NaCl and extracted with
4 volumes of ethyl acetate (3 times). The ethyl acetate fractions were
collected and dried by adding anhydrous Na2SO4 and dried with a
rotary evaporator at 40�C. Folin-Ciocalteau reagent (5 mL) was added
to 1 mL of different concentrations of gallic acid or sample tubes, and
after 5 minutes, 4 mL of 7.9% sodium carbonate solution was added.
Sample and standard tubes were covered for 2 hour at room tempera-
ture and away from strong light. Absorbance was read at 740 nm. All
experiments were done in triplicate.28
Animals
Male Wistar rats (190 + 20 g) were purchased from Razi Vaccine and
Serum Research Institute (Shiraz, Iran). The experiments were con-
ducted in accordance with ethical guidelines for animal experimenta-
tion approved by the ethical committee of Animal Laboratories in
Shiraz University of Medical Sciences. The rats (6 per group) were
housed in the Shiraz School of Pharmacy Animal Laboratory under
a 12-hour light–dark cycle, with room temperature maintained at
25�C, humidity at 55%, and food and water available ad libitum. The
rats were deprived of food, but not water, for 24 hours prior to experi-
mental procedures.
Induction of Colitis
Colitis was induced by rectal instillation of a 4% solution of acetic
acid according to the method reported earlier.29 Under light ether
anesthesia, an 8-cm-long polyethylene tube was inserted in the rectum
and through that acetic acid solution was instilled to the colon. After
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instillation of the solution, rats were kept in separate cages with free
access to water and rat chow pellet.
Administration of Drugs
Animals were divided into 8 groups, each containing 6 rats.
Drug administration was performed according to the following
protocol:
Normal group: without induction of colitis that received normal sal-
ine solution rectally at the day of induction.
Negative control group: with induction of colitis that received dis-
tilled water orally.
Positive control group: with induction of colitis that received
prednisolone at dose 5 mg/kg/day orally prior to induction of
colitis.
Treatment group I: with induction of colitis that received aqueous
fraction of Malva sylvestris at dose 200 mg/kg/day orally prior
to induction of colitis.
Treatment group II: with induction of colitis that received ethanolic
fraction of Malva sylvestris at dose 200 mg/kg/day orally prior
to induction of colitis.
Treatment group III: with induction of colitis that received n-hex-
ane fraction of Malva sylvestris at dose 200 mg/kg/day orally
prior to induction of colitis.
Treatment group IV: with induction of colitis that received the iso-
lated polysaccharide of Malva sylvestris at dose 200 mg/kg/day
orally prior to induction of colitis.
In groups I to IV treatment was performed for 5 days consecutively
before induction of colitis.
Treatment group V: with induction of colitis that received the iso-
lated polysaccharide of Malva sylvestris at dose 200 mg/kg/day
orally for 5 days after induction of colitis.
The dosage regimen for animal study was selected based on
our preliminary studies (data not shown).
Macroscopic and Microscopic Examination of Colon
Twenty-four hours after the last dose of the drug in treatment group V
and 24 hours after induction of colitis in other groups, rats were eutha-
nized with a high dose of ether and an incision was made to abdomen.
Eight-centimeter length of the end part of the colon was removed and
used for macroscopic and microscopic evaluations.
For macroscopic evaluation, ulcer area and colon weigh-to-length
ratio (mg/cm) were measured. A macroscopic scoring was also used to
present the severity of damage as follows29: normal appearance, 0;
erythema without ulcer, 1; inflammation and ulcer, 2; and severe ulcer
with necrosis, 3.
Colon samples were then fixed in 10% formalin solution, stained
by hematoxylin and eosin, and observed by light microscope. Severity
and extent of damage was examined and scored based on the para-
meters presented in Table 1.30
Statistical Analysis
Kolmogorov–Smirnov statistical test was used for controlling
normality and homogeneity of the resulting data. Colon weight-
to-length ratio and ulcer area in different groups was tested by
one-way analysis of variance and Dunnett’s post hoc test. Nonpara-
metric variables (ordinal variables of scoring) were compared by
Mann–Whitney U test. Statistical analysis was performed by SPSS
16 software, and P value less than 0.05 was considered significant
for all experiments.
