Chitosan Loaded Mucoadhesive Microspheres of Gliclazide - Journal

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Nitin M et al./ Influence of Vitamin C with Lansoprazole in Pylorus Ligation Induced Ulcer Model in Rats 8 In pylorus ligation induced ulcer model , the rats were divided into 3 groups of 6 animals each. The animals of Group I were treated with vehicle and the animals of Group II were treated with standard, i.e., lansoprazole 0.54 mg/200 g b.w orally. The animals of Group III were treated with vitamin C and lansoprazole i.e., 4.5 mg and 0.54 mg per 200 g b.w orally respectively. The drugs were administered daily for 5 days. On th 5 day, the rats were fasted for 24 h before pyloric ligation. Care was being taken to avoid coprophagy at the end of 24 h, the rats were anaesthetized with anesthetic ether. Abdomen was opened by a midline incision. The stomach was lifted out and a ligature was placed at the pyloric sphincter without causing any damage to its blood supply. The stomach was replaced carefully and abdomen wall was sutured in two layers. After 6 h, the rats were sacrificed with excess of anesthetic ether, and the stomachs were dissected out. Gastric juice was collected and drained into test tubes and then centrifuged at 1000 rpm for 10 min and the volume of supernatant was noted. The pH of the gastric juice was recorded by pH meter. Then the contents were subjected for the analysis of free and total acidity. The stomachs were opened along the greater curvature then washed under running water to see the ulcers in the glandular portion of the stomach. The number of ulcers per stomach was noted and scoring was done microscopically with the help of hand lens 9 (10x) . The recording was 0 for normal colored stomach, 0.5 for red coloration, 1 for spot ulcer, 1.5 for hemorrhagic streaks, 2 for ulcer ≥ 3 ≤ 5 and 3 for ulcer > 5. The mean ulcer score for each animal is expressed as ulcer index and the percentage protection was calculated by using the following 10 formula : % Protection = [(UI control – UI treated) /UI control] x 100 9 Determination of free acidity and total acidity 1 ml of gastric juice was pipetted into 100 ml conical flask. It was diluted to 10ml with distilled water and 2 –3 drops of Topfer's reagent was added and titrated with 0.01 N sodium hydroxide until all traces of red color disappear and the color of the solution turns to yellowish orange. The volume of the alkali added was noted. This volume corresponds to free acidity. Then 2 - 3 drops of phenolphthalein solution was added and titration was continued until a definite red tinge reappears. Again the total volume of alkali added was noted. The volume corresponds to total acidity. Acidity was calculated by using the following formula: Volume of NaOH x Normality of NaOH x 100 Acidity = ------------------------------------------------------------- mEq/L/100 gm 0.1 11 Histopathological evaluation The stomachs were immersed in 10 % formalin solution embedded in paraffin wax, sections of thickness of about 5 128 µm were cut and stained with haemotoxylin and eosin. Sections were sections were examined for histopathological changes such as congestion, haemorrhage, necrosis, inflammation, infiltration, erosion and ulcers. Statistical analysis The results were expressed as mean ± SEM, (n=6). Statistical analysis was performed using student 't' test. P value less than 0.05 was considered to be statistically significant. RESULTS It is evident from Table 1 that the effect of combination group i.e., vitamin C and lansoprazole showed significant (P < 0.001) reduction in all biochemical parameters like volume, free acidity, total acidity and increase in pH of gastric juice, while ulcer index also showed significant (P < 0.05) reduction when compared to control and standard lansoprazole. The percentage protection of combination group was found to be 94.3% when compared to control and standard lansoprazole alone (82.8%). The histopathological examination using haematoxylene and eosin staining also revealed the protective activity of combination group when compared to control and standard lansoprazole (Fig. 5 to Fig. 8). DISCUSSION Ulcer is a recurrent disease affecting large populations in all geographical regions, and reactive oxygen species have been implicated in the pathogenesis of a wide variety of clinical disorders and gastric damage. Peptic ulcers result from an imbalance between defensive (cytoprotective) and offensive factors (gastric acid), association with Helicobacter pylori infection and increased use of NSAIDs like aspirin and 12 indomethacin , causing damage by inhibiting the 13 biosynthesis of cytoprotectve prostaglandins . 7 The Shay model of pylorus ligation is a simple, reproducible and highly predictable model for the screening and evaluation of antiulcer drugs. It utilizes neither the exogenous ulcerogens nor the induced exogenous interfering factors. In this model gastric ulceration may be due to increased secretion of acid pepsin which leads to auto digestion of gastric mucosa, decreased mucosal blood flow and breakdown of mucosal 14 barrier . In addition pyloric ligation may reduce glutathione 15 levels of gastric mucosa and increase the lipid peroxidation . There is one more report that pepsin is active in lower pH. Therefore the reduced gastric ulcer in this model may occur due to the reduction in acid secretion and increased gastric pH. In this condition the activity of pepsin is minimized and consequently the digestion of mucosal barrier is prevented. In the present study, from (Table 1), and (Fig. 1 to Fig. 8) the significant reduction in ulcer index and other biochemical parameters of gastric juice like volume, free acidity, total acidity and increase in pH by combination group suggests that RJPS, Jul - Sep, 2011/ Vol 1/ Issue 2
Nitin M et al./ Influence of Vitamin C with Lansoprazole in Pylorus Ligation Induced Ulcer Model in Rats Table 1: Effect of Vitamin C and Lansoprazole in pylorus ligation induced gastric ulcer model in rats: Gr. Treatment Dose/200 g Volume of Free Total pH of Ulcer % No. rat gastric acidity acidity gastric Index Protec juice (ml) (mEq/l)/ (mEq/l)/ Juice tion 100 g 100 g 1 Control Distilled water 0.5 ml 8.133 ± 0.11 112.0 ± 0.73 124.3 ± 0.55 1.800 ± 0.07 5.833 ± 0.10 - 2 Lansoprazole 0.54 mg 4.567 ± 0.14 56.67 ± 0.55 67.67 ± 0.55 6.767 ± 0.09 1.000± 0.18 82.8% 3 Vitamin C + (0.54+4.5) 2.467 ± 0.15*** 34.00 ± 1.31*** 44.00 ± 0.96*** 7.467 ± 0.11*** 0.333 ± 0.10* 94.3% Lansoprazole mg Values are the mean ± S.E.M.n=6, *P < 0.05 and ***P < 0.001 compared with lansoprazole. Fig. 1: Stomach epithelium of normal rat in pylorus ligation model Fig. 1: Stomach epithelium of control rat in pylorus ligation model Redness and ulcer Fig.3: Stomach epithelium of standard lansoprazole treated rat in pylorus ligation model 129 Fig. 4: Stomach epithelium of standard lansoprazole treated rat in pylorus ligation model Fig. 5: Histopathological slide of normal rat showing normal histology. Fig. 6: Histopathological slide of control rat showing redness, congestion, hemorrhagic streaks, edema, ulceration, necrosis and dilation of blood vessels. RJPS, Jul - Sep, 2011/ Vol 1/ Issue 2
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R G U H S Dr. Amit Agarwal, India D
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