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International Food Research Journal 23(3): 1164-1172 (2016)
Journal homepage: http://www.ifrj.upm.edu.my
1*Nyam, K.L., 1Tang, J. L. K. and 2Long, K.
1Department of Food Science and Nutrition, Faculty of Applied Sciences,
UCSI University, 56000 Kuala Lumpur, Malaysia
2Malaysian Agricultural Research & Development Institute (MARDI)
PO Box 12301, 50774 Kuala Lumpur
Anti-ulcer activity of Hibiscus cannabinus and Hibiscus sabdariffa seeds in
ulcer-induced rats
Abstract
The aim of this research is to determine the antiulcer properties and percentage protection of
Hibiscus cannabinus and Hibiscus sabdariffa seed samples towards ulcer-induced Sprague
dawley rats. Rats were divided into six groups each for each ulceration method and fed with
distilled water, Omeprazole, H. cannabinus seed oil (HCSO), H. cannabinus seed extract
(HCSE), H. sabdariffa seed oil (HSSO) and H. sabdariffa seed extract (HSSE), respectively
via oral administration. Among the two plants tested, H. cannabinus showed the best protection
percentage towards ethanol, non-steroidal anti-inammatory drugs (NSAIDs) and cold restrain
stress induced ulcers. H. cannabinus seed extract (HSSE) exhibited an exceptionally high ulcer
protection of 74.98 ± 0.78% against NSAIDs induced ulcer. The gastric lesions were controlled
primarily by both mucosal protection and acid inhibition. In conclusion, addition of these seeds
to the daily diet may reduce free radical activity in the body and reduce the risk of developing
peptic ulcer disease.
Introduction
Approximately four million people worldwide
are affected by peptic ulcer disease (PUD) every year
due to the imbalance between the aggressive and
defensive factors (Zelickson et al., 2011). Around 10
to 20% of patients’ encountered complications while
10 to 40% are mortality cases (Lau et al., 2011). This
disease is considered common around the world and
therefore deserves more attention in controlling the
widespread of PUD. The ingestion of non-steroidal
anti-inammatory drugs (NSAIDs) and Helicobacter
pylori (H. pylori) infection are two major factors that
increases the complications and risk of developing
PUD. Synthetic drugs for instance antacids, proton
pump inhibitors and histamine-2 blockers are
used to treat ulceration. However, the side effects
are unavoidable such as diarrhoea, headache and
constipation (Srinivas et al., 2013). Despite this,
herbal medicine is still the mainstay of 75-80% of
population and is considered a better option as they
have fewer side effects, better compatibility with
the human body and thus, potential for new drugs
(Parekh et al., 2005).
Kenaf (Hibiscus cannabinus L.) is a warm-
season annual ber crop that has been commercially
cultivated in Asia, such as China, India, Malaysia
and Thailand (Liu, 2000). Many researches around
the globe have been focusing on the utilization of
H. cannabinus stems for papermaking and pulping
as it was an excellent cellulose ber source while
the potential for using H. cannabinus seeds as a
source of edible oil is overlooked. H. cannabinus
seed is high in oil content ranging from 21.4% to
26.4% (Nyam et al., 2009). The oil is very similar
to cotton seed oil. Edible oil is yield from seeds
of this plant and is used as rst-class cooking oil
and margarine production. This oil is nutritionally
benecial to health because of the abundant amount
of monounsaturated and polyunsaturated fatty acids
available (Coetzee et al., 2008). The seed oil is edible
and can be kept for it has a long shelf life. Besides,
the oil is a rich source of bioactive compounds
lled with high antioxidative, anticancer and lipid
lowering cholesterol properties (Nyam et al., 2009).
