Adaptogenic and in vitro antioxidant activity of Pedalium murex Linn. in acute and chronic stress paradigms in rodents
M. Pharm Dissertation Protocol Submitted to
Rajiv Gandhi University of Health Sciences, Karnataka
Bangalore – 560041
By
Mr. Vijay Padasalagi, B. Pharm.
Under the Guidance of
Dr. P. V. Habbu, M. Pharm., Ph. D.
Professor
Post Graduate Department of Pharmacognosy,
SET’S College of Pharmacy,
S. R. Nagar, Near Microwave tower, Dharwad,
Karnataka – 580 002.
RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES,
KARNATAKA, BANGALORE.
ANNEXURE-II
PROFORMA FOR REGISTRATION OF SUBJECTS FOR DISSERTATION
1 / NAME OF THE CANDIDATE AND ADDRESS / Mr. VIJAY PADASALAGIPOST GRADUATE DEPARTMENT OF PHARMACOGNOSY
SET’S COLLEGE OF PHARMACY
S.R.NAGAR,
DHARWAD – 580 002,
KARNATAKA.
2 / NAME OF THE INSTITUTION / SET’S COLLEGE OF PHARMACY
S.R.NAGAR,
DHARWAD – 580 002,
KARNATAKA.
3 / COURSE OF STUDY AND SUBJECT / MASTER OF PHARMACY IN PHARMACOGNOSY
4 / DATE OF THE ADMISSION / JUNE 2009
5 /
TITLE OF THE TOPIC
“ADAPTOGENIC AND IN VITRO ANTIOXIDANT ACTIVITY OF PEDALIUM MUREX LINN. IN ACUTE AND CHRONIC STRESS PARADIGMS IN RODENTS”6 / BRIEF RESUME OF THE INTENDED WORK
6.1 / Need for study
Stress can be described as the sum total of all the reactions of the body, which disturb the normal physiological condition and result in a state of threatened homeostasis. Stress is an internationally recognized phenomenon fortified by advancement of industrialization in a demanding civilization1. Stress has been postulated to be involved in the pathogenesis of variety of diseased states, ranging from psychiatric disorders like depression and anxiety, immunosuppression, endocrine disorders including diabetes mellitus, impotency and cognitive dysfunctions, diseases like peptic ulcer, hypertension and ulcerative colitis2.
Benzodiazepines and anxiolytics, despite having significant antistress activity, have not proved effective against chronic stress induced adverse effects on immunity, behavior cognition, male sexual function, during pregnancy and lactation. Additionally, the problem of tolerance and physical dependence on their prolonged use, limits the clinical utility of these drugs. Therefore there is a need for an effective herbal anti-stress agent in the therapy of stress induced disorders2.
Since the introduction of adaptogens i.e. plant derived biologically active substances, which appear to induce a state of non-specific increase of resistance of the organism to diverse aversive assaults which threatened internal homeostasis and which improve physical endurance for doing work even in adverse circumstances and in difficult environmental condition. These agents are basically preventive rather than curative3. The rasayanas in the Indian system of medicine are the agents which are reputed to promote the health and longevity by augmenting defense against diseases, arresting the aging process, revitalizing the body in debilitated condition increasing the capability of individual to resist the adverse environmental factors and by creating a sense of mental well being. The properties described to ayurvedic rasayanas are similar to those said to be present in adaptogens, which appear to increase nonspecific resistance of body against diverse stressors and help to promote physical and mental state of individual 4.
Plants like Panax ginseng5, Withania somnifera6, Vitis venefera1, Butea monosperma7, Boerhaavia difussa8, Trichopus zeylenicus9, Rubia cordifolia10 are found to be potential antistress herbs in acute and chronic stress models. The present research proposal aims to explore the adaptogenic/antistress activity of a rasayana herb Pedalium murex Linn. using animal models.
6.2 / Review of literature
India has a rich heritage of medicinal herbs which are used by the local population and traditional practitioners for the treatment of several conditions.
Pedalium murex Linn.( Pedaliaceae), commonly called Bara-Gukhru, is distributed in the coastal areas of South India11. In Indian system of medicine the whole plant is used as diuretic, anti-inflammatory, antispasmodic, tonic and rejuvenating (adaptogen) drug12. An infusion or extract prepared from the leaves, stem and fruits in cold water is demulcent and a diuretic found useful in the disorders of urinary systems such as gonorrhea, dysuria and incontinence of urine13,14.
