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Antioxidant and Anti-Inflammatory Effects of Gel and Aqueous Extract of Melilotus officinalis L. in Induced Ulcerative Colitis: A Rattus norvegicus Model

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Background: Ulcerative colitis (UC) is a chronic condition of intestine illness accompanied by some unknown etiology with different immune, genetic and environmental factors. Objectives: The current study aimed to evaluate the antioxidant and anti-inflammatory effects of the Melilotus officinalis L. in the acetic acid induced UC in rats. Materials and Methods: The plant aqueous extraction and high performance liquid chromatography and Ferric Reducing Antioxidant Power (FRAP) assay were performed on aqueous extract to identify its compounds and antioxidant activities. Also, 70 adult male rats were and UC was induced using 3% acetic acid solution. They received different daily doses of M. officinalis L. in two forms (orally, 500 and 1000 mg/kg) and gel extract (10% and 20%). On the 7th day, the colon tissues were examined regarding the macroscopic and histopathology lesions plus oxidative stress and compared to the positive and negative control groups. Results: HPLC analysis revealed that five grams of the flower powder contained 9.7 mg gallic acid, 99 mg catechin, 21.9 mg caffeic acid, 0.86 mg chlorogenic acid, 1.13 mg quercetin, 548.9 mg cinnamic acid, 289 mg coumarin and 126 mg p-coumaric acid. The FRAP value of the extract was 2.91 ± 0.14 μM/g. There were significant differences between the group of rats which received the gel or aqueous extract of the flower compared to the negative control group using normal saline and the base gel and they had no significant differences with the positive control group using the Asacol, regarding the pathologic, malondialdehyde, and weight improvements. Conclusions: It can be concluded that the M. officinalis L. extract can be used as an effective medicine to treat UC in animal model and also in human subjects. Keywords: Colitis, Ulcerative; Inflammation; Extract; Ulcerative Colitis; Asacol
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Ann Colorectal Res. 2015 June; 3(2): e29511. DOI: 10.17795/colorectalresearch29511
Published online 2015 June 1. Research Article
Antioxidant and Anti-Inflammatory Effects of Gel and Aqueous Extract of
Melilotus officinalis L. in Induced Ulcerative Colitis: A Rattus norvegicus Model
Ali Reza Safarpour 1; Fatemeh Kaviyani 2; Masood Sepehrimanesh 1,*; Nasrollah Ahmadi 3;
Omid Koohi Hosseinabadi 4; Nader Tanideh 2; Najmeh Showraki 5
1Gastroenterohepatology Research Center, Shiraz University of Medical Sciences, Shiraz, IR Iran
2Transgenic Technology Research Center, Shiraz University of Medical Sciences, Shiraz, IR Iran
3Department of Pathology, School of Veterinary Medicine, Shiraz University, Shiraz, IR Iran
4Laboratory Animals Center, Shiraz University of Medical Sciences, Shiraz, IR Iran
5Department of Oral Medicine, School of Dentistry, Shiraz University of Medical Sciences, Shiraz, IR Iran
*Corresponding author: Masood Sepehrimanesh, Gastroenterohepatology Research Center, Research Tower, Nemazee Teaching Hospital, Shiraz University of Medical Sciences, P. O.
Box: 7193-51311, Shiraz, IR Iran. Tel: +98-7136281442, E-mail: sepehrimaneshmasood@gmail.com
Received: May 3, 2015; Revised: June 9, 2015; Accepted: June 12, 2015
Background: Ulcerative colitis (UC) is a chronic condition of intestine illness accompanied by some unknown etiology with different
immune, genetic and environmental factors.
Objectives: The current study aimed to evaluate the antioxidant and anti-inflammatory effects of the Melilotus officinalis L. in the acetic
acid induced UC in rats.
Materials and Methods: The plant aqueous extraction and high performance liquid chromatography and Ferric Reducing Antioxidant
Power (FRAP) assay were performed on aqueous extract to identify its compounds and antioxidant activities. Also, 70 adult male rats were
and UC was induced using 3% acetic acid solution. They received different daily doses of M. officinalis L. in two forms (orally, 500 and 1000
mg/kg) and gel extract (10% and 20%). On the 7th day, the colon tissues were examined regarding the macroscopic and histopathology
lesions plus oxidative stress and compared to the positive and negative control groups.
Results: HPLC analysis revealed that five grams of the flower powder contained 9.7 mg gallic acid, 99 mg catechin, 21.9 mg caffeic acid,
0.86 mg chlorogenic acid, 1.13 mg quercetin, 548.9 mg cinnamic acid, 289 mg coumarin and 126 mg p-coumaric acid. The FRAP value of
the extract was 2.91 ± 0.14 μM/g. There were signicant dierences between the group of rats which received the gel or aqueous extract of
the flower compared to the negative control group using normal saline and the base gel and they had no significant differences with the
positive control group using the Asacol, regarding the pathologic, malondialdehyde, and weight improvements.
Conclusions: It can be concluded that the M. officinalis L. extract can be used as an effective medicine to treat UC in animal model and also
in human subjects.
Keywords: Colitis, Ulcerative; Inflammation; Extract; Ulcerative Colitis; Asacol
Copyright © 2015, Colorectal Research Center and Health Policy Research Center of Shiraz University of Medical Sciences. This is an open-access article distributed
under the terms of the Creative Commons Attribution-NonCommercial 4.0 International License (http://creativecommons.org/licenses/by-nc/4.0/) which permits
copy and redistribute the material just in noncommercial usages, provided the original work is properly cited.
1. Background
Inflammatory bowel diseases (IBD), including Crohn’s
disease (CD) and ulcerative colitis (UC) are chronic in-
flammatory disease of the gastrointestinal tract. In fact,
UC is a relapsing non-transmural inflammatory disease
restricted to the colon and is one of the idiopathic dis-
eases of IBD (1). The incidence of UC remained stable
over the past 30 years, but, the disease become more
prevalent due to the early age of onset and low mortal-
ity, albeit with significant morbidity (2-4). The etiolo-
gies of the IBD are unknown and therefore there are no
causal treatments. In addition, patients with UC have
substantial impairments in health-related quality of
life (QoL) due to their symptoms, particularly during
periods of disease activity (5). On the other hand, the
current treatment is not completely successful and it is
frequently connected with adverse effects (6). It is sug-
gested that the plant and its products may provide an
option for the alternative or supplementary treatment
of the patients with IBD (7).
There are several ways to induce UC in the animal
model; they include using trinitrobenzene sulfonic acid
(TNBS) (8), dextran sulfate sodium (9), and acetic acid
(10, 11). Induction of UC using acetic acid had no mortal-
ity and the general health of the rats was similar to that
of the controls (12). Therefore, this method is widely sug-
gested as a useful model for the studies on the patho-
physiology and treatment of UC and it was performed
in the current study.
A number of traditional plants were reported in the
literature, but there was little scientific effort to validate
their effects. One of them is the genus of Melilotus, fam-
ily of Fabaceae. Data regarding traditional phytotherapy
Safarpour A et al.
Ann Colorectal Res. 2015;3(2):e295112
show that the Melilotus species (sweet clover) were used
to reduce spasm, in the liver diseases and as a diuretic
(13). Melilotus officinalis L. also named yellow sweet clover
is a plant not only used as food and forage but also as a
medicine. The preventive effect of the whole plant ex-
tract on the experimental atherosclerosis in rabbits was
reported (14). However, the number of reports on the M.
officinalis L. application in animal and human diseases is
scarce and there are no reports on the efficacy of M. offici-
nalis L. in UC treatment.