It has been reported by Yazan et al. (2013) that H.
cannabinus seed oil was cytotoxic towards ovarian
cancer and colon cancer cell lines. Alpha-linolenic
acid (omega-3 fatty acid) was also found in the seed
which acts as a precursor of eicosanoids with anti-
inammatory and antithrombotic activity (Ruiz et
al., 2002). However, a higher antioxidant yield and
Keywords
Ethanol-induced ulcer
Non-steroidal anti-
inammatory drug
(NSAID)-induced ulcer
Cold-restraint stress induced
ulcer
Ulcer index
Gastric juice
Article history
Received: 21 May 2015
Received in revised form:
2 September 2015
Accepted: 21 September 2015
1165 Nyam et al./IFRJ 23(3):1164-1172
increased in antioxidant activity can be obtained
from H. cannabinus seed extract (HCSE) (Wong
et al., 2014) . The ndings of Lee et al. (2007)
indicated that ethanol extract of H. cannabinus may
produce promising anti-inammatory properties and
is capable of treating macrophage related diseases
for example septic shock, arteriosclerosis and
rheumatoid arthritis.
Roselle (Hibiscus sabdariffa L.) is a tropical wild
plant widely used in many countries as a medical
herb for its antioxidant, antibacterial (Oboh and
Elusiyan, 2004) and lipid-lowering properties (Faraji
and Tarkhani, 1999). Jadhavet et al. (2009) reported
that the leaves and owers promote hair growth and
aid in ulcer healing. H. sabdariffa seeds are used in
China for vegetable oil production, sauces and as
coffee substitute. H. sabdariffa seed oil (HSSO) is
composed of mainly oleic and linoleic fatty acids
(Mohamed et al., 2007). According to Lautenschlager
(2003), linoleic acid is commonly used for cosmetic
production. It moisturises the skin due to its high
penetrability, aids in healing dermatoses and
sunburns and may help in reducing acne lesions. It
has anti-inammatory effects, improves blood lipids
and lowers blood pressure. The therapeutic effect of
this extract will encourage its use in the treatment of
inammation. Lee et al. (2007) indicated that ethanol
extract of H. cannabinus may produce promising anti-
inammatory properties and is capable of treating
macrophage related diseases, for example septic
shock, arteriosclerosis and rheumatoid arthritis.
Plants like H. cannabinus and H. sabdariffa are
regaining the attention of many researches due to
their many potential benets for human health and
prevention of diseases. This provides an alternative
strategy and offers a bright future in the development
of antiulcer drugs for the treatment of gastric
ulcer. The objectives of this study was to compare
the antiulcer properties of H. cannabinus seed oil
(HCSO), H. cannabinus seed extract (HCSE), H.
sabdariffa seed oil (HSSO) and H. sabdariffa seed
extract (HSSE) in ulcer induced Sprague dawley rats.
Materials and Methods
Materials
Ten kilograms of dried H. cannabinus seeds were
obtained from Malaysian Agricultural Research and
Development Institute (MARDI), Serdang, Selangor,
Malaysia. Ten kilograms of H. sabdariffa seeds
were sourced from the Department of Agriculture
Plantation, Rembau, Negeri Sembilan, Malaysia.
Male Sprague dawley rats were obtained from the
institutional animal house of University Kebangsaan
Malaysia, Bangi, Malaysia for the antiulcer study.
H. cannabinus (HCSE) and H. sabdariffa seed
extracts (HSSE) preparation
A 500 mL Scott bottle was prepared and lled with
80% ethanol to a total volume of 500 mL. A total of
50 g of grinded H. cannabinus seed was added to the
solvent, followed by ultrasonic extraction (Ultrasonic
Homogenizer Labsonic P, 400 W, Sartorius, AG)
for 30 min with a 5 min pulse duration period and
a 5 min pulse interval period. The extraction was
repeated for 3 cycles. The H. cannabinus seed extract
(HCSE) collected was centrifuged at 3500 rpm for
10 min. The supernatant of the kenaf seed extract
was collected and ltered; the pellet was discarded.
The ltered supernatant was subjected to rotary
evaporation (Rotavapor R-200, Buchi, Switzerland)
(Wong et al. 2014). The same extraction procedures
were repeated for H. sabdariffa seed to obtain H.
sabdariffa seed extract (HSSE).
H. cannabinus seed oil (HCSO) and H. sabdariffa
seed oil (HSSO) preparation
The HCSO were extracted from the seeds with
soxhlet extractor using hexane at 60 ºC for 3 hours.
