Pharmacological and toxicological evaluation in rats of Dissotis rotundifolia, a medicinal plant traditionally used in ghana for the management of peptic ulcer
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Date
OCTOBER 2015
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Abstract
The ethnopharmacological importance of Dissotis rotundifolia (Melastomacea) is
evidenced by its traditional use by various tribes in Ghana for several illness
including gastrointestinal disorders. Despite its widespread use in the treatment
of peptic ulcer, the plant extract has not been subjected to any systematic
pharmacological investigations to ascertain its efficacy and safety. This study
aimed at evaluating the gastroprotective activity, gastrohealing effects and
establishing the possible mechanism of action of the extract of whole plant of
Dissotis rotundifolia (DRE) using chemical and physiological stress-induced ulcer
models in rats. Peptic ulcer was induced using, acetylsalicylic acid, ethanol and
cold restraint stress and either a pre-treatment with DRE (100, 300 and 500
mg/kgbwt) and positive drugs (30 mg/kg bwt omeprazole and 50 mg/kgbwt
ranitidine) before ulcer induction or post-treatment with DRE and positive drugs
after ulcer induction. The antisecretory effect of DRE was evaluated in a bid to
examining the possible antiulcer mechanism. The in vitro anti-Helicobacter
pylori activity of the extract was also assessed in addition to antioxidant effects in
vitro and in vivo. The phytochemical screening of the major chemical
constituents of DRE was also done. The potential toxicological effect of the
extract in Sprague-Dawley rats was evaluated in acute and sub-acute toxicity
studies. The results show a marked percentage protection and curative indices of
pre-treated and treated groups in the cold stress, acetylsalicylic acid and ethanol
induced ulcers in rats. This was coupled to a significant reduction in ulcer
indices. The 300 mg/kg bwt extract pre-treated / post-treated group recorded a
significant reduction in ulcer index compared to DRE doses of 100 and 500
mg/kgbwt group and ranitidine group. Meanwhile statistical analysis showed no
significant difference between the 300 mg/kgbwt DRE and 30 mg/kgbwt
omeprazole treated / pretreated group. The observation was seen in almost all the
ulcer models employed. Also in the antisecretory studies it was observed that the
pretreatment with 300 mg/kg DRE significantly increased gastric secretion (pH,
mucus and titrable acidity) and showed a marked reduction in gastric juice
volume (p < 0.01), when compared with the groups treated with ulcerogen
(acetylsalicylic acid) only without any form of pretreatment. In contributing to
elucidating the possible mechanism of action, results from anti-Helicobacter
pylori activity indicates that plant extracts (200-800 mg/ml) and standard
antibiotic drugs (amoxicillin, clarithromycin) showed antimicrobial activity
against clinical isolates of H. pylori. The zones of inhibition ranged from 13-30
mm. DRE showed statistically lower zones of inhibition compared to standard
drugs (clarithromycin, amoxicillin) (p<0.05). Metronidazole revealed no zone of
inhibition indicating resistance by Helicobacter pylori. Meanwhile maximal
effect was found at 400 mg/mL concentration of DRE. In vivo activity of H+ / K+
- ATPase in negative control CRS, ASA, and EtOH ulcerated rats was found to be
elevated compared to normal, omeprazole and DRE treated groups (p< 0.05). It
was also observed that H+ / K+ - ATPase enzyme activity was low in DRE and
omeprazole treated rats in all models compared to negative control groups
(p<0.05). A marked increase in glycoproteins (sialic acid, hexose, hexosamine
and fucose) was observed in all the pretreated groups compared to negative
iv
control ulcer groups. There was also an increase in the total carbohydrate /
protein ratio of the gastric mucosa in DRE treated rats. Histopathological
examination of stomach of ulcerogens (cold, ASA, EtOH) without drug
pretreatment animals indicated that there was disruption of the surface epithelium
and glandular structure. Drugs (DRE, omeprazole and ranitidine) pretreated
group showed mild damage to the gastric mucosa indicative by a mild disruption
of the surface epithelium with mild edema and leucocyte infiltration into the
submucosal layer. This is indicative of a marked inhibition of gastric ulcer by
DRE, omeprazole and ranitidine. This result demonstrates that the plant extracts
possesses antiulcer effect against cold, acetylsalicylic acid and ethanol induced
ulcers in rats. The anticholinergic experiment showed that the extract decreased
the propulsive movement of charcoal meal through the gastrointestinal tract
(GIT). This observation was significantly (P < 0.01) different from what was
seen in the negative control group. The delayed gastric emptying time increases
the absorption of orally administered drugs. In vitro free radical scavenging
activity suggest that aside the other mechanisms, DRE possesses antioxidant
activity evidenced by an excellent DPPH, nitric oxide, superoxide, hydroxyl
radical-scavenging activities and profound Fe2+ - ascorbate lipid peroxidation
inhibitory activity. Results from the effects of DRE on the gastric mucosa
antioxidant in vivo in ethanol induced ulcer model shows that there was a
significant increase in catalase (CAT) and superoxide dismutase (SOD) activities,
a rise in reduced glutathione (GSH) content and a marked decrease in
malondialdehyde (MDA) levels in DRE and omeprazole pretreated group
compared to negative control (p< 0.05). The preliminary phytochemical
screening, showed the presence of tannins, saponins, terpenoids, flavonoids,
alkaloids and reducing sugars. In the toxicological study, there was no significant
difference in almost all the parameters measured. This was confirmed by the
absence of any damage observed in micrographs obtained from histopathological
examination. The results showed that DRE had no overt organ specific toxicity in
rats within the 14-day administration of the extract thus have a high safety profile.
In conclusion the study demonstrates that the extract is able to perform its
antiulcer effect through inhibition of H+ / K+ - ATPase, increasing mucus content
and mucin activity, acts as an antagonist at muscarinic cholinergic receptor and
inhibition of Helicobacter pylori. The study also shows that DRE performs its
antiulcer effect by acting as a scavenger of Hydroxyl, nitric oxide, DPPH radical
and also as anti-lipid peroxidative. The in vitro scavenging property was
confirmed by results obtained from the in vivo antioxidant experiment indicating
that DRE significantly inhibited the effects of ethanol on lipid peroxidation,
gastric GSH levels depletion and increased CAT and SOD activities. Implicitly it
can be stated that the antiulcer effects of DRE is multifaceted. DRE is acting as
an antiulcer agent by serving as a proton pump inhibitor, increasing mucin
activity, gastric mucus content, pH and titrible acidity, inhibiting growth of H.
pylori, serving as a scavenger of OH, DPPH, NO2 radicals and anti-lipid
peroxidative. Overall these findings provide substantial evidence-based data to
support the traditional medicinal use of the whole plant extract of Dissotis
rotundifolia in the management of peptic ulcer.
Description
A Thesis submitted to the Department of Pharmacology, Faculty of Pharmacy and
Pharmaceutical Sciences, College of Health Sciences in fulfillment of the requirements for the degree of Doctor of Philosophy.