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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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.
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.