Pharmacological review revealed the antihyperlipidemic15 Hepatoprotective16, ovicidal and ovipositional effect17 of the plant using invitro and invivo studies. An aqueous extract from the plant has been evaluated for its analgesic and antipyretic activities18 (Muralidharan and Balamurugan, 2008).
Phytochemical review demonstrated the presence of 2',4',5'-trihydroxy-5,7-dimethoxyflavone,triacontanyl dotriacontanoate, Luteolin, rubusic acid, nonacosane, tritriacontane, triacontanoic acid, tritriacontanoic acid and sitosterol-β-D-glucoside from the fruits of Pedalium murex Linn.19
6.3 / Objectives of the study
By considering the ethnomedicinal contribution of the plant in traditional system of medicine and based on the pharmacological and phytochemical review the present protocol was designed with the following objectives-
Ø To assess adaptogenic/antistress activity of whole plant crude fractions of Pedalium murex Linn. in acute and chronic stress model (Immobilization induced stress) by following parameters,
(a) Biochemical estimation of glucose, triglycerides, cholesterol, alanine aminotranseferase (ALT), aspartine aminotransferase (AST), and creatine kinase in plasma.
(b) Weight of spleen, adrenal glands and thymus.
(c) Gastric ulcer index.
(d) Effect on sexual behavior.
Ø To assess adaptogenic activity of fractions of Pedalium murex Linn. on Swimming endurance test model by weighing adrenal glands and estimating ascorbic acid and cortisol content of adrenal gland.
Ø In-vitro antioxidant activity of potential fraction(s).
7 / MATERIALS AND METHODS
7.1 / Sources of data
Ø Pharmacology Biochemistry and Behaviour
Ø Journal of Ethnopharmacology
Ø Indian Journal of Experimental Biology
Ø Indian material Medica
Ø Indian medicinal plants- a compendium of 500 species
Ø International Journal of Pharmacology
Ø Pubmed
Ø J-gate@Helinet consortium (RGUHS)
7.2 / Method of collection of Data
Data will be generated using laboratory experimental techniques. In vivo evaluation activities are carried out on rats and mice of either sex. The results and data are obtained from the present study will be statistically analyzed using the one way analysis of variance (ANOVA) test followed by post hoc Tukey’s multiple comparison test.
7.3 / Pharmacological Studies
A. Immobilization induced stress20:
In our experiment the stress is produced by restraining individual animal inside an acrylic hemicylindrical plastic tube (4.5 cm diameter, 12 cm long) for a period of 150 min once only in AS and once daily for seven consecutive days in CS as describer earlier.After the treatment protocol serum will be separated and used to estimate glucose, cholesterol, alanine aminotranseferase (ALT), aspartine aminotransferase (AST), and creatine kinase using autoanalyser. The adrenals spleen and thymus will be dissected and weigh. The stomach will be dissected out and cut open along the greater curvature for scoring the incidence of ulcer. Sexual function text will be performed in a separate group of animals.
B. Swimming endurance test21:
Swiss albino mice (15-20gm) of either sex will be used for swim endurance test. Animals are forced to swim in glass chamber (30cm x 30cm x 15 cm) containing water at room temperature after the drug treatment for 7 days. The mice will allowed to swim till they got exhausted and the moment they drowned will be considered as the end point. Animal will be sacrificed and weight of adrenal gland and amount of ascorbic acid and cortisol in adrenal gland will be estimated.
C. In vitro antioxidant activity22:
v DPPH (Diphenyl picryl hyrazyl) assay.
v Hydroxy radical scavenging activity.
v Lipid peroxide scavenging activity.
7.4 / Does the study require any investigation or interventions to be Conducted on patients or other humans or animals? If so please describe briefly.
The above studies require investigation to be done on the albino rats and Swiss mice strain for the determination of adaptogenic activities. The studies are planned in accordance with the procedure reported in the literature.
7.5 / Has ethical clearance been obtained from your institution in case of 7.4?
The study has been given clearance from the ethical committee of institution (copy enclosed).