2. Objectives
The current study aimed to determine the healing ef-
fects of M. officinalis L. as dietary and gel form and com-
pare them with that of Asacol on the tissue histopatho-
logical changes and malondialdehyde (MDA) level in
male rats with experimentally acetic acid induced UC.
3. Materials and Methods
3.1. Ethical Statement
This study was approved by the animal care and use
committee of Shiraz university of medical sciences, Shi-
raz, Iran. All efforts were made to prevent any unneces-
sary and harmful animal handling.
3.2. Plant Materials and Aqueous Extraction
Melilotusofficinalis L. was collected from the Productive
Farm of Baradan Medicinal Plant Co., Abadeh, Iran (GPS
coordinates: 31.160833, 52.650556, with Altitude of 2500
m) during the spring season (May, 2013). All plants were
in the flowering stage of developing and the taxonomic
identification of each plant was confirmed by the Fars
agriculture Jihad educational complex, Fars, Iran and the
local herbarium number 14684. The plant also matched
with the digital herbarium of Botanical Garden and Bo-
tanical Museum Berlin-Dahlem, Freie university, Berlin
(http://ww2.bgbm.org/herbarium/ (Barcode: B -W 14163
-01 0 / ImageId: 385175).
3.3. Aqueous Extraction
The provided plants were dried for five days at room
temperature and then powdered by grinder. Finally, 100
g of plant powder was used for aqueous extraction based
on previous reported procedures (15).
3.4. High Performance Liquid Chromatography
(HPLC)
Phenolic acid constituents of M. officinalis L. were
identified by high performance liquid chromatogra-
phy (HPLC) according to the method reported by Kwok
et al. (16), with some modifications. Briefly, 5 g of plant
powder was dissolved in 50 mL of HPLC grad water and
after 24 hours, the solution was ltered across 0.22 μm
filter. HPLC analyses were performed using an Agilent
1200 HPLC system (Milford, MA, USA) coupled to a pho-
todiode array detector. The samples were separated on
a reversed-phase C18 column (150 mm × 4.6 mm). The
mobile phase consisted of water (solvent A) and formic
acid (solvent B). At time = 0, the solvent A and B were in
1:9 (v/v) ratio. The gradient mobile phase was gradually
changed to 89:11 (v/v) A to B ratio in 20 minutes. Mobile
phase rate was 1 mL/minute and the column tempera-
ture was adjusted at 30°C. The eluent was monitored
by a UV detector and the detection wavelength was set
at 280 and 360 nm. Galic acid, chlorogenic acid, caffeic
acid, quercetin, cumarin, p-cumaric acid, and carvacrol
were used as internal standards.
3.5. Ferric Reducing Antioxidant Power Assay
FRAP assay was performed on aqueous extract accord-
ing to the previous methods (17, 18) with some modifica-
tions. A 20 μL sample of extract with dierent concen-
trations, 31.25, 62.5, 125, 250, 500 mM and 1 M was mixed
with 180 μL of FRAP reagent in the pale wells. Blank
samples also were prepared; then, both real and blank
samples were incubated in water bath for 10 minutes
at 37°C and the absorbance of the samples was deter-
mined against blank ones at 593 nm. Series of stock so-
lution at 12.5, 25, 50, 100 and 200 μg/mL were prepared
(r2 = 0.999) using aqueous solution of FeSO4.7H2O as the
standard curve. The values obtained were expressed as
nanomole (μM) of ferrous equivalent Fe (II) per gram of
freeze dried sample.
3.6. Animal Housing, UC Induction and Treatment
Seventy 250 - 280 g male Sprague Dawley rats were pur-
chased from the Laboratory Animals Center, Shiraz Uni-
versity of Medical Sciences. Animals were randomly allo-
cated into seven equal independent groups and treated
according to the procedure shown in Figure 1.
UC was induced according to the previously reported
protocols (10, 11). Briefly, all animals were fasted over-
night and their bowels were cleaned before induction
of colitis. A polyurethane cannula (2 mm diameter) was
applied for the rectal entrance of acetic acid and the
tip was inserted up to 8 cm proximal to the anus verge.
Two milliliters of 3% acetic acid was administered tran-
srectally into the colon by a cannula during 30 seconds
to induce UC under ketamine and xylazine anesthesia.
The main substance of gel was carboxymethyl cellulose
(CMC) and the gel was produced by mixing 2% sodium
CMC in 5% glycerol and it was continuously stirred with
a mixer in order to prepare a gel-forming agent. In the
next step, M. officinalis L. extracts (10%, and 20%) were
added to deionized water. The mixtures were gradually
added to the Na-CMC with glycerol and finally the pre-
pared gel was homogenized for 30 minutes and all for-
mulations were collected in an aluminum tube in the
refrigerator (11).
Safarpour A et al.
3
Ann Colorectal Res. 2015;3(2):e29511
70 male SD rat
10 male SD rat
10 male SD rat
M. oficinalis
Gel 10%
Enema
2 ml per day
M. oficinalis
Gel 20%
Enema
2 ml per day
M. oficinalis
Aqueous extract
(500 mg/kg)
Oral
1 ml per day
M. oficinalis
Aqueous extract
(1000 mg/kg)
Oral
1 ml per day
10 male SD rat 10 male SD rat 10 male SD rat
10 male SD rat 10 male SD rat
Grouping and UC induction
Group I
Group IV Group VGroup VI Group VII
Group II Group III
Normal saline
Oral
1 ml per day
Asacol
Enema
2 ml per day
Gel base
Enema
2 ml per day
• Daily weigh measurement
• Histopathological evaluations
• MDA assay
Colon tissue sampling After 7 days
Figure 1. Experimental Setup and Rat Groupings
3.7. Tissue Sampling, Macroscopic and Microscop-
ic Evaluations
After seven days, rats were euthanized under deep
ether anesthesia. Then, the distal 8 cm of the colon was
removed and opened by longitudinal incision. The mu-
cosal surface was washed with saline buffer and the mu-
cosal injury was macroscopically assessed using Morris
et al. (19) grading scale. Colon tissue was processed and
stained according to the previously reported procedures
(10, 20-25). Briefly, colon tissues were free of surrounding
fat and fixed in 10% buffered formalin solution for histo-
pathological examination. Formalin-fixed tissues were
processed routinely and embedded in paraffin. Blocks
were cut at 5 μm and stained with hematoxylin-eosin. All
sections were studied and photographed using a light
microscope. The degree of inflammation of the colon was
graded as previously described (26).
3.8. Malondialdehyde Measurement
Colon tissue MDA assessment was performed by mea-
suring thiobarbituric acid reactive substances (TBARS)
in PBS tissue homogenate (27); since MDA is one of the
end-products of lipid peroxidation (LPO), and the extent
of LPO is most frequently measured by estimating MDA
levels (28).
3.9. Statistical Analysis
The results were presented as mean. Differences among
the groups were determined using a one-way ANOVA and
Duncan post-hoc test. All the statistical analyses were per-
formed by SPSS software (version 18, Chicago, IL, USA). P
value less than 0.05 was considered significant.
4. Results
The current study aqueous extraction efficiency was
Safarpour A et al.
Ann Colorectal Res. 2015;3(2):e295114
39%. Chromatogram of M. officinalis L. in 280 nm is shown
in Figure 2. As demonstrated, the comarin had the high-
est concentration in the M. officinalis L. phenolic acid con-
stituents; and comarin levels in aqueous extract of M. offi-
cinalis L. are presented in Table 1. Catechin and cinnamic
acid were the highest compounds in the phenolic acid
and comarin constituents of M. officinalis L. Antioxidant
activity of aqueous extract of M. officinalis L. was mea-
sured based on the FRAP assay.