The oil was then recovered by evaporating off the
solvent using rotary evaporator Model N-1 (Eyela,
Tokyo Rikakikal Co., Ltd., Japan) and residual
solvent was removed by ushing with 99.9% (Ng
et al. 2014). The above procedure was repeated to
obtain H. sabdariffa seed oil (HSSO) by replacing
H. cannabinus seed powder with H. sabdariffa seed
powder.
Evaluation of antiulcer activity
Experimental animals
All Sprague dawley rats weighing about 200-
250 g were housed in standard cages of three to a
cage. The animals were taken care under the standard
laboratory conditions with controlled lighting (12
hours light and dark cycles) and at room temperature
(25 ± 3ºC).
The animals were fed with meat-free rat and
mouse-diet from Speciality Feeds (Glen Forrest,
Western Australia) and have free access to water ad
libitum throughout the duration of the experiment.
Animals were allowed to acclimatise for two
weeks prior to experiment to ensure animals are in
a healthy state and to minimise the stress factors
cause by transportation of animals. All experimental
procedures were conducted after the approval of
the Faculty’s research ethics committee of UCSI
University, Cheras, Malaysia with the approval code
Nyam et al./IFRJ 23(3):1164-1172 1166
Proj-FAS-EC-13-038 and were in strict accordance to
the institutional animal ethical committee guidelines
for care and use of laboratory animals.
Experimental design
The animals were grouped into six groups
comprising of ve animals each. Food pellets were
withdrawn 24 hours before the start of the experiment
and while the animals had free access to water ad
libitum. The compositions of experimental treatments
were as shown in Table 1.
Ethanol-induced ulcer
After 24 hours of fasting, the animals were treated
with treatments as shown in Table 1. One hour after
the treatment, all six groups (n=5) of rats were orally
administered with 5 mL/kg of ethanol each according
to Robert (1979) with modications. After one hour,
the rats were euthanized with excess chloroform and
their stomachs were immediately excised and opened
along the greater curvature, cleared of residual matter
with saline and the inner surface was examined for
ulceration.
Non-steroidal anti-inammatory drug (NSAID)-
induced ulcer
The procedures were similar to that used in
ethanol induced ulceration except that the control
group (Group I) received indomethacin only. Lesions
were induced according to the method of Nawafor
et al. (2000) with modications. After 24 hours of
fasting, the animals were treated with treatments as
shown in Table 1. One hour after the treatment, all six
groups (n=5) of rats received indomethacin 100 mg/
kg orally. After four hours, the rats were euthanized
with excess chloroform and their stomachs were
immediately excised and opened along the greater
curvature, cleared of residual matter with saline and
the inner surface was examined for ulceration.
Cold-Restraint Stress Induced Ulcer
Four groups (Group I, II, III and IV) (n=5) of
rats were used. After 24 hours of fasting, Group
I (control) received distilled water (10 mL/kg),
Group II was given omeprazole (30 mg/kg), Groups
III and IV were fed with 500 mg/kg of HCSO and
HCSE (p.o), respectively. The method described by
Senay and Levine (1967) was adopted with slight
modications. One hour after treatment, gastric
ulceration was induced by immobilising the animals
in a closed cylindrical restrainer immersed vertically
up in a water bath of (18 ± 2)ºC to the xyphoid
level, in the presence of intense light for three hours
to induce stress ulcer. After three hours, the rats
were euthanized with excess chloroform and their
stomachs were immediately excised and opened
along the greater curvature, cleared of residual matter
with saline and the inner surface was examined for
ulceration.
Measurement of ulcer index
The average length (mm) of all lesions of each
Table 1. Effect of HCSO, HCSE, HSSO and HSSE on various parameters in ethanol induced,
indomethacin-induced, and cold restraint stress-induced gastric ulcer in rats
Values are expressed as mean ± SD (n=5). Statistical analyses were performed using ANOVA followed by
Tukey test. Mean values at the same column with different superscript letters are signicantly different at
p < 0.05.
1167 Nyam et al./IFRJ 23(3):1164-1172
stomach were measured and classied into three
levels as shown in Table 1. The ulcer index (UI) was
determined as follows:
Ulcer index (UI) = 1 × (number of ulcers level I) + 2
× (number of ulcers level II) + 3 × (number of ulcers
level III)
Percentage of ulcer protection was calculated as
below (Navarrete et al. 1998):
% Protection = (Ulcer index Control - Ulcer index Test) × 100
Ulcer index Control
Determination of pH
Gastric juice (1 mL) was diluted with 1 mL
distilled water and was measured using a pH meter
(Dashputre and Naikwade, 2011).
