Investigation of the effects of the water extract of the plant muscari neglectum on gastric damage in rats with some biochemical parameters

The objective of this thesis was to find out the antioxidant activity and phytochemical content of the water extract of the above-ground part of Muscari neglectum (MN) from Sakarya city, to determine its acute toxicity in the living body, and to evaluate its positive effect against gastric ulcer induced by indomethacin (IND) by biochemical parameters and histopathological observations. The above-ground parts of the Muscari neglectum plant utilised in this study were collected from the Sakarya region, thoroughly washed to remove any extraneous matter, and then dried in the shade at room temperature. For water infusion, the plant was extracted at 60°C for 30 minutes with a plant: water ratio of 1:10. The extract solution was filtered with Whatman type filter paper and frozen at -80°C for 24 hours and then dried in a lyophiliser under cold pressure. After extraction, free radical scavenging activity (DPPH), reducing antioxidant capacity (CUPRAC), iron (II) ion chelating activity (MCA), reducing capacity (FRAP), total phenolic compounds and flavonoids, were analysed by spectrophotometric method and phytochemical contents were identified by LC-MS/MS (liquid chromatography-tandem mass spectrometry). A total of 60 male Sprague-Dawley rats were utilised in the course of the study, with 24 rats employed in acute toxicity models and 36 in gastric ulcer models (n=6). For acute toxicity test, rats were divided into 4 groups according to healthy, 500, 1000, 2000 mg/kg body weights by giving Muscari neglectum plant. The toxic effect symptoms were followed for 24 hours. Rats were distributed into 6 groups as healthy; indomethacin (25 mg/kg); reference [40 mg/kg famotidine (FAM)] and the different concentrations of the Muscari neglectum above- ground water extract (100, 200, 400 mg/kg) by selecting 1/5, 1/10, 1/20 of the given limit dose. In all groups except the healthy group, indomethacin was given orally to the rats 5 min after the doses were administered. The rats were sacrificed under the influence of mild sevofluorane anaesthesia six hours after the experimental application. The stomachs of all the used animals were cut out along the large quartur, rinsed with saline and observed macroscopically. Sections taken from the tissues were histopathologically evaluated by haemotoxylated eosin staining. Biochemically, the activities of the antioxidan enzymes [superoxide dismutase (SOD) and catalase (CAT)] and glutathione (GSH), malonaldehyde (MDA) levels in stomach tissues were analysed using sandwich enzyme-linked immunosorbent assay (ELISA) kits. The rutls showed that the lyophilised water extract of the above-ground part of Muscari neglectum plant yielded 14.974 g%. Total phenolic and flavonoid comppounds were 26.0±1.21 mg GAE/g and 0.45±0.05 mg KUE/g, respectively. The % inhibition values of, free radical scavenging activity (DPPH), iron (II) ion chelating activity (MCA), reducing capacity (FRAP), analyses showed a linear increase with increasing concentration. Our reducing antioxidant capacity (CUPRAC), results were found as 0.06 µM TE. According to LC-MS/MS analysis results, fumaric acid, protocatechuic acid, phloridzin dihydrate and myricetin compounds (0.30281874; 0.16369329; 0.02561198; 0.01061646 mg/g extract, respectively) were found intensely, while acetohydroxamic acid, caffeic acid, syringic acid, resveratrol, kaempferol, gallic acid, 4-hydroxybenzoic acid. It is noteworthy that acid, salicylic acid, quercetin and luteolin compounds (0.0066244; 0.00280178; 0.00575919; 0.00322667; 0.00270288; 0.00455979; 0.00475702; 0.0015531; 0.00627964; 0.00278937 mg/g extract, respectively) are also present. It was determined that the doses of Muscari neglectum administered in the experimental studies did not cause any behavioural alterations (e.g. strange behaviours, eating habits, sleep, tremor, abnormal vocalisation, sensitivity to sound) or result in fatality in rats as a consequence of a 24-hour acute toxicity evaluation. These findings demonstrated that the above-ground water extracts of Muscari neglectum plant did not cause any toxic effects at a dose of 2000 mg/kg. The findings of the study demonstrated that all concentrations of the water extract of the above-ground part of Muscari neglectum plant were able to prevent indomethacin-induced gastric mucosal damage, the strongest antiulcer effect was found in the 400 mg/kg group with 69% (p<0.001), showing similar effects with the reference group. When the Muscari neglectum groups were compared with the IND groups, a statistically significant (p<0.0001) increase was observed in SOD activity in the 400MN group, CAT activity in all groups and GSH in 200MN and 400MN, whereas a statistically significant (p<0.0001) decrease in MDA was observed in the 200MN and 400MN groups. The high antioxidant content of the water extract of the above-ground part of the Muscari neglectum plant has been demonstrated to offer gastroprotective effects in a model of gastric ulceration induced by indomethacin in rats and the most effective dose was found to be 400 mg/kg. When the histopathological observations of Muscari neglectum water extract in gastric tissue were compared with indomethacin, it was found that 200 MN and 400 MN extracts significantly (p˂0.05) reduced the damage findings (lamina epithelialis erosion, epithelial degeneration and necrosis, hyperaemia in vessels). Consequently, it was demonstrated that water extracts of the above-ground part of the Muscari neglectum plant did not cause toxic effects at a dose as high as 2000 mg/kg, and the phenolic components contained in its content had a protective effect on indomethacin-induced gastric damage in rats. It showed this effect by decreasing MDA levels and increasing SOD, CAT, GSH levels. It was also found that the gastroprotective effect increased depending on the dose of water extract of the plant. These effects were also supported by histopathological findings. The protective effect of three different doses (100, 200, 400 mg/kg) of the aqueous extract of Muscari neglectum aerial part against gastric ulcer induced experimentally by indomethacin was investigated by biochemical parameters (SOD, CAT enzyme activities, GSH and MDA levels) and histopathological observations. It was found that the above-ground water extract of Muscari neglectum had a protective effect against gastric ulcer and this effect is thought to be due to the biomolecules with antioxidant properties in the plant component. In the life of living organisms, radical compounds that are toxic to living organisms are formed by various effects during external influences and metabolic reactions, and these cause aging and cell-based harmful effects in living organisms. Phenolic compounds are the most effective molecules in preventing these effects. Furthermore, phenolic compounds have been shown to possess antioxidant properties and to exhibit antiulcer effects. In view of these findings, the combinations of phenolic components in the content of Muscari neglectum above-ground water extracts could be considered as a distinct alternative treatment modality to drugs such as FAM, which is used as a reference, by acting on multiple targets (antioxidant + acid regulator + anti-inflammatory). Further studies are recommended to isolate the synergistic effects of the compounds in Muscari neglectum content from the plant and to conduct clinical studies. It is suggested that this plant can be used as a protective against gastric damage in humans. The results of invitro antioxidant analyses of Muscari neglectum water extract and the existence of phenolic compounds in its content showed that the water extract of Muscari neglectum has very good antioxidant effect. In the context of this study, biochemical analyses and histopathological examinations of in vivo studies on gastrit ulcer model mice showed protective effect of Muscari neglectum water extract against gastric ulcer. It is believed that this positive effect is due to the antioxidant biomolecules present in the Muscari neglectum plant. A thorough examination of the available literature revealed no scientific studies on the effects of water extracts from Muscari neglectum plants on stomach diseases. It is predicted that this study will be an important reference for other researches to be conducted with Muscari neglectum.