The current study analysis demonstrated that the FRAP
value for this extract was 2.91 ± 0.14 μM/g. Percentage of
weight changes in different groups during the experi-
mental period is shown in Figure 3. The most weight
change was observed in the normal saline control group
(13.67%), while, the lowest weight changes were observed
in the Asacol control, 20% extract enema, and 1000 mg
oral extract with 1.44%, 1.62%, and 2.31%, respectively.
Mean comparison of macroscopic and microscopic
changes as quantitative values plus MDA level in differ-
ent groups are presented in Table 2. As observed, normal
saline control group had the highest and Asacol control,
20% extract enema, and 1000 mg oral extract showed the
lowest pathological changes in both macroscopic and
microscopic evaluations. Also, changes of MDA levels in
response to different treatments indicated the antioxi-
dant activity of M. officinalis L. extract. The lowest MDA
level (highest antioxidant level) was detected in the gel
20% in enema rout of administration, which was compa-
rable to that of Asacol treatment (Table 2).
mAU
800
700
600
500
400
300
200
100
0
0510 15 20 25 30 min
10.864
12.124
16.640
17.048
17.919
15.450
17.49
3
Figure 2. Chromatogram of Aqueous Melilotus officinalis Extract Provided
by High Performance Liquid Chromatography
Table 1. Characterization of Melilotus officinalis Compounds by
High Performance Liquid Chromatography
Compounds Level (mg/L)
Phenolic acid compounds
Gallic acid 9.75
Catechin 99.06
Caffeic acid 21.99
Chloregenic acid 0.87
Quercetin 1.13
Comarin compounds
Cinnamic acid 548.91
Comarin 289.00
p-comaric acid 126.00
Rutin -
Carvacerol -
Figure 3. Rate of Weight Change and its Trend in Different Groups Dur-
ing the Study Period
16
14
12
8
6
4
2
0
Percentage of weight change
Control (Normal saline)
Control (Base gel)
Control (Asacoi)
OE 500
OE 1000
EE 10%
EE 20%
Group
a
13.67 b
12.16
d
5.69 d
4.81
c
2.31
c
1.44
c
1.62
Differences among the groups were determined using a one-way ANOVA
and Duncan post-hoc test. Significant difference between the groups is
indicated by different superscript letters (P < 0.05).
Table 2. Comparison Between Macroscopic and Microscopic Scores Plus Malondialdehyde Levels in Different Experimental Groupsa, b
Groups Macroscopic Score Microscopic Score b MDA Level (μM)
Control groups
Normal saline 19.0 10.7 1.80
Base gel 18.1 9.9 1.75
Asacol 1.9 1.2 0.93
Melilotus officinalis
500 mg/kg oral extract 8.2 4.7 1.13
1000 mg/kg oral extract 2.1 1.0 1.00
10% gel enema 7.5 4.2 1.13
20% gel enema 1.8 0.8 0.93
a Differences among the groups were determined using a one-way ANOVA and Duncan post-hoc test.
b Significance: P < 0.05.
Safarpour A et al.
5
Ann Colorectal Res. 2015;3(2):e29511
Comparison of histopathological lesions in the colon
tissue of healthy rats and seven different experimental
groups are shown in Figure 4. As indicated, acetic acid in-
duced UC in the colon, demonstrated by lack of epithelial
cells plus infiltration of inflammatory cells in the sub-
mucosal layers. However, by administration of oral and
enema of M. officinalis L., the healing process of the colon
tissue was improved and using 1000 mg/kg oral aqueous
extract or gel 20% enema can completely cure the colon
tissue, comparable to Asacol treatment.
Figure 4. Histopathological Evaluations of the Colon Tissues
A, healthy rat colon with complete crypt, submucosal layer and muscular layer plus healthy epithelium; B, normal saline control group, lack of epithelial
cells plus inflammatory cells infiltration; C, base gel control group, damage of epithelium and inflammation of subepithelial layers; D,500 mg/kg oral
aqueous extract of Melilotus officinalis, formation of epithelial cells but existence of inflammatory cells in the submucosal layers; E, enema of Melilotus
officinalis L. 10% gel, formation of epithelial cells but existence of inflammator y cells in the submucosal layers; F, enema of M. officinalis L. 20% gel, approxi-
mately complete formation of epithelial cells plus crypts and muscularis mucosa formation, mild inflammation of the submucosal layers; G, 1000 mg/kg
oral aqueous extract of Melilotus officinalis L., complete formation of epithelial cells, crypts and muscularis mucosa, very mild inflammation of the sub-
mucosal layers; H, Asacol control group, complete formation of epithelial cells, crypts and muscularis mucosa without any inflammation (H& E staining).
Safarpour A et al.
Ann Colorectal Res. 2015;3(2):e295116
5. Discussion
The current study compared the healing effects of M. of-
ficinalis L. extract in the 500 and 1000 mg/kg orally and
10% and 20% gel forms against acetic acid induced UC by
measuring tissue histopathology and MDA level in rats.
Also, the phenolic acid compounds of the plant and an-
tioxidant activity of aqueous extract were determined
using HPLC and FRAP test. The analysis revealed that cat-
echin and cinnamic acid were the highest phenolic acid
and comarin constituents of this plant. Also findings of
the current study demonstrated that both dietary and gel
forms had significant obvious healing effects and these
effects were dose dependent. However, the antioxidant
activity of the 20% gel in enema rout was clearly higher
than those of the other concentrations and routs of ad-
ministration.
Although, the UC symptoms can sometimes reduce
their own, the disease usually requires treatment to go
into remission. There are several strategies to treat UC
including pharmacotherapy (aminosalicylates, glucocor-
ticoids, immunosuppressive) and surgical procedures,
but the current therapies are not completely successful
and frequently cause adverse effects (7). Anti-inflamma-
tory drugs, such as aminosalicylates, corticosteroids and
immunosuppressive agents, were frequently applied to
treat this disease. However, there were serious side effects
and the recrudescence rates of IBD were rather high (29,
30). Therefore, scientists noticed the use of medicinal
plants as alternative or complementary therapies for UC.
The current study authors previously reported the bene-
ficial effects of some medicinal plants in different animal
species. The plants included Pistacia atlantica (11), Berberis
vulgaris (31), strawberry (32) and Hypericum perforatum
(10) in rats and Teucrium polium (33) and Calendula offici-
nalis (34) in dogs. However, no reports were issued on the
healing effects and antioxidant properties of M. officinalis
L. extract and its mechanism on UC until now.
Authors found that the aqueous extract of M. officinalis
L. contained high level of catechin and cinnamic acid as
the phenolic acid and comarin compounds. Catechin is a
type of natural phenol and antioxidant found in medici-
nal plants Catechins is reported to have anti-inflammatory
effects on TNBS and acetic acid induced rat colitis (35, 36).
This may be through selective immunomodulatory effects
mediated, at least in part, by inhibition of nuclear factor-
κB (NF-κB) and activator protein-1 (37). It is clarified that cat-
echins can exert their antioxidant activities via decreasing
nitric oxide, MDA, and increasing superoxide dismutase.
They also ameliorate mucosal inflammation by inhibit-
ing the production of TNF-α, IFN-γ and NF-κBp65 (36). The
current study findings regarding beneficial anti-colitis
activities of M. officinalis L. are in line with the above men-
tioned reports. It can be concluded that catechin, with the
highest amount of phenolic acid compound in the plant
extract, plays antioxidant and anti-inflammatory activities
due to the reported mechanisms.