Statistical analysis
All experiments were performed in triplicate,
unless stated otherwise. The results were expressed as
mean ± standard deviation (SD). Statistical analyses
were performed by one-way analysis of variance
(ANOVA) followed by Tukey test. A p-value of
<0.05 was considered statistically signicant. The
statistical analyses were performed using Minitab
software, version 16.1.
Results and Discussion
Based on Table 1, the oral administration of
HCSO, HCSE, HSSO and HSSE demonstrated gastro
protective effects against gastric lesions induced by
ethanol. They showed signicant reduction in ulcer
index, as well as increased in protection from gastric
lesion. Rats fed with HCSO exhibited the highest
protection percentage of 54.54 ± 2.74% while HSSO
showed the lowest protection percentage of only
31.78 ± 3.83%.
All the four samples showed signicant (p <
0.05) antiulcer effect at the dose of 500 mg/kg per
oral dose. This may be due to the cytoprotective
effect of the samples via antioxidant effects. Ethanol
at a dose of 5 ml/kg, produced red coloration, sported
ulcer and severe gastric hemorrhagic erosions in the
control rat.
The development of the haemorrhagic lesions and
necrotic tissue injury as observed in may be caused
by stasis in gastric blood ow, in which gastric blood
ow cease. Alcohol is able to penetrate the gastric
mucosa rapidly, causing cell rupture and plasma
membrane damage thus the intracellular membrane
permeability to sodium and water was increased. The
stomach appears to be larger in size and was lled
with gastric contents. Besides, when an extreme
amount of calcium is accumulated in the intracellular,
this results in development of gastric mucosal injury
which led to cell death and exfoliation in the surface
epithelium (Raju et al., 2009). The direct damage of
gastric mucosal cells that resulted in the development
of free radicals and hyperoxidation of lipids would
increase the gastric lesions.
From this study, ethanol is known to be very
harmful to the stomach since the gastric mucosa was
affected by the disruption of protective barrier and
provoking prominent microvascular changes within a
few minutes after application (Moleiro et al., 2009).
Therefore, mucus secretion is vital in protecting
the gastric mucosa from gastric lesions. The results
clearly indicated a signicant cytoprotective and
gastric antiulcer activity of H. cannabinus and H.
sabdariffa. Flavonoids are among the cytoprotective
materials able to increase mucus, bicarbonate and
prostaglandin secretion, strengthening of gastric
mucosal barrier and scavenging of free radicals which
are very important in preventing ulcerative and erosive
lesions of gastrointestinal tract (Sachin and Archana,
2009). The results of the present study suggested that
HCSO may be benecial in the treatment of gastric
lesions. The effectiveness of HCSO in preventing
peptic ulcer was further investigated in non-steroidal
anti-inammatory drugs (NSAIDs) induced ulcer and
cold restraint water immersion stress induced ulcer.
Based on Figure 1 (A), severe injuries are seen
in the gastric mucosa. Ethanol produced extensive
visible hemorrhagic necrosis of gastric mucosa. Rat
pre-treated with 10 ml/kg omeprazole (30 mg/kg)
had injuries of the gastric mucosa. However they
are milder compared to the injuries seen in negative
control rats (Figure 1 (B)). Mild injuries are seen in
the gastric mucosa of rat pre-treated with HCSO (500
mg/kg) and HCSE (500 mg/kg). The oil and extract
reduces the formation of gastric lesions induced by
acidied ethanol (Figure 1 (D)). Moderate injuries
are seen in the gastric mucosa of rat pre-treated with
HSSO (500 mg/kg). The oil was able to reduce the
formation of gastric lesions induced by acidied
ethanol (Figure 1 (E)). Sported injuries are seen in
the gastric mucosa, but they are milder compared to
the injuries seen in negative control rats when pre-
treated with 500 mg/kg of HSSE (Figure 1 (F)).