Results
Phytochemical Analysis
The yield for polysaccharide extraction was 9.6% (w/w). The
gas chromatography–mass spectrometry analysis revealed that
the isolated polysaccharide has an acidic nature and consisted
of galactose, glucose, uronic acids, arabinose, and rhamnose
Table 1. Histopathologic Scoring of Colon Damage.30
Variables 0 1 2 3
Ulceration No ulcer Erosion or single ulceration not
exceeding lamina muscularis
mucosa
Multifocal ulcerations not exceeding
the submucosa
Ulcerations exceeding the
submucosa
Mucus cell
depletion
Preserved mucus cell Mild depletion in a few cells Moderate depletion (�50% of cells) Severe depletion or
complete disappearance
of mucosa
Crypt
abscesses
No abscesses 1-3 abscesses/slide 4-9 abscesses/slide 10 or more abscesses/slide
Inflammatory
cysts
No cyst 1-3 cysts/slide 4-9 cysts/slide 10 or more cysts/slide
Mucosal
atrophy
Normal thickness Mild atrophy (<10%) Moderate atrophy (10% to 50%) Severe atrophy (450%)
Edema
(submucosa)
Normal thickness Mild edema (submucosal
expansion <10%)
Moderate edema (submucosal
expansion, 10% to 100%)
Severe edema (submucosal
expansion >100%)
Inflammatory
cell
infiltration
No inflammatory cell
infiltration
Mild inflammatory cell infiltration Moderate (distributed but not dense)
inflammatory cell infiltration
Dense inflammatory cell
infiltration
Vascular
dilatation
Normal blood
vessels
Mild dilatation of single blood
vessel
Moderate dilatation of several blood
vessels
Severe dilatation of several
blood vessels
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in the molar ratio of 4:5:14:6:1. The molecular weight of the
polysaccharide was 1.3 � 106 kD.
Yields and phenolic contents of each fraction are shown in
Table 2.
Macroscopic Evaluation of ColonResults of measuring colon weight-to-length ratio and ulcer area
are presented in Table 3. Data of negative control group show
that acetic acid instillation was an efficient method for inducing
ulcerative colitis as reported in previous studies. In the positive
control group, which received prednisolone 5 mg/kg/day for
5 days before induction of colitis, the macroscopic parameters
were not changed significantly, which may be due to the pre-
treatment. Prednisolone cannot prevent the damage made by
acetic acid in the form of pretreatment. Efficacy of treatment
by herbal fractions was higher in treatment group I, which
received aqueous fraction and decreased for the groups that
received ethanolic and n-hexane fractions. In the groups that
received the isolated polysaccharide as pretreatment, efficacy
in decreasing macroscopic damage was very similar to treatment
group I. However, the treatment plan of group V, which received
the isolated polysaccharide after induction of colitis, was not as
effective as the aqueous fraction.
A same trend was observed for all the macroscopic damage
parameters studied. Macroscopic appearance of colon speci-
mens after treatment in different groups is presented in Figure 1.
Microscopic Evaluation of Colon
Results of the histopathologic study confirmed the results of the
macroscopic evaluation. As presented in Table 4, considerable
inflammation and ulcer was observed in negative and positive
control groups, and prednisolone did not decrease the
Table 3. Results of Macroscopic Damage of Colitis in Different
Groups.
Macroscopic Parameters of Colitisa
Treatment
Groups
Colon Wet Weight-
to-Length Ratio
(mg/cm)
Ulcer Area
(mm2)
Damage
Score
Normal 0.642 + 0.026 No ulcer 0
Control 1.09 + 0.13 193.45 + 78.98 2.00 + 0.37
Prednisolone 0.90 + 0.08 371 + 65 1.83 + 0.40
Treatment I 0.65 + 0.02 No ulcer 1.00 + 0.17
Treatment II 0.71 + 0.03 No ulcer 0.80 + 0.20
Treatment III 0.72 + 0.04 126.33 + 41.59 1.50 + 0.34
Treatment IV 0.68 + 0.02 132.67 + 54.85 1.00 + 0.37
Treatment V 0.82 + 0.05 458.40 + 47.30 2.40 + 0.24
aResults are presented as mean + standard error of mean (N ¼ 6).