Finally, MDA is frequently used to measure lipid perox-
ide levels and, exhibiting good correlation with degree
of lipid peroxidation. In the present study, the MDA lev-
els decreased in all M. officinalis L. treated groups com-
pared with those of the control groups. This decrease
may in fact support the protective effect of M. officinalis
L. against lipid peroxidation in UC. Although, scientific
information concerning antioxidant properties of the M.
officinalis L. extract is still rather scarce (13), such effects
against lipid peroxidation were previously reported in
cell culture experiments (38).
In conclusion, the mucus layer contributes significantly
to gut barrier function and protection from pathogens
(39) and the current findings are interesting as a strategy
to select the medicinal plants to treat UC. The antioxidant
and anti-inflammatory effects of M. officinalis L. extract
seem to be clear, especially by identifying catechin and
cinnamic acid as the most phenolic acid and comarin
constituents, but its molecular anti-inflammatory mech-
anisms are not fully investigated yet. Therefore, perform-
ing further studies to identify this mechanism in cell
culture and also animal and human studies are highly
recommended.
Acknowledgements
The animals were kindly provided by Mahjoob Vahedi
at the Laboratory Animals Center, Shiraz University of
Medical Sciences, Shiraz, Iran. Authors wish to thank all
people who kindly helped with the technical and scien-
tific phases of this study.
Authors’ Contributions
Ali Reza Safarpour: Study concept and design, analysis
and interpretation of data, critical revision of the manu-
script for important intellectual content; Fatemeh Kavi-
yani: Gathering, analysis and interpretation of data, Draft-
ing the manuscript; Masood Sepehrimanesh: Gathering,
analysis and interpretation of data, drafting the manu-
script, critical revision of the manuscript for important
intellectual content, statistical analysis, administrative,
technical, and material support, study supervision; Nas-
rollah Ahmadi: Data gathering, study supervision; Omid
Koohi Hosseinabadi: Gathering, analysis and interpre-
tation of data, drafting the manuscript; Nader Tanideh:
Study concept and design, drafting the manuscript, criti-
cal revision of the manuscript for important intellectual
content, study supervision.
Funding/Support
The study was supported by Shiraz University of Medi-
cal Sciences, Shiraz, IR Iran.
References
1. Baumgart DC, Sandborn WJ. Inflammatory bowel disease:
clinical aspects and established and evolving therapies. Lancet.
Safarpour A et al.
7
Ann Colorectal Res. 2015;3(2):e29511
2007;369(9573):1641–57.
2. Loftus CG, Loftus EJ, Harmsen WS, Zinsmeister AR, Tremaine
WJ, Melton L3, et al. Update on the incidence and prevalence of
Crohn's disease and ulcerative colitis in Olmsted County, Minne-
sota, 1940-2000. Inflamm Bowel Dis. 2007;13(3):254–61.
3. Safarpour AR, Hosseini SV, Mehrabani D. Epidemiology of inflam-
matory bowel diseases in iran and Asia; a mini review. Iran J Med
Sci. 2013;38(2 Suppl):140–9.
4. Taghavi SA, Safarpour AR, Hosseini SV, Noroozi H, Safarpour M,
Rahimikazerooni S. Epidemiology of Inflammatory Bowel Dis-
eases (IBD) in Iran: A Review of 740 Patients in Fars Province,
Southern Iran. Ann Colorectal Res. 2013;1(1):17–22.
5. Casellas F, Arenas JI, Baudet JS, Fabregas S, Garcia N, Gelabert
J, et al. Impairment of health-related quality of life in patients
with inflammatory bowel disease: a Spanish multicenter study.
Inflamm Bowel Dis. 2005;11(5):488–96.
6. Biasi F, Astegiano M, Maina M, Leonarduzzi G, Poli G. Poly-
phenol supplementation as a complementary medicinal ap-
proach to treating inflammatory bowel disease. Curr Med Chem.
2011;18(31):4851–65.
7. Vochyanova Z, Bartosova L, Bujdakova V, Fictum P, Husnik R,
Suchy P, et al. Diplacone and mimulone ameliorate dextran sul-
fate sodium-induced colitis in rats. Fitoterapia. 2015;101:201–7.
8. Mascaraque C, Gonzalez R, Suarez MD, Zarzuelo A, Sanchez de
Medina F, Martinez-Augustin O. Intestinal anti-inflammatory
activity of apigenin K in two rat colitis models induced by trini-
trobenzenesulfonic acid and dextran sulphate sodium. Br J Nutr.
2015;113(4):618–26.
9. Boussenna A, Cholet J, Goncalves-Mendes N, Joubert-Zakeyh J,
Fraisse D, Vasson MP, et al. Polyphenol-rich grape pomace ex-
tracts protect against dextran sulfate sodium-induced colitis in
rats. J Sci Food Agric. 2015.
10. Tanideh N, Nematollahi SL, Hosseini SV, Hosseinzadeh M, Mehra-
bani D, Safarpour A, et al. The Healing Effect of Hypericum per-
foratum Extract on Acetic Acid-Induced Ulcerative Colitis in Rat.
Ann Colorectal Res. 2014;2(4):e25188.
11. Tanideh N, Masoumi S, Hosseinzadeh M, Safarpour AR, Erjaee H,
Koohi-Hosseinabadi O, et al. Healing effect of pistacia atlantica
fruit oil extract in acetic Acid-induced colitis in rats. Iran J Med
Sci. 2014;39(6):522–8.
12. Fabia R, Willen R, Ar'Rajab A, Andersson R, Ahren B, Beng-
mark S. Acetic acid-induced colitis in the rat: a reproducible
experimental model for acute ulcerative colitis. Eur Surg Res.
1992;24(4):211–25.
13. Plesca-Manea L, Parvu AE, Parvu M, Taamas M, Buia R, Puia M. Ef-
fects of Melilotus officinalis on acute inflammation. Phytother
Res. 2002;16(4):316–9.
14. Hirakawa T, Okawa M, Kinjo J, Nohara T. A new oleanene glucuro-
nide obtained from the aerial parts of Melilotus officinalis. Chem
Pharm Bull (Tokyo). 2000;48(2):286–7.
15. Kazemipour N, Nikbin M, Davarimanesh A, Sepehrimanesh M.
Antioxidant activity and mineral element contents of Calotropis
procera from Iran: a traditional medicinal plant in Middle East.
Comp Clin Pathol. 2014:1–4.
16. Kwok C, Wong CN, Yau MY, Yu PH, Au ALS, Poon CC, et al. Con-
sumption of dried fruit of Crataegus pinnatifida (hawthorn)
suppresses high-cholesterol diet-induced hypercholesterolemia
in rats. J Funct Foods. 2010;2(3):179–86.
17. Kazemipour N, Nikbin M, Maghsoudlou MT, Sepehrimanesh M.
Antioxidant properties of extracts of Scutellaria lute-caerulea In
vitro. Online J Vet Res. 2014;18(2):75–81.
18. Benzie IF, Strain JJ. Ferric reducing/antioxidant power assay:
direct measure of total antioxidant activity of biological fluids
and modified version for simultaneous measurement of total
antioxidant power and ascorbic acid concentration. Methods En-
zymol. 1999;299:15–27.
19. Morris GP, Beck PL, Herridge MS, Depew WT, Szewczuk MR, Wal-
lace JL. Hapten-induced model of chronic inflammation and
ulceration in the rat colon. Gastroenterology. 1989;96(3):795–803.
20. Tamadon A, Nikahval B, Sepehrimanesh M, Mansourian M, Naei-
ni AT, Nazifi S. Epididymis ligation: a minimally invasive tech-
nique for preparation of teaser rams. Vet Surg. 2010;39(1):121–7.