The gastric pH of rats that treated with omeprazole
(positive control) at 30 mg/kg was 8.29 ± 0.77,
slightly higher compared to the Group I (negative
control). While the rats treated with HSSE showed
the lowest gastric pH of only 4.12 ± 0.35 (Table 1).
From Table 1, the rats that pre-treated with HCSO
Nyam et al./IFRJ 23(3):1164-1172 1168
and HSSO had gastric pH of 6.79 ± 0.23 and 6.78 ±
0.53, respectively, which was nearer to neutral (pH
7).
Non-steroidal anti-inammatory drugs (NSAIDs)
induced ulcer
In this study, HCSO and HCSE were able to
produce a signicant reduction in gastric mucosal
damage induced by indomethacin, indicating a
possible increase of prostaglandin synthesis. The
analgesic effect of NSAIDs has caused NSAID
induced ulcer to be painless, thus endangering
consumers. They are usually prescribed as pain killers
to soothe headaches, sprains and arthritis symptoms.
The protective action of HCSE against
indomethacin induced gastric lesions may be due to
5-lipoxygenase inhibitory effect of cyclooxygenase
(Rainsford, 1987). Besides, it may have stimulated
prostaglandin secretion or produce prostaglandins
like substances to protect the stomach. Omeprazole
was incorporated into the study to examine the
participation of proton pump inhibitor on NSAIDs
induced ulceration. The signicant percentage of
protection of gastric ulcer in the rats pre-treated with
HCSE was compatible with omeprazole, the standard
drug used in this experiment to cure ulcer. The acidity
of gastric content in rats treated with HCSO, HCSE,
HSSO and HSSE were higher compared to negative
control group (Table 1).
The gross appearances of excised stomachs of
indomethacin induced gastric lesions are shown in
Figure 2. The lesions in the negative control group
stomach appeared as elongated bands of thick, black
coloured lesions with yellowish mucus. Mucus
secretion is vital in protecting the gastric mucosa
from gastric lesions. The ulcer lesions produced in
HCSE fed rats were not as prominent compared to
the other samples. Omeprazole, as a proton pump
inhibitor has been widely used by people around
the world as an acid inhibitor agent for managing
gastric acid secretion related disorders and protects
the stomach gastric mucosa (Li et al., 2004). The
mechanism of omeprazole works in a way that it is
able to bind very specically to a single subunit of
the gastric H+, K+-ATPase and inactivates it. This
inhibits the acid secretion despite of the source of
secretory stimulation. Omeprazole is very benecial
and effective for both long and short-term use in
treating peptic ulcer disease (AlRashdi et al., 2012).
HSSO portrayed the highest gastric pH of 4.47
± 0.21, followed by HCSO, HCSE and HSSE with
gastric pH of 3.81 ± 0.33, 3.33 ± 0.20 and 1.86 ±
0.12, respectively in comparison to negative control
(Group I) with the lowest pH (Table 1). The oils
Figure 1. Gross appearance of the gastric mucosa in
ethanol induced rats. (A) 10 mL/kg distilled water
(negative control). (B) Omeprazole (30 mg/kg). (C)
HCSO (500 mg/kg). (D). HCSE (500 mg/kg). (E). HSSO
(500 mg/kg). (F) HSSE (500 mg/kg)
Figure 2. Gross appearance of the gastric mucosa in
NSAIDs induced rats. (A) 10 mL/kg distilled water
(negative control). (B) Omeprazole (30 mg/kg). (C)
HCSO ( 500 mg/kg). (D) HCSE (500 mg/kg). (E) HSSO
(500 mg/kg). (F) HSSE (500 mg/kg)
1169 Nyam et al./IFRJ 23(3):1164-1172
and extracts showed protection against gastric
lesions produced by indomethacin. Omeprazole
also accelerates ulcer healing by reducing the acid
secretion thus pH of gastric juice increases. NSAID
such as indomethacin have the ability to cause gastro-
duodenal ulcer by reducing prostaglandin synthesis,
which is important for increasing blood ow,
secretion of mucus and bicarbonate and in inhibiting
the secretion of hydrochloric acid (Konturek et al.,
2005). Thus, the gastric juice pH for indomethacin
induced ulcer method was much lower compared to
ethanol induced ulcer method.