Table 2. Yield and Phenolic Content of Fraction Obtained From
Malva sylvestris.a
n-Hexane
Fraction
Ethanolic
Fraction
Aqueous
Fraction
Percentage of extraction
yield (w/w)
1.85 + 0.23 1.60 + 0.15 10.83 + 2.70
Phenolic content (mg/g of
dried fraction)
0.47 + 0.04 1.05 + 0.06 3.39 + 0.31
aData are shown as mean + standard deviation.
Figure 1. Macroscopic appearance of colon in different groups:
(a) Normal, (b) Control, (c) Prednisolone, (d) Treatment I, and
(e) Treatment IV.
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microscopic parameters of inflammation by pretreatment.
Between different groups treated with herbal fractions, the aqu-
eous fraction provided the highest efficacy in preventing signs
of inflammation and ulcer and the effect of n-hexane fraction
was the lowest.
Figure 2 shows the histological appearance of colon of rats
in different groups. Pretreatment with polysaccharides of the
plant also was effective in reducing the signs of inflammation,
but posttreatment with the isolated polysaccharide was not as
beneficial. Comparison between microscopic results of differ-
ent groups is summarized in Figure 3.
Discussion
In this study, anti-inflammatory effects of different fractions of
Malva sylvestris were evaluated against acetic acid–induced
ulcerative colitis in rats. Anti-inflammatory effects of the tested
fractions in this study seem to be related to their polysaccharide
and polyphenol content. Therefore, the content of polyphenols
in different obtained fractions was determined spectrophoto-
metrically and used as the method for standardization of the
fraction. Also, the structure of the isolated polysaccharide was
characterized. Polyphenols including flavonoids, anthocyanins,
and tannins are secondary plant metabolites with diverse biolo-
gical activity such as antioxidant, anti-inflammatory,12,16-19
anticarcinogenic, antimicrobial, vasodilatory, and wound heal-
ing properties.11,31,32 The highest phenolic content was found
in the aqueous fraction (3.39 mg/g), which was lower than pre-
vious reports on this plant.12,33 It may be due to different meth-
ods applied for total phenol determination and variation in
plant growth condition. The yield of polysaccharide extraction
in our study was higher than the optimized condition suggested
by Samavati.34 The characterized polysaccharide in this study
had an acidic nature with a high molecular weight. These data
are in accordance with previous reports on Malva sylvestris
polysaccharides.17,35,36
The protective effect of the aqueous fraction of Malva syl-
vestris on reducing microscopic and macroscopic parameters
of colitis was significantly higher than prednisolone, which is
a conventional medicine for the treatment of acute episodes
of ulcerative colitis. Indeed, ulcers were not observed in rats
treated with aqueous and ethanolic fractions of Malva sylves-
tris, and the damage scores in these 2 groups were significantly
lower than the others. The aqueous extract of different parts
(flowers, leaf, and roots) of this plant was reported to be rich
in mucilaginous polysaccharides with monosaccharide con-
tents of glucuronic acid, galacturonic acid, rhamnose, galac-
tose, fructose, glucose, sucrose, and trehalose.17,35-38 The
mucilaginous proteoglycans of this plant were reported to have
anticomplementary activity,17 which may be one of the prob-
able mechanisms for the observed results. A considerable
amount (12.8% dry weight) of arabinogalactans has been
extracted from cultures of Malva sylvestris callus cells.39
These polysaccharides are believed to be the main effective
components of the aqueous fraction of the plant. The anti-
inflammatory effect of extracted Malva sylvestris polysacchar-
ides was also studied as pre- and post-treatment. The results
showed that these polysaccharides have exhibited beneficial
effects against acid-induced ulcerative colitis when used as pre-
treatment. To our knowledge, there is no report on effects of
Malva sylvestris polysaccharides to compare the findings but
polysaccharides of Rheum tanguticum, Angelica sinensis, and
Vaccinium oxycoccos have been studied against this disease.