21. Sepehrimanesh M, Azarpira N, Saeb M, Nazifi S, Kazemipour N,
Koohi O. Pathological changes associated with experimental
900-MHz electromagnetic wave exposure in rats. Com Clin Pathol.
2014;23(5):1629–31.
22. Panahi M, Karimaghai N, Rahmanifar F, Tamadon A, Vahdati A,
Mehrabani D, et al. Stereological evaluation of testes in busulfan-
induced infertility of hamster. Comp Clin Pathol. 2014:1–6.
23. Koohi-Hosseinabadi O, Moini M, Safarpoor A, Derakhshanfar A,
Sepehrimanesh M. Effects of dietary Thymus vulgaris extract
alone or with atorvastatin on the liver, kidney, heart, and brain
histopathological features in diabetic and hyperlipidemic male
rats. Comp Clin Pathol. 2015:1–5.
24. Rajaian H, Nazifi S, Mansourian M, Poorbaghi SL, Sepehri-
manesh M, Ghezelbash A. Pathological changes associated with
experimental salinomycin toxicosis in calves. Online J Vet Res.
2008;12:15–22.
25. Tafti AK, Nazifi S, Rajaian H, Sepehrimanesh M, Poorbaghi
SL, Mohtarami S. Pathological changes associated with ex-
perimental salinomycin toxicosis in sheep. Com Clin Pathol.
2008;17(4):255–8.
26. Shamsa F, Ahmadiani A, Khosrokhavar R. Antihistaminic and an-
ticholinergic activity of barberry fruit (Berberis vulgaris) in the
guinea-pig ileum. J Ethnopharmacol. 1999;64(2):161–6.
27. Koohi-Hosseinabadi O, Andisheh-Tadbir A, Bahadori P, Sepehri-
manesh M, Mardani M, Tanideh N. Comparison of the therapeu-
tic effects of the dietary and topical forms of Zizyphus jujuba
extract on oral mucositis induced by 5-fluorouracil: a golden
hamster model. J Clin Exp Dent. 2015;7:e304–9.
28. Lata H, Ahuja GK, Narang AP, Walia L. Effect of immobilisation
stress on lipid peroxidation and lipid profile in rabbits. Indian J
Clin Biochem. 2004;19(2):1–4.
29. Abdallah DM, Ismael NR. Resveratrol abrogates adhesion mol-
ecules and protects against TNBS-induced ulcerative colitis in
rats. Can J Physiol Pharmacol. 2011;89(11):811–8.
30. Liu X, Wang J. Anti-inflammatory effects of iridoid glycosides
fraction of Folium syringae leaves on TNBS-induced colitis in
rats. J Ethnopharmacol. 2011;133(2):780–7.
31. Tanideh N, Afaridi E, Mehrabani D, Azarpira N, Hosseinzadeh
M, Amini M, et al. The Healing Effect of Berberis vulgaris in Ace-
tic Acid-Induced Ulcerative Colitis in Rat. Middle East J Sci Res.
2014;21(8):1288–94.
32. Tanideh N, Akbari Baseri F, Jamshidzadeh A, Ashraf MJ, Kuhi
O, Mehrabani D. The healing effect of strawberry extract on
acetic acid-induced ulcerative colitis in rat. World Appl Sci J.
2014;31(3):281–8.
33. Mehrabani D, Bahrami F, Hosseini SV, Ashraf MJ, Tanideh N,
Rezaianzadeh A, et al. The Healing Effect of Teucrium polium in
Acetic Acid-Induced Ulcerative Colitis in the Dog as an Animal
Model. Middle East J Dig Dis. 2012;4(1):40–7.
34. Mehrabani D, Ziaei M, Hosseini SV, Ghahramani L, Bananzadeh
AM, Ashraf MJ, et al. The effect of calendula officinalis in therapy
of acetic Acid induced ulcerative colitis in dog as an animal mod-
el. Iran Red Crescent Med J. 2011;13(12):884–90.
35. Sato K, Kanazawa A, Ota N, Nakamura T, Fujimoto K. Dietary sup-
plementation of catechins and alpha-tocopherol accelerates the
healing of trinitrobenzene sulfonic acid-induced ulcerative coli-
tis in rats. J Nutr Sci Vitaminol (Tokyo). 1998;44(6):769–78.
36. Ran ZH, Chen C, Xiao SD. Epigallocatechin-3-gallate amelio-
rates rats colitis induced by acetic acid. Biomed Pharmacother.
2008;62(3):189–96.
37. Abboud PA, Hake PW, Burroughs TJ, Odoms K, O'Connor M,
Mangeshkar P, et al. Therapeutic effect of epigallocatechin-3-gal-
late in a mouse model of colitis. Eur J Pharmacol. 2008;579(1-3):411–7.
38. Braga PC, Dal Sasso M, Lattuada N, Marabini L, Calo R, Antonacci
R, et al. Antioxidant activity of Melilotus officinalis extract inves-
tigated by means of the radical scavenging activity, the chemilu-
minescence of human neutrophil bursts and lipoperoxidation
assay. J Med Plant Res. 2013;7(7):358–65.
39. Yazdani A, Poorbaghi SL, Habibi H, Nazifi S, Rahmani Far F, Sepeh-
rimanesh M. Dietary Berberis vulgaris extract enhances intesti-
nal mucosa morphology in the broiler chicken (Gallus gallus).
Comp Clini Pathol. 2013;22(4):611-5.
... Naringenin (14) on improving UC. In addition, Safarpour et al. reported (38) some beneficial effects of M. officinalis in treating UC. In this study, we evaluated the healing effects of M. officinalis at the higher doses to investigate the probable side effects. ...
... This can be one of the important and underlying mechanisms of M. officinalis protection against UC. Similar to findings of this study, Safarpour and et al. revealed that treatment by M. officinal in colitis rats decreased the level of MDA and improved UC symptoms (38). ...
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Background: Ulcerative colitis (UC) is a form of inflammatory bowel disease (IBD) that is characterized by acute and chronic inflammation.The etiology and pathophysiology of IBD is unidentified, and there are many obstacles on the definite treatment ofthis disease. Recently, the useful effects of some herbal medicine on improving UC have been studied. Melillotus officinalis L. (M.officinalis) is an herb with antioxidant and anti-inflammatory effects used as food, forage and medicine.Objectives: This study evaluated the antioxidant effects of M. officinalis aqueous extracts in the acetic acid- induced ulcerative colitisin rats.Methods: Fifty rats were randomly divided into five equal groups. Group I (Control healthy group) received 1 mL/kg of normalsaline orally. Group II (control colitis group) received 1 mL/kg of normal saline orally. Group III (positive control) received 3 mg/kgprednisolone orally. Group IV received 1000 mg/kg M. officinalis aqueous extracts orally. Group V received 2000 mg/kg M. officinalisaqueous extracts orally. Ulcerative colitis was induced by intra-rectal acetic acid (3% v/v) administration. All treatments were done24 hours after induction of colitis and continued for seven days. On the eighth day, the rats were sacrificed and colonic biopsieswere taken for histopathological and biochemical studies. Data analysis was performed, using SPSS software and significance levelwas set at P≤0.05.Results: Treatment with M. officinalis aqueous extract could enhance colonic antioxidant capacity and decrease inflammation andacute colonic injury induced by acetic acid, which is dose-dependent. In addition, administrating the extract significantly (P≤0.05)reduced the colonic level of malondialdehyde and myeloperoxidase, and significantly (P ≤ 0.05) increased the level of reducedglutathione (P≤0.05). The extract had more effects at the dose of 2000 mg/kg than 1000 mg/kg dosage and prednisolone.Conclusions: This study revealed that M. officinalis had apparent curative effects on treating UC because of its antioxidant and antiinflammatoryactivities.