According to Figure 2 (A), rat pre-treated with
10 mL/kg distilled water (negative control) had
severe injuries in the gastric mucosa. Indomethacin
produced elongated bands of thick, black coloured
lesions with yellowish mucus. There are some red
dots seen on the gastric mucosa in rat pre-treated with
omeprazole (30 mg/kg) however, they were much
milder compared to the injuries seen in negative
control rats (Figure 4.2 (B)). Dotted erosions are seen
in rat fed with HCSO (500 mg/kg) (Figure 4.2 (C)).
Rat pre-treated with HCSE (500 mg/kg) showed mild
dotted lesions (Figure 4.2 (D)). Moderate injuries
are seen in rat pre-treated with HSSO (500 mg/kg)
(Figure 4.2 (E)). For the rats pre-treated with 500
mg/kg HSSE, erosion injuries on the gastric mucosa
are milder compared to the injuries seen in negative
control rats (Figure 4.2 (F)).
Cold restraint water immersion stress induced ulcer
The cytoprotective activity of HCSO and
HCSE was examined on cold restrained water
immersion stress rats. This is because HCSO and
HCSE portrayed the best protection percentage in
ethanol induced ulceration and NSAID induced
ulceration, respectively. Cold restrained water
immersion stress provides both emotional stress
as well as physiological stress to the animals. It is
one of the best methods for inducing stress-induced
gastric lesions in rats. Some of the factors known to
play a role in inducing gastric lesions are by auto-
digestion of gastric mucosal barrier, accumulation of
hydrochloric acid and generation of oxygen derived
free radicals. Stress-related mucosal bleeding has
known to be the cause of morbidity and mortality
in critically ill patients (Metz 2004). Therefore,
necessary prevention steps are important to avoid
stress-related ulceration. Arakawa et al. (1997) stated
that stress causes increases in plasma norepinephrine
and epinephrine levels, whereas stress-induced
syndromes are reduced by inhibitors of adrenaline
receptors or sympathetic nerve excitation.
Cold restrained water immersion stress produced
visible gastric lesions in all rats after three hours
as anticipated. In the cold restraint stress induced
gastric ulcer rats, a signicant reduction (14.00 ±
1.00 and 15.00 ± 1.00) in ulcer index was observed
in rats treated with 500 mg/kg of HCSO and HCSE
in comparison with negative control group (Group
I) (Table 1). The average ulcer index in negative
control group (Group I) was very high (35.33 ± 2.52).
The ulcers produced are most likely mediated by
production of histamine followed by acid secretion
and reduction in mucus production. The blood ow
reduces drastically in cold stress immobilisation
leading to local hypoxia and ischemia (Hase and
Moss, 1973). Lesions observed were clearly visible
and were spread throughout the stomach area. The
percentages of protection against ulcer were 60.38 ±
0.33, 57.54 ± 0.38 and 95.33 ± 1.44 for groups treated
with HCSO, HCSE and omeprazole, respectively.
Thus, omeprazole had more antiulcer effect in cold
restrained water immersion stress induced rats
followed by HCSO and HCSE.
According to Table 1, animal treated with
omeprazole portrayed the highest gastric pH of 6.74
± 0.22, followed by HCSE and HCSO with gastric
pH of 4.83 ± 0.33, 3.52 ± 0.33, respectively in
comparison to negative control (Group I) with the
lowest pH (Table 1). There is a correlation between
pH and % protection as shown in Table 1. As gastric
pH increases, % protection increases. The blood ow
may decline when stress is present and this would
enhance the production of certain substance present
in the stomach such as acid, bile and pepsin. Thus,
the gastric pH dropped due to the over production
of gastric acid and this would cause gastro-duodenal
Figure 3. Macromorphological presentation of the gastric
mucosa in cold restraint water immersion stress induced
ulcer in rats. (A) 10 mL/kg distilled water (negative
control). (B) Omeprazole (30 mg/kg). (C) HCSO (500 mg/
kg). (D) HCSE (500 mg/kg)
Nyam et al./IFRJ 23(3):1164-1172 1170
ulceration. Rats fed with omeprazole (Group II)
portrayed a signicant (p < 0.05) increase in gastric pH
of 6.74 ± 0.22 and increase in protection percentage
of 95.33 ± 1.44 compared to negative control group.