Liu et al reported that treatment with Rheum tanguticum poly-
saccharides at dose 200 mg/kg significantly reduced mortality,
colon mass, and ulcer areas.40 Wong et al suggested that the
protective effects of Angelica sinensis polysaccharides (in a
pretreatment protocol) may be because of the prevention of
oxidative stress in the pathological process of ulcerative coli-
tis.41 Beneficial effects of pectic polysaccharides from the
common cranberry (Vaccinium oxycoccos) on acetic acid–
induced colitis with the possible mechanism of reduction of
neutrophil infiltration and polysaccharide antioxidant proper-
ties was also reported by Popov et al.42 Polysaccharides of
Malva sylvestris were reported to have antioxidant activity,34
and this effect may be attributed to the observed activity in the
present study.34 Although we did not evaluate the mechanism
of the observed results, our findings in macroscopic and micro-
scopic studies are in agreement with the results of above-
mentioned studies about other polysaccharides. This could be
Table 4. Results of Microscopic Damage of Colitis in Different Groups.
Microscopic Parameters of Colitisa
Treatment
Groups Ulceration
Mucous Cell
Depletion
Crypt
Abscess
Inflammatory
Cyst
Mucosal
Atrophy
Edema Sub
mucosal
Inflammatory
Cell
Vascular
Dilatation
Normal 0.3 (0) 0.2 (0) 0.0 (0) 0.0 (0) 0.2 (0) 0.5 (0) 1.2 (0) 0.5 (0)
Control 2.0 (3) 1.7 (3) 1.3 (0) 0.8 (0) 1.3 (2) 2.3 (3) 2.3 (3) 2.2 (3)
Prednisolone 2.7 (3) 2.8 (3) 2.3 (3) 1.8 (3) 2.8 (3) 3.0 (3) 2.8 (3)2.8 (3)
Treatment I 0.3 (0) 0.2 (0) 0.0 (0) 0.0 (0) 0.0 (0) 0.3 (0) 1.8 (2) 0.8 (1)
Treatment II 1.7 (2) 1.3 (2) 1.0 (0) 0.5 (0) 1.2 (2) 2.2 (3) 2.7 (3) 2.3 (3)
Treatment III 2.5 (3) 1.8 (2) 1.5 (0) 1.0 (0) 1.7 (2) 2.5 (3) 3.0 (3) 2.7 (3)
Treatment IV 0.6 (0) 1.0 (0) 0.6 (0) 0.6 (0) 1.2 (0) 1.6 (2) 2.4 (3) 2.0 (3)
Treatment V 1.3 (2) 1.8 (2) 0.8 (0) 0.5 (0) 1.5 (1) 1.8 (2) 2.3 (3) 2.0 (2)
aResults are mean of 6 rats. The value in the parentheses is the mode of data.
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related to immunomodulatory,43 anti-inflammatory,44,45 and wound
healing,46 as well as antioxidant properties of polysaccharides.
The aqueous fraction of Malva sylvestris showed higher
protective effects against ulcerative colitis than the isolated
polysaccharide, which suggests that other constituents in the
aqueous fraction provided synergistic effects with the polysac-
charide. Magro et al reported that the aqueous extract of Malva
sylvestris leaves was the only preparation of this plant that had
significant antimicrobial properties against fungi,47 and also it
was reported to inhibit the prostaglandin synthesis by cycloox-
ygenase pathway.
The aqueous extract of fresh leaves of Malva sylvestris was
previously reported to contain different terpenoids including
linalool, linalool-1-oic acid, different megastigmene deriva-
tives, blumenol A, dehydrovomifoliol, and malvone A.23,48
Also, fructose, glucose, trehalose, ascorbic acid, and some gly-
cosidic phenolics were identified in the aqueous extract of this
plant.12,49 Geerling et al reported that intake of fructose, mag-
nesium, vitamin C, and fruit was associated with reduced
ulcerative colitis risk.50 On the other hand, some of these
components were reported for exhibition of antioxidant, anti-
inflammatory, and antioxidant properties that may have a role
Figure 2. Histopathologic appearance of colon in different groups: (a) Normal, (b) Control, (c) Prednisolone, (d) Treatment I, and (e) Treat-
ment IV.