... There are several models for induction of UC in animals such as use of trinitrobenzene sulfonic acid (TNBS) (7), dextran sodium sulfate, 8 and acetic acid. 9 Rectal administration of acetic acid can mimic the conditions which occurred in human UC 10 and related UC is a reproducible laboratory animal model and is useful for screening of effectiveness of drugs. 11 Use of medicinal plants and their derivatives has an ancient basis. ...
... 21 Improving of pathological conditions of colon, increase of weight and decline in MDA level were seen in rat suffered from UC in response to strawberry extract in dose-response type. 22 Similar findings are reported by our group and other scientists for Berberis vulgaris, 23 Melilotus officinalis, 9 Hypericum perforatum, 24,25 Calendula officinalis 26 and Pistachia atlantica 27 in line with findings of this study. However, the reports about the beneficial effects of using of Z. jujuba in inflammatory diseases are scarce. ...
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Objective: To investigate the effects of hydroalcoholic extract of Ziziphus jujuba on the histopathological, tissue oxidative stress and inflammation plus to antioxidant pathways of colon tissue in rat with induced Ulcerative colitis. Materials and methods: Ulcerative colitis was induced in 80 rats those divided into 8 equal groups. Group 1 and 2 were negative controls receiving 1 mL/day of normal saline in enema and oral; group 3 and 4 as positive control 1 and 2 received 10 mg/kg of intra-colonic asacol and oral mesalazine; groups 5 and 6 received 20% and 40% of hydroalcoholic extract of Z. jujuba trans-rectally; group 7 and 8 received 1500 and 3000 mg/kg of hydroalcoholic extract of Z. jujuba orally, respectively. After 7 days, animals were evaluated for colon tissue histopathology, levels of malondialdehyde and IL-1β, and activities of superoxide dismutase, glutathione peroxidase and myeloperoxidase in colon tissue. Results: Hydroalcoholic extract of Z. jujuba in both forms of trans-rectal and oral administration especially in the higher doses could result into a more healing effect in damaged colonic tissue, more reduce glutathione peroxidase and IL-1β level. Also, these two doses (gel 40% and oral 3000 mg/kg) could more decrease the myeloperoxidase activity and stimulate superoxide dismutase and glutathione peroxidase activities. Also, gel 40% in transrectal administration was more potent than administration 3000 mg/kg in oral. Conclusion: The results of the present study indicated that Z. jujube may be considered as a treatment of choice for Ulcerative colitis especially in gel form and also in dose-dependent pattern.
... IL-6 is another predominant cytokine found in inflamed areas from UC patients, and the other predominant cytokines were related to a 1 profile. Different studies confirmed that induced UC in rats increases immunomodulator indexes confirming that oxidative stress could be correlated to disease [36,[64][65][66][67]. Under normal conditions, ROS is naturally neutralized by the endogenous antioxidant system, including GSH, SOD, CAT, and GPx. ...
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Background. Ulcerative colitis is a worldwide chronic gastrointestinal disease characterized by variable extensions of colon mucosal inflammation. The available drugs have an incomplete response with various side effects and socioeconomic impacts. Aloe barbadensis Miller (Aloe vera) is a well-known medicinal plant with diverse pharmacological and therapeutic activities. As a result, in the current study, Aloe vera was selected to evaluate its therapeutic effects on experimental colitis in rats. Methods. This study is intended to evaluate the possible beneficial effect of Aloe vera for the treatment of experimental colitis. Trinitrobenzenesulfonic acid (TNBS) was used to induce experimental colitis in 60 of 70 Wistar rats. The rats were grouped in 7 clusters including healthy control, negative, positive control (received sulfasalazine), and test groups treated with Aloe vera extracts via oral or rectal routes. Macroscopic and histologic factors as well as the biochemical parameters were evaluated on day 7. Results. In the present study, it was found that serum levels of tumor necrosis factor-α (75 vs. 44 pg./ml), interleukin-6 (41 vs. 21 pg/ml), and nitric oxide (24 vs. 6 μm/ml) in TNBS-induced untreated colitis treatment were significantly increased as compared to healthy control. Similar patterns were also observed in malondialdehyde (76.41 vs. 236.35 μg/mg) and myeloperoxidase (4.24 vs. 29.38 U/mg) in colonic tissue. Among different treatments, rectal administration of Aloe vera extract (400 mg/kg) exhibited the best result in which serum concentration of tumor necrosis factor-α (55 pg/ml), interleukin-6 (24 pg/ml), and nitric oxide (10 μm/ml) and the levels of malondialdehyde (102.67 μg/mg), as well as myeloperoxidase (12.29 U/mg) in colon tissue, were reduced as compared to the untreated group. Also, the body weight and colon weight/length ratios were more improved in the treated group with 400 mg/kg Aloe vera extract, rectally. Conclusion. Aloe vera extract exhibited a therapeutic effect in TNBS-induced colitis, and local, rectal administration of Aloe vera extract was more effective than oral administration. 1. Introduction Ulcerative colitis (UC) as an annoying chronic problem is one of the two major subtypes of inflammatory bowel diseases (IBDs) with different geographic prevalences and worldwide distribution [1, 2]. Although UC may present insidiously, its hallmark is subacute bloody diarrhea, accompanied by anemia and fatigue. It also may change to acute severe colitis, presenting temperature above 37.5°C, heart rate above 90/min, and hemoglobin concentration below 10.5 g/dL with more than 6 bloody stools daily [3, 4]. Its manifestation is due to continuous inflammation of the rectum with the variable extension but usually with decreasing severity up to the cecum [5]. Its etiology and exact underlying pathophysiologic aspects are unclear, but most probably is due to aberrant deregulated mucosal immune responses (humoral and cellular immunity) to environmental factors in a genetically susceptible population. Following epithelial barrier dysfunction and immune cell activation, inflammatory cytokines and mediators (interferon-γ (IFN-γ), interleukin-2 (IL-2), IL-4, IL-5, IL-10, IL-13, IL-17, IL-23, and tumor necrosis factor α (TNF-α)) are released which may be used as disease activity indicators [6, 7]. The diagnosis of UC is based on clinical presentation and chronic colon inflammation confirmed by histology [8]. Uncertain definitive pathogenesis, variable presentation, natural course, and lack of standard disease activity index are obstacles for definite therapeutic effect assessment. In any case, the accepted therapeutic goals are (a) accentuating induction of remission and maintenance period, (b) improving the nutritional status, (c) decreasing disease complications, and (d) considering side effects and cost effectiveness. In current medicine, the main treatments are focused on 5-ASA and steroids. Biologic therapies such as antitumor necrosis factor antibodies are prescribed for resistant patients. Antiadhesion molecules and kinase inhibitors are under research for UC treatment [9–12]. The inadequate response, frequent relapse, steroid dependency, and side effects result in developing a new candidate as the second line of treatment. Considering therapeutic effects of some herbal medicine such as heartleaf houttuynia [13, 14], boswellic acid [15, 16], diamonnium glycyrrizhinate [17], slippery elm [18], fenugreek [18], devil’s claw [18], tormentil [18], and wei tong ning [18] in various diseases, especially in China, Middle-East, and Africa, new research in this field is rational. Aloe vera (Aloe barbadensis Mill.) belongs to the Aloeaceae family with thick, tapered, green lance-shaped, juicy, sharp, and edged leaves [19]. Aloe vera grows in dry regions of Africa, Europe, Asia, and America. Aloe vera is probably the most applied medicinal plant commercially and the most popular plant worldwide [20]. Various parts of the plant contain amino acids, sugars, enzymes, vitamins, minerals, saponins, anthraquinones, lignin, and salicylic acid. Also, the leaves are the source of various organic acids, phenolic compounds, minerals, and vitamins [21]. Therapeutic effects of Aloe vera in wound healing [22], inflammation, intestinal absorption, and reducing oxidative status were assessed in recent