Results showed that rats pre-treated with HCSO
and HCSE before stressing had signicantly reduced
in ulcer index compared to rats pre-treated with
distilled water (negative control, Group I). It was
observed that the protection of gastric mucosa was
more prominent in rats pre-treated with 500 mg/
kg HCSO as demonstrated in Table 1. The plant
oil and extracts are used to stimulate the mucosal
defence mechanism of the stomach by increasing the
amount of mucus production to protect and facilitate
in repairing of damaged epithelial cells (Goel and
Sairam, 2002). HCSO signicantly acts as a rst
line defence against cold restraint stress-induced
gastric ulcers by showing cytoprotective property.
There may be some antioxidant properties present in
HCSO that provides some protection towards the rats
induced with cold restrained water immersion stress.
Rats fed with omeprazole (Group II) showed
a signicantly (p < 0.05) lower ulcer index (1.67
± 0.58) compared to negative control group. As
according to Brzozowaski et al. (2000), proton pump
inhibitors and histamine H2-receptor antagonists are
known for speeding up the healing process of gastric
lesions and in inhibiting mucosal injury. Therefore,
gastric ulcer was signicantly inhibited showing a
protection percentage of 95.33 ± 1.44. The stomach
surface remained pink, showing there was neither
inammation nor bleeding (Figure 3). The pH of
gastric contents in HCSO was signicantly lower
compared with HCSE (pH 3.52 ± 0.33 vs. pH 4.83
± 0.33). Omeprazole showed the highest gastric pH
of 6.74 ± 0.22. The auto digestion of gastric mucosal
barrier caused by the overproduction and build-up
of gastric acid is known to be the most important
reason for the formation of gastric lesions caused by
stress (Sairam et al., 2003). This decreases the gastric
mucus in reducing the mucosal damage of stomach.
Apart from that, the gastric acid secretion plays an
important role in the progression from an erosive
mucus layer to a gastric lesion.
The protective activity against cold restraint
stress may be due to the antioxidant activity present
in HCSO, which helps in strengthening the rats’
physiological competence to reduce stress induced
ulcers. The results obtained indicate that the HCSO
had the ability to maintain the cell membrane integrity,
to protect the gastric mucosa against oxidative
damage and to strengthen the mucosal barrier, which
acts as the rst line defense against ulcer agents. The
diagrams in Figure 3 show the excised stomach of
respective groups and the severity of ulcer on the
lining.
Figure 4.3 shows that the (A) Rat pretreated
with 10 mL/kg distilled water (negative control)
suffered from severe injuries in the gastric mucosa.
Cold restrained water immersion stress produced
brownish-black coloured lesions with mucus. Not
much gastric ulcer was formed in rat pre-treated
with 10 ml/kg omeprazole (30 mg/kg) however,
there were a few small dots on the surface of the
stomach [Figure 4.3 (B)]. Dotted erosions are seen
in the gastric mucosa of rat pre-treated with HCSO
(500 mg/kg) and HCSE (Figure 4.3 (C) and 4.4 (D)).
HCSO and HCSE reduced the formation of gastric
lesions induced by cold restrained water immersion
stress.
Conclusion
Both plant (H. cannabinus and H. sabdariffa)
seed oils and extracts reduced the ulcer index and
increased percentage of protection against ulceration,
suggesting their role in protecting against gastric
ulcer. This suggests the potential of H. cannabinus
and H. sabdariffa in strengthening the stomach
defense against inammation and further progress to
gastric ulcer. All samples showed protective activity
against ethanol, NSAIDs and cold restrain induced
ulcer activities. However, H. cannabinus samples
produced better protection against ulcer compared
to H. sabdariffa samples. HCSO showed the highest
protection against ethanol and cold restrained stress
ulcer, whereas HCSE exhibited the highest protection
against NSAID ulcers. Hence, this result has revealed
that H. cannabinus seed is a superior potential of an
antiulcer agent.
Acknowledgement
Financial support of this work by internal funding
from CERVIE UCSI University (Proj-In-FAS-022) is
gratefully acknowledged.
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