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in the observed efficacy of this fraction. As far as we know,
this is the first report on the beneficial effects of Malva syl-
vestris in the treatment or prevention of ulcerative colitis
but the anti-inflammatory and anti-ulcerogenic properties
of the aqueous extract of Malva sylvestris against gastric
ulcers were previously reported in oral administration by
Sleiman and Daher.19 They showed that the aqueous extract
of Malva sylvestris enhanced the mucous production in
mucosal tissue and improved elastin promoter activity,51
enhancing the elasticity of mucosal tissues by inhibition of
elastase and trypsin.52
The ethanolic fraction and to a lower extent the hexane frac-
tion showed a reduction in evaluated macroscopic and micro-
scopic parameters by pretreatment. The hexane extract of
Malva sylvestris was previously reported to contain carote-
noids, tocopherols, and different fatty acids with the consider-
able portion (50% to 80%) of polyunsaturated fatty acid.
Although carotenoids and tocoferols have the potential of anti-
oxidant properties and may be responsible for the efficacy of
these fractions, Geerling et al reported that high intake of
mono- and polyunsaturated fatty acids may increase the risk
of developing ulcerative colitis and this may explain the lower
beneficial results (compared to aqueous and ethanolic frac-
tions) observed in present study.50
The aqueous fraction of Malva sylvestris provided the
highest extraction yield and the highest protective effects
compared with the other fractions and isolated polysacchar-
ide. Moreover, this plant has been known as an edible wild
vegetable in many countries. The study by Sleiman and
Daher19 on the safety of oral administration of the plant
showed that the oral intake of Malva sylvestris was not asso-
ciated with serious side effects.
Conclusion
Overall, according to the result of this study, it can be con-
cluded that fractions of Malva sylvestris have decreased the
inflammatory symptoms of experimental colitis induced by
acetic acid in rats. Further pharmacognostic and pharmacolo-
gical studies on these fractions can result in finding new
active constituents for treatment of inflammatory bowel dis-
ease. Considering that aqueous fraction of Malva sylvestris
presented the highest extraction yield and protective effects
compared to other fractions and the isolated polysaccharide,
preparations of the aqueous fractions of this plant can be sug-
gested as a safe remedy for patients with inflammatory bowel
disease. Also, this vegetable can be suggested to be used as a
functional food or a protective supplement for patients with
ulcerative colitis to improve their health condition or to
reduce the risks of adverse effects of their conventional med-
ication; however, more in vivo and clinical studies are needed
to support this hypothesis.
Author Contributions
Azadeh Hamedi (supervising pharmacognosy part and writing part of
the article), Hossein Rezaei (conducting the animal study and prepar-
ing herbal extracts), Negar Azarpira (conducting the histopathological
study), Mehrnaz Jafarpour (conducting mass spectroscopy and other
experimental analysis and writing part of the article), and Fatemeh
Ahmadi (supervising animal study and writing and finalizing the
article).
Declaration of Conflicting Interests
The authors declared no potential conflicts of interest with respect to
the research, authorship, and/or publication of this article.
Figure 3. Comparison of results of microscopic evaluation of colitis in different groups.
*P < 0.05 for all parameters. **P < 0.01 for all parameters.
Hamedi et al 7
 at UNIVERSITE DE MONTREAL on September 2, 2015chp.sagepub.comDownloaded from 
http://chp.sagepub.com/
Funding
The authors disclosed receipt of the following financial support for the
research, authorship, and/or publication of this article: This study was
part of the PharmD thesis project of Hossein Rezaei and was financed
by Shiraz University of Medical Sciences (Grant No. 90-01-36-3994).
Ethical Approval
The experiments were conducted in accordance with ethical guide-
lines for animal experimentation approved by ethical committee of
Animal Laboratories in Shiraz University of Medical Sciences.
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