research [23]. It also has been used empirically to increase high-density lipoprotein, reduce low-density lipoprotein, and decrease glycemia in diabetics [19]. Furthermore, the anti-inflammatory effects of Aloe vera in the human colon were confirmed in vitro by Langmead et al. [24]. In 2017, the healing effect of the aqueous leaf extract of Aloe vera in an animal model of experimentally induced colitis was investigated. The favorable effects confirmed through the significant reduction in Bax mRNA expression and elevation in Bcl-2 mRNA expression when compared with the colitis group without treatment [25]. In another study, 50 and 300 mg/kg of Aloe vera gel extract were used to evaluate the improvement in the symptoms of UC in rats. According to microscopy and macroscopic observations, the symptoms of UC were improved significantly [26]. Park et al. showed that 0.1% and 0.5% aloesin supplement (one of the compounds of Aloe vera) decreased the myeloperoxidase (MPO) activities as well as TNF-α and interleukin-1β (IL-1β) mRNA expressions on the UC rat colitis model [27]. In another study, glucomannan extracted from Aloe vera balanced pro- and anti-inflammatory cytokines regulated the expressions of TLR-2 and improved the health state of colitis in mice [28]. Similarly, assessments on polysaccharides extracted from Aloe vera on UC-animal models depicted an improvement in colitis, via JAK2, p-JAK2, STAT-3, and p-STAT3 protein expression [29]. In a randomized, double-blind, placebo-controlled trial, oral Aloe vera gel (100 mL twice daily for 4 weeks, in a 2 : 1 ratio) was administered for active UC patients. The supplement reduced the clinical colitis activity index and histological scores significantly during treatment with Aloe vera [30]. However, it seems that further evaluation about the therapeutic potential of Aloe vera extract on UC as well as its effect on new biochemical factors related to UC is needed to get more insight into signaling pathways. Furthermore, in the current study, for the first time, the different routes and doses of Aloe vera administrations (intragastrically and rectally) were studied. Regarding the therapeutic dose of Aloe vera used in the previous studies with no report of toxicity in the tested range, 200 and 400 mg/kg Aloe vera extract were chosen for further study [31–35]. This study was designed to evaluate and compare the dose and route treatment of Aloe vera extract on colitis in rats and its impacts on proinflammatory cytokines. 2. Materials and Methods 2.1. Ethical Statement The animal experiments were performed in accordance with the guidelines of the Laboratory Animal Center of Shiraz Medical University (No. 91-01-36-4560). All the experimental procedures were strictly conducted according to the international standards and national legislation on animal care and the Animal Research Reporting In Vivo Experiments (ARRIVE) guidelines. Experimental research on the plant was under international legislation and guidelines of the Pharmacognosy Department of Shiraz University of Medical Sciences, Shiraz, Iran. At the end of the study, rats were euthanized with the rapid and humane method using a 70% volume displacement rate of CO2 increased to around 100% in the induction chamber. 2.2. Study Design and Induction of Colitis The Laboratory Animal Center of Shiraz University of Medical Sciences with a pathogen-free environment, constant temperature (23 ± 2), and acceptable humidity (55 ± 5%) provided us with 70 male Wistar rats (175–215 grams) supplied with a balanced diet along with free access to water. The rats were fasted with free access to water for 24 h before induction of colitis. After rats were anesthetized with ketamine (50 mg/kg i.p), the rubber-tipped gavage needle was inserted into the anus of rats (7 cm) and 1 ml solution of 2,4,6-trinitrobenzenesulfonic acid (TNBS, 150 mg/kg dissolved in ethanol) was slowly injected into the colon while the control group received only ethanol. Animals were held in the head-down position for 30 seconds and then returned to their cages [36–38]. Later, water and food were available. 12 hours after colitis induction, the treatments were started and continued one a day for six consecutive days. The effectiveness of treatment was assessed by clinical, macroscopic, biochemical, and histopathological assessments. The rats’ general conditions were assessed daily. 2.3. Experimental Animals A total of 70 Sprague Dawley male rats (aged 10-12 weeks, weighing initially 220 ± 20 gram) were obtained from the Laboratory Animal Center of Shiraz University of Medical Sciences. Animals were divided into seven groups (10 rats per group, n = 10). Group (1): healthy control group Group (2): TNBS-induced colitis untreated rats Groups (3): TNBS-induced colitis treated rats who received 200 mg/kg Aloe vera extract once a day, intragastrically Groups (4): TNBS-induced colitis treated rats who received 400 mg/kg Aloe vera extract once a day, intragastrically Groups (5): TNBS-induced colitis treated rats who received 200 mg/kg Aloe vera extract once a day, rectally Groups (6): TNBS-induced colitis treated rats who received 400 mg/kg Aloe vera extract once a day, rectally Group (7): TNBS-induced colitis treated rats who received 500 mg/kg sulfasalazine once a day, intragastrically as a positive control group The dose of Aloe vera extract for treatments was selected according to previously reported research [33–35]. According to the published articles, evaluation on the acute and subacute toxicity of Aloe vera in rats indicated that the methanol extract at the doses of 1, 2,4, 8, and 16 g/kg B.wt did not produce significant toxic effects [31]. In the other study, assessments on the subacute toxicity test showed that Aloe vera did not produce marked subacute toxic effects up to a maximum concentration of 3330 mg/kg body weight on rats with no mutagenic activity in ICR mice exposed to 10000 mg Aloe vera/kg body weight [32]. As a result, at the tested dose of Aloe vera extract, the toxic effect in rats without colitis was not assessed. Sulfasalazine also was purchased from Merck chemical company. The appropriate amount of extract or sulfasalazine based on the treated group was dissolved in sterile water. Intragastric administration was used in conscious rats with biomedical needles (length 76.2 mm, diameter 3 mm, straight). To prepare extract for rectal administration, 5% glycerol was mixed with 2% sodium carboxymethyl cellulose (NaCMC) as an inert preservative substance [39]. Next, 200 and 400 mg/kg body weight of the dried extract was dissolved in deionized water, and the mixtures were gradually added to the glycerol-NaCMC solution. The gel was homogenized for 30 minutes, and the gel was collected in an aluminum tube in the refrigerator. For rectal administration, the gavage needle was inserted into the anus of rats (7 cm) and 1 ml of the prepared gel was injected [40]. 2.4. Plant Extract Aloe vera leaves were obtained in Shiraz, Fars Province, Iran, and its species was endorsed by SUMS taxonomists at a pharmacy school. 100 g dried Aloe vera was powdered and percolated with 70% ethanol (3 times), at room temperature, and the extracts were filtered and evaporated under reduced pressure to acquire 9.8 g of dried extracts (9.8% yield). This procedure was repeated several times to get enough amounts of extract for in vitro and in vivo studies. 2.5. Macroscopic Scoring The dosage and period of treatments were accompanied by daily body weights, gross stool evaluation for visible and/or occult bleeding. On the last day of the experiment (7th), the degree of colonic inflammation and damage was scored (Table 1) as described by Morris et al. with slight modifications [41, 42]. Score Gross morphology 0 No damage 1 Localized hyperemia, but no ulcers or erosions 2 Ulcers or erosions with no significant inflammation 3 Ulcers or erosions with inflammation at one site 4 Two or more sites of ulceration and/or inflammation 5 Two or more major sites of inflammation and ulceration or one major site of inflammation and ulceration extending >1 cm along the length of the colon
... Current chemical drugs used for the treatment of UC are associated with many side effects. Recent complementary medicines, especially herbal remedies, are used increasingly by patients with inflammatory bowel conditions and have been reported to be effective [19][20][21][22]. Quercus brantii (QB) also known as Oak, from the Fagaceae family, is widely grown in Zagros mountainous region in the west and southwest of Iran [23,24]. ...
