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screening of antiulcer agents

This document summarizes screening methods for antiulcer agents, including both in vivo and in vitro methods. It begins with an introduction to peptic ulcers and classifications of antiulcer drugs. In vivo screening methods described include the pylorus ligation model in rats, ethanol-induced gastric ulcer model, water immersion stress model, and indomethacin-induced ulcer model. The key in vitro screening method discussed is the H+/K+-ATPase inhibition assay to measure inhibition of proton pump activity. The document provides details of procedures and evaluations for each screening model. It concludes with references for further information.

Education◦
SCREENING OF ANTIULCER AGENTS 1 Dr. V.V.P.F’S COLLEGE OF PHARMACY AHMEDNAGAR (2018-2019 ) GUIDED BY Prof. Dr.P.M.GAIKWAD M.Pharm, Ph.D (Head, Dept of Pharmacology) PRESENTED BY Miss. Joshi Uttara L. M.Pharm (sem 1st ) (Pharmacology) Roll no.08
Contents • Introduction • Symptoms of peptic ulcer • Classification • Screening methods 1. In vivo methods 2. In vitro methods • Reference 2
Introduction • Peptic ulcer is one of the most common gastrointestinal diseases. • The drug has been use to prevention of stress ulcer. 3
Symptoms of Peptic Ulcer • Burning abdominal pain • Nausea • Vomiting • Pain in stomach • Weight loss • Constipation 4
PATHOPHYSIOLOGY 5
Classification of Anti-Ulcer Drug 1) H2 receptor antagonist e.g. Cimetidine , Ranitidine , Famotidine 2) Proton pump inhibitors - e.g. Omeprazole ,Pantoprozole,Lansoprazole 3) Anticholinergics e.g. pirenzepine , Telenzepine, Propanthalin 4)Prostaglandin analogues:Misoprostal ,Enprostil 5) Antacids Syatemic:Sodium bicarbonate Nonsystemic:Mg hydroxide 6) Ulcer protective e.g. Sucralfate 6
7)Ulcer Healing drugs : carbenoxolone sodium 8)Anti H pylori drugs: Amoxicillin, Clarithromycin,Metronidazole,tinidazole, Tetracycline. 7
PROTON PUMP INHIBITORS - Most effective drugs in antiulcer therapy. - Prodrug requiring activation in acid environment . - Activated forms binds irreversibly to H+K+ ATPase and inhibit it. - Eg –Omeprazole Pantoprazole - The parietal cell of the stomach is activated by histamine acetylcholine and gastrin. - In addition to the direct action of gastrin and acetylcholine on the parietal cell these two agents may release histamine form a histamine form storage in the gastric mucosa. 8
MECHANISM OF ACTION - Prodrugs inactive at neutral pH . - At pH < 5 rearranges to two charged cationic forms t that bind covalently with SH groups of H+/K+ATPase and inactivate it irreversible. - Also inhibits gastric mucosal carbonic anhydrase. 9
Screening methods • In vitro method 1) Gastric binding assay . 2) H+/K+ ATPase inhibition assay • In vivo methods 1) Pylorus ligation in rats 2) Stress ulcer model . 3) Histamine induced gastric ulcer . 4) Acetic acid induced gastric ulcer 5) Ethanol induced gastric ulcer 6) Indomethacin induced ulcers in rats 10
In vitro methods 11
1) H+/K+ -ATPase inhibition assay • Proton pump > final step in the synthesis of acid by parietal call. Procedure – Incubation of guinea pig gastric mucosa buffer ATP and test compound 12
Add malachite green (colorimetric agent ) after 30 minutes Add 15% sodium citrate after 10 sec . For 45 minutes Released of orthophosphate from ATP quantified by colorimeter at 570 nm Evaluation: • Percentage inhibition of H⁺/K⁺-ATPase is calculated. 13
In vivo method 14
1) Pylorus ligation in rats • Purpose –  A simple and reliable method for production of gastric ulceration in the rat. Based on ligature of the pylorus has been published by shay et al 1945 . The ulceration is caused by accumulation of gastric the stomach 15
Midline abdominal incision is given Ligation of pylorus Test compounds are given either orally or injected S.C. About 17-19 hours after pyloric ligation 16 PROCEDURE Wistar Rat are saturated for 48 hrs. access to water
17 Rats are sacrificed and stomachs are dissected out volume of gastric acid pH no. of ulcer index and severity is noted.
Evaluation • ID50 values can be calculated by probit analysis. • An Ulcer Index (UI) is calculated. • UI = UN+US+UP × 10-1 18 Where, Un=Average number of ulcer per animal, Us= Average of severity score , Up= Percentage of animal with ulcer
2)Ethanol Induced mucosal damaged in Rats PURPOSE Intra gastric application of absolute ethanol is a reproducible method to produce gastric lesions in experimental animal • These lesion can be at least partially inhibited by various drugs such as some prostaglandins • The protective effect against various irritants has been called cytoprotective activity. 19
Wistar Rat are saturated for 18 hrs. access to water After 30 min 99.80% alcohol is given orally. Rate are sacrificed and stomach dissected out A circular full thickened area is cut Evaluated using a light transmission densitometer. The optical density of the test tissues is determined. 20 PROCEDURE
EVALUATION  The significance of differences in optical density Test drug is given orally. . 21
22 Principle: •Exposure of cold conditions to restrained animals •Accelerates the occurrence of gastric ulcers. •Shortens the immobilization time 3) Water immersion induced stress model
PROCEDURE Stress induced ulcers were induced by force swimming in the glass cylinder (height 45cm diameter 35cm) containing water up to 350C maintained at 350C for 3 hrs (Wistar rat) Animals were fasted 24 hrs prior to the experiment 23
All stomachs were opened along the greater curvature ulcer index and % inhibition was calculated. After the drug treatment (standard/test) animals were allowed to swim for 3hrs then animal were dissected stomachs were removed 24
25 Principle: NSAID induced gastric damage ; by blocking COX enzyme, endogenous prostaglandin production inhibited 4) Indomethacin induced ulcer model
PROCEDURE All the animals were fasted 36 hours before administration of indomethacin The animals were divided into groups. each rat was administered with the 20mg/kg indomethacin orally. 30 min prior to the administration of the indomethacin standard/test drug was administered. 26
By titrating with 0.01M NAOH using phenolphthalein as an indicator. Gastric juice estimated for pepsin Ulcer index and % inhibition were calculated The rats were anaesthetized with ether 1 hour latter the stomach was incised through the greater curvature and examined for the number of lesion under the dissecting microscope 27
Reference: 1) H.G. Vogel and W.H. Vogel ,“ Drug Discovery and evaluation pharmacological assay ”, Springer , New York. Page no-1236-1242. 2) K.D. Tripathi Medical Pharmacology Sixth edition, Page no-151-161. 3) S.k.kulkarni,̏ Text Book of Practical Pharmacology ̋,6th edition , Vallabh Prakashan, Page No-107 -108 28
29 4. Datta GK, Sairam K, Priyambada S, Debnath PK, Goel RK: Antiulcerogenic activity of Satavari mandur-An ayurvedic herbomineral preparation. Indian J Expl Biol 2002; 40:1173-77. 5.Mishra L.Wagnar H. Lipid derivatives from mucona pruriens seeds Indian journal of chemistry 2006,45(B):801-804 6.Chopra RN ,Nayar SL , Chopra IC Glossry of indian medicinal Plants ,CSIR , New Delhi 1956 .
Thank you 30