EVALUATION OF TEST FORMULATION FOR ANTI-ULCERATIVE ACTIVITY USING STRESS INDUCED ULCER IN EXPERIMENTAL RAT

                                                                     STUDY PROTOCOL

EVALUATION OF TEST FORMULATION FOR ANTI-ULCERATIVE ACTIVITY USING STRESS INDUCED ULCER IN EXPERIMENTAL RAT

1.0 INTRODUCTION:

Gastric ulceration is a benign lesion on the mucosal epithelium upon exposure of the stomach to excess acid and aggressive pepsin activity. It possibly due to disturbance in balance between the aggressive factors like pepsin, acid and H. pylori and the defensive factors such as bicarbonate, gastric mucus, and prostaglandins.

 

2.0 TEST SYSTEM DETAILS:

Species             : Rattus norvegicus

Strain                : Sprague dawley; Wistar.

Age                   : 6-8 Weeks

Body Wight        : 150-200g

Sex                   : Male or Female

No. of animals    : 8 /Group

Total animal       : 56 + 5 Extra = 61

3.0  ALLOCATION OF GROUPS:

Groups	Treatment	Dose; ROA	No. of Animals
G1	Normal Control	0.9% Normal saline /Na-CMC	8
G2	Disease Control	0.9% Normal saline/Na-CMC	8
G3	Reference Drug- Cimetidine	100 mpk; p.o.	8
G4	Test Formulation-1	X mpk; p.o.	8
G5	Test Formulation-1	XX mpk; p.o.	8

# 5 extra animals will be taken extra due to variability in the disease development and possibilities of animal mortalities

 

4.0 METHOD:

·       Animals will be acclimatized for 7 days prior to main experiment and maintained all standard condition as per CPCSEA guidelines.

·       Animals will be randomly divided into seven groups containing eight animals in each.

·       To induce ulcer the 36h fasted animals will be cold immobilized individually in restraint cages and will kept at 3˚±1˚C for 4 h.

·       Group of animals G1 will be served as normal control (without ulceration) and administered with 0.25% sodium CMC at 10ml/kg.

·       Group of animals G2 will be served as ulcerated control and administered with 0.25% sodium CMC at 10ml/kg.

·       Group of animals G3 will be served as reference drug and administered with omeprazole at the dose of 30mg/kg p.o.

·       Similarly, group of animals G4 to G7 will be administered with test compound/test sample at different dose level.

·       All drug, vehicle and test sample will be administered 1h prior to induction of ulcer.

·       After 4h post ulcer induction, the animals will be humanely sacrificed by high dose of thiopental sodium.

·       The stomachs will be removed, opened along the greater curvature for further analysis.

·       5ml of distilled water will be added and the resultant solution will be centrifuged at 3000 rpm for 10 min. The supernatant will be separated and stored for biochemical analyses.

·       The cleaned stomachs will be preserved in 0.1 M phosphate saline buffer (1:4 (w/v), pH 7.4) prior to macroscopic examination and homogenization.

·       Cleaned stomachs will be scored using dissecting microscope with square-grid eyepiece based on grading on a 0–5 scale (depicting severity of vascular congestions and lesions/hemorrhagic erosions).

·       Mean ulcer score for each animal will be expressed as ulcer index (U.I) and the percentage of inhibition against ulceration will be determined using the expressions:

 

U.I. = [Ulcerated area/total stomach area] × 100.

 

%Ulcer inhibition = [U.I. in control − U.I. in test] × 100/U.I. in control

 

5.0 End Point Parameter:

·       Ulcer Index

·       % Ulcer inhibition

·       Histopathology of stomach tissue

·       pH of GI content

·       Superoxide dismutase (SOD) and malondialdehyde (MDA) (In stomach homogenate)

 

6.0  REFERENCE(S):

 

6.1 Paulo Tamashiro Filho, Balogun Sikiru Olaitan, Danielle Ayr Tavares de Almeida, Joaquim Corsino da Silva Lima, Poliana Guerino Marson-Ascencio, Se´ rgio Donizeti Ascencio, Fabrı´cio Rios-Santos, Domingos Tabajara de Oliveira Martins. Evaluation of antiulcer activity and mechanism of action of methanol stem bark extract of Lafoensia pacari A. St.-Hil. (Lytraceae) in experimental animals. Journal of Ethnopharmacology 144 (2012) 497–505.

6.2 Ketan Vinayakrao Hatwarea Sanjay Sharmaa Kiran Patila Meghanath Shetea Sravani Karria Gaurav Gupta. Evidence for gastroprotective, anti-inflammatory and antioxidant potential of methanolic extract of Cordia dichotoma leaves on indomethacin and stress induced gastric lesions in Wistar rats. Biomedicine & Pharmacotherapy Volume 103, July 2018, Pages 317-325.

6.3 Woretaw Sisay Zewdu1 Tezera Jemere Aragaw. Evaluation of the Anti-Ulcer Activity of Hydromethanolic Crude Extract and Solvent Fractions of the Root of Rumex nepalensis in Rats. Journal of Experimental Pharmacology 2020:12 325–337.

                                   

                                 END OF DOCUMENT

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1. List of All SOPs and Documents for In-vivo Laboratory

2. List of All SOPs and Documents for Study Protocol

3. List of Pharmacological Study Models with details about disease Inducing agent, Standard or Reference Drug and related parameters.

4.List of OECD Guidelines or Toxicological Studies

5.List of Pharmacopeial Test or Quality Control Test or Batch Testing Parameters (In-vivo)

6.STUDY PROTOCOL: EVALUATION OF ANTI-ULCER ACTIVITY OF NATURAL PROBIOTIC FORMULATION USING EXPERIMENTAL ANIMALS

7.EVALUATION OF TEST FORMULATION FOR ANTI-ULCERATIVE ACTIVITY USING ALCOHOL INDUCED ULCER IN EXPERIMENTAL RAT

8.EVALUATION OF TEST FORMULATION FOR ANTI-ULCERATIVE ACTIVITY USING INDOMETHACIN INDUCED ULCER IN EXPERIMENTAL RAT

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