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Objectives: Ulcerative colitis is a common subtype of persistent inflammatory bowel disease with high morbidity consequences. Despite unknown definite pathogenesis, multiple anti-inflammatory medications are used for its treatment. Traditionally, Quercus brantii (QB), mostly available in the Middle East, has been used for gastrointestinal disorders. Other beneficial effects associated with QB include reduction of oxidative stress, inflammations, homeostatic instability, and improvement in clinical conditions. Materials and methods: This experimental study was designed to assess the possible therapeutic effects of QB on UC and compare its effects with those of sulfasalazine. Of the 70 Wistar rats clustered in seven groups, ten received only alcohols and sixty were confirmed to be suffering from trinitrobenzene sulfonic acid- (TNBS-) induced colitis. Four groups received different dosages of QB extract via oral and rectal routes, one received sulfasalazine, and the other remaining two groups received nothing. The effects of QB were evaluated by assessing macroscopic and histologic scoring, measuring inflammatory mediators, and determining oxidative stress markers. Results: Comparing to the untreated TNBS-induced control groups, QB-treated groups showed a dose- and route-dependent improvement comparable with sulfasalazine. Treating rats with QB reduced the microscopic and macroscopic damage, decreased TNF-α, IL-6, NO, MPO activity, and MDA content, increased superoxide dismutase (SOD) activity, and reduced body weight loss. Conclusions: Our data recommended the anti-inflammatory and antioxidant effects of QB extract in a dose-dependent manner.
... For instance, the aqueous extract of Melilotus officinalis (yellow sweet clover) could decrease pathologic lesions and malondialdehyde (MDA) content and improve weight gain in 3% acetic acid-induced ulcerative colitis in rat. This may be due to its active substances such as cinnamic acid, coumarin, p-coumaric acid, catechin, caffeic acid, gallic acid, quercetin, and chlorogenic acid and also its high ferric reducing antioxidant power (Safarpour et al. 2015). The hydroalcoholic extract of Calendula officinalis (marigold) in both oral and enema application also significantly increased weight gain, resolved acute inflammation and granular atrophy, increased myeloperoxidase activity, and decreased MDA level after UC induction . ...
... For instance, I and coworkers reported the beneficial effects of some plant species in diabetes mellitus [1][2][3][4], ulcerative colitis [5][6][7][8], thyroid abnormal functions [9,10], hyperlipidemia [3], and intestinal morphology [11]. A similar trend is also existed about the OM. ...
... Animals were off fed for 36 hours before induction of UC to empty the colon. UC was induced according to the previously reported protocols (12). All animals were fasted overnight and their bowels were cleaned before induction of UC. ...
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Background: Ulcerative colitis (UC) is a multi-factorial disease with unknown etiology and has many clinical manifestations.Objectives: The current study aimed to evaluate the effects of sesame oil (SO) and grapeseed oil (GSO) on acetic acid-induced UC inrats.Materials and Methods: Eighty male rats were divided into eight groups as health control (HC1), received normal saline; HC2, receivedSO; HC3, received GSO; negative control (NC), UC and normal saline; positive control (PC), UC and mesalamine; SO, UC andSO; GSO, UC and GSO, and SO + GSO. The daily weight changes, serum levels of oxidative stress markers and lipid profile plus colonmacroscopic and microscopic histological changes were measured at the end of the seventh day.Results: Significant differences were detected betweenHC1 and PC on the 3rd (P = 0.002), 4th (0.013) and 6th days (0.014) and betweenHC1 and NC on the 4th day (0.027) in weight of rats. Use of GSO alone or in combination with SO decreased the extent of the changesboth in macroscopic and microscopic indices and also at the inflammation level. The most significant decrease in the MDA leveland the most obvious increase in the TAC belonged to the GSO group in comparison to the NC group. The lowest cholesterol (51.43± 5.62 mg/dL) and HDL levels (29.29 ± 6.24 mg/dL) were detected in response to SO consumption in comparison to NC group (P =0.030 and P = 0.257, respectively).Conclusions: GSO in combination with SO may be considered as the treatment of choice for UC based on antioxidant andhistopathological evaluations.
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This study was done to investigate the synergistic impacts hydro extract of jujube fruit in combination with Mesalazine (orally) and Asacol (intra-colonic) administration in ameliorating animal model of ulcerative colitis (UC). After the induction of UC and with the development of signs, the treatment groups daily received the hydro extract of jujube fruit (200 mg/kg, orally, enema), Mesalazine (30 mg/kg, orally) and Asacol (10 mg/kg, enema). After 10 days, rats were euthanized and were studied. Findings indicated a significant increase in Myeloperoxidase (161.66 ± 10.40), Nitric oxide (216.01 ± 17.55), IL-6 (138.54 ± 7.02), and TNF-α (123.87 ± 9.80) colon tissue levels and pathological damage of positive control group compared with the negative control group. Hydro extract of jujube fruit in combination with Mesalazine (orally) and Asacol (intra-colonic) group represented a higher capability in significantly decreasing Myeloperoxidase (73.33 ± 9.07), Nitric oxide (81.66 ± 10.50), IL-6 (51.69 ± 5.19), TNF-α (30.59 ± 5.50) levels and pathological damage in compared with the other treatment groups. Considering accessibility and affordability of jujube fruit and the side effects of routine drugs, taking a combination of jujube fruit with low doses of routine pharmaceutical drugs can improve and cure ulcerative colitis disease.
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Drug delivery across the skin is used for several millennia to ease gastrointestinal (GI) ailments in Traditional Persian Medicine (TPM). TPM topical remedies are generally being applied on the stomach, lower abdomen, lower back and liver to alleviate GI illnesses such as dyspepsia, gastritis, GI ulcers, inflammatory bowel disease, intestinal worms and infections. The aim of the present study is to survey the topical GI remedies and plant species used as ingredients for these remedies in TPM. In addition, pharmacological activities of the mentioned plants have been discussed. For this, we searched major TPM textbooks to find plants used to cure GI problems in topical use. Additionally, scientific databases were searched to obtain pharmacological data supporting the use of TPM plants in GI diseases. Rosa × damascena, Pistacia lentiscus, Malus domestica, Olea europaea and Artemisia absinthium are among the most frequently mentioned ingredients of TPM remedies. β-asarone, amygdalin, boswellic acids, guggulsterone, crocin, crocetin, isomasticadienolic acid, and cyclotides are the most important phytochemicals present in TPM plants with GI-protective activities. Pharmacological studies demonstrated GI activities for TPM plants supporting their extensive traditional use. These plants play pivotal role in alleviating GI disorders through exhibiting numerous activities including antispasmodic, anti-ulcer, anti-secretory, anti-colitis, anti-diarrheal, antibacterial and anthelmintic properties. Several mechanisms underlie these activities including the alleviation of oxidative stress, exhibiting cytoprotective activity, down-regulation of the inflammatory cytokines, suppression of the cellular signaling pathways of inflammatory responses, improving re-epithelialization and angiogenesis, down-regulation of anti-angiogenic factors, blocking activity of acetylcholine, etc.
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