EVALUATION OF HEPATOPROTECTIVE ACTIVITY OF HERBAL FORMULATION(S) USING ALCOHOL INDUCED HEPATOTOXICITY IN EXPERIMENTAL ANIMALS

                                                                STUDY PROTOCOL

EVALUATION OF HEPATOPROTECTIVE ACTIVITY OF HERBAL FORMULATION(S)
 USING ALCOHOL INDUCED HEPATOTOXICITY IN EXPERIMENTAL ANIMALS
 
 
1.0 Introduction 

Alcoholic liver disease (ALD) is characterized by morphological changes ranging from hepatic steatosis,

 inflammation and hepatic necrosis to progressive fibrosis. Ethanol (EtOH) oxidation generates toxic metabolites, 

free radicals and induces a state of oxidative stress characterized by an enhancement of lipid peroxidation and 

depletion of endogenous antioxidant enzyme activities such as superoxide dismutase (SOD), catalase (CAT) and 

glutathione peroxidase, which contribute to the pathogenesis of ALD.

2.0 TEST SYSTEM DETAILS:

Species             : Rattus norvegicus (Rats)

Strain                : Wistar or Sprague Dawley

Age                   : 8-10 weeks

Body Wight        : 120-200 g

Sex                   : Male or Female

No. of animals    : 8 /Group

3.0  ALLOCATION OF GROUPS:

Groups	Treatment	Dose; ROA	No. of Animals
G1	Normal Control	0.25% Na- CMC; qdX8W	8
G2	Ethanol Control	0.25% Na- CMC; qdX5W	8
G3	Positive Control- Silymarin	100 mpk; qdX5W	8
G4	Plant Extract/Herbal Formulation-1	X mpk, p.o	8
G5	Plant Extract/Herbal Formulation-1	XX mpk, p.o	8
G6	Plant Extract/Herbal Formulation-2	X mpk, p.o	8

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

4.0 METHOD:

·       Rats weighing 150-200gm will be included in the present investigation.

·  The animals will be housed in polypropylene well-aerated cages at normal atmospheric temperature (21 ± 3°C) and normal 12 h light/dark cycle.

·     Rats will be free access to water and supplied daily with laboratory standard diet of known composition ad libitum.

·       Animals of group G1 and G2 will be administered with 0.25% sodium carboxymethylcellulose (CMC).

·      All the animals (except G1) will be orally administered with ethanol at 5ml/kg body weight, daily for 5 weeks to induce chronic liver and kidney injury.

·       Animals of Group G3 will be received reference drug Silymarin at the dose of 100mg/kg b.wt p.o.

·       Animals of group G4, G5 and G6 will be administered with test sample at different dose level.

·       All the animals will be treated for 5 weeks with vehicle or Silymarin or test samples at different dose levels.

·       Animals will be anaesthetized and blood sample will be collected on week 1, 2, 3, 4 and 5 for Albumin, Globulin, Total Bilirubin, AST, ALT, Total protein, albumin, ALP, LDH analysis.

     Furthermore, at the end of the experiment all the animals will be humanely sacrificed, liver and kidneys will be isolated weighed and stored in 10 % buffered formalin and further processed for histopathology.

     5.0 END POINT PARAMETER(S):

·       Clinical observation

·       Feed water consumption

·       Body Weight

·       Liver Function Profile - Albumin, Globulin, Total Bilirubin, AST, ALT, Total protein, albumin, ALP, LDH, GGT and CRP

·    Anti-oxidant Profile – Catalase (CAT), malonaldehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPX) and glutathione (GSH).

·       Liver Lysosomal enzymes- Cathepsin D and β-Galactosidase

·       Histopathology of Live tissue

6.0  REFERENCE(S):

6.1 Qigui Mo , Gao Zhou , Baibo Xie, Bingxin Ma , Xinyu Zang, Yuxin Chen , Linyou Cheng, James Hua Zhou and Youwei Wang. Evaluation of the hepatoprotective effect of Yigan mingmu oral liquid against acute alcohol-induced liver injury in rats. BMC Complementary Medicine and Therapies (2020) 20:32 https://doi.org/10.1186/s12906-020-2817-9.

6.2 Tong Zhou, Yu-Jie Zhang, Dong-Ping Xu, Fang Wang, Yue Zhou, Jie Zheng, Ya Li, Jiao-Jiao Zhang, and Hua-Bin Li. Protective Effects of Lemon Juice on Alcohol-Induced Liver Injury in Mice. BioMed Research International Volume 2017, Article ID 7463571, 8 pages https://doi.org/10.1155/2017/7463571.

6.3 Poonam Lodhi, Neeraj Tandan, Neera Singh, Divyansh Kumar, and Monu Kumar. Camellia sinensis (L.) Kuntze Extract Ameliorates Chronic Ethanol-Induced Hepatotoxicity in Albino Rats. Evidence-Based Complementary and Alternative Medicine Volume 2014, Article ID 787153, 7 pages http://dx.doi.org/10.1155/2014/787153.

6.4 Wang, Xu, Dong, Ke, Ma, Yujing, Jin, Qizhi, Yin, Shujun and Wang, Shan. "Hepatoprotective effects of chamazulene against alcohol-induced liver damage by alleviation of oxidative stress in rat models" Open Life Sciences, vol. 15, no. 1, 2020, pp. 251-258. https://doi.org/10.1515/biol-2020-0026

6.5 Abdelaziz Souli, Hichem Sebai, Latifa Chehimi, Kaı Rtibi, Haifa Tounsi , Samir Boubaker, Mohsen Sakly, Jamel El-Benna5 and Mohamed Amri. Hepatoprotective effect of carob against acute ethanol-induced oxidative stress in rat. Toxicology and Industrial Health 1–9

                                   END OF DOCUMENT

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