8 / REFERENCES
1. Satyanarayana S, Srinivas N, Rajabhanu K, Sushruta K, Krishna M. Adaptogenic and nootropic activities of aqueous extract of Vitis vinifera (grape seed): an experimental study in rat model. BMC Compl Altern Med 2005;5:1.
2. Muruganandam AV, Vikas K, Bhattacharya SK. Effect of poly herbal formulation, EuMil, on chronic stress induced homeostatic perturbation in rats. Ind J Exp Biol 2002;40:1151-60.
3. Bhattacharya SK, Arunabh B, Amit C. Adaptogenic activity of Siotone, a poly herbal formulation of ayurvedic rasayanas. Ind J Exp Biol 2000;38:119-28.
4. Bhattacharya SK and Muruganandam AV. Adaptogenic activity of Withania somnifera: an experimental study using rat model of chronic stress. Pharmacol Biochem Behav 2003;75:547-55.
5. Emilia N, Mariana A, Angelo AI. The aphrodisiac and adaptogenic properties of ginseng. Fitoterapia 2000;71:S1-S5.
6. Jayant ND. Adaptogenic and cardioprotective action of Ashwagandha in rats and frog. J Ethnopharmacol 2000;70:57-63.
7. Bhatwadekar AD, Chintawar SD, Logade NA, Somani RS, Veena SK, Kasture SB. Antistress activity of Butea monosperma flowers. Ind J Pharmacol 1999;31:153-55.
8. Krupavaram B, Venkatrao N, Nandan kumar K, Gowda TS, Shalam MD and Shantakumar SM. A study on adaptogenic activity of extract of Boerhaavia diffusa (linn). Ind Drugs 2007;44(4).
9. Singh B, Gupta DK, Chandan BK. Adaptogenic activity of a glycol-peptido- lipid fraction from the alcolic extracts of Tricopus zeylenicus Gaertn. Phytomed 2001;8(4)283-91.
10. Rupali AP, Swati CJ, Sanjay BK. Antihyperglycemic, antistress and nootropic actvityof roots of Rubia cordifolia Linn. Ind J Exp Biol 2006;44:987-92.
11. Nadakani KM. Indian material Medica 3rd Edn. Vol-II,Popular Prakashan Bombay.
12. Warrier PK, Nambiar VPK, Thakur RS, Ramankutty C. Indian medicinal plant- a compendium of 500 species. Orient Longman, Delhi 1994;4:230-32.
13. Chopra RN, Nayar SL, Chopra IC. Glossary of Indian Medicinal Plants. Ist Edn. National Institute of Science Communication (CSIR) New Delhi 1999;10.
14. Shukla YN, Khanuja SPS. Chemical pharmacological and botanical studies on Pedalium murex Linn. J Med Aroma Plant Sci 2004;26:64-69.
15.Mukund NB, Muralidharan P, Balamurugan G. Antihyperlipidemic activity of Pedalium murex(Linn)fruits on high fat diet fed rats Int J Pharmacol 2008;4(4):310-13.
16. Ladani K, Patel NJ, Patel N, Solanki A. Hepatoprotective activity of aqueous-alcoholic extract of Pedalium murex(Bada gokharu)in ethanol and isonizid hepatotoxic rats Ind J Pharmacol 2008;40(S2):S87.
17. Sahayraj K, Joe AR, Francis B. Ovicidal and Ovipositional effect of Pedalium murex Linn. (Pedaliaceae) root extracts on Dysdercus cingalatus (Fab.) Entomon 2006;31(1):57-60.
18. Muralidharan P, Balamurugan G. Analgesic and anti-inflammatory activities of aqueous extract of Pedalium murex Linn.2008;28:84-87.
19. BhakuniRS, ShuklaYN, ThakurRS. Flavonoids and other constituents from Pedalium murex Phytochem 1992;31(8):2917-18.
20. Deepak R, Gitika B, Gautam P, Raghvendra P, Satyawan S, Hemant K S. Adaptogenic effect of Bacopa monniera. Pharmacol Biochem Behav 2003;75:823-30.
21. Kannur DM, Hukkeri VI, Akki KS. Adaptogenic activity of Caesalpinia bondus seed extract in rats. J Ethnopharmacol 2006;108(3):327-31.
22. Veerapur VP, Prabhakar KR, Vipin KP, Machendar RK, Ramakrishana S, Mishra B et al. Ficus racemosa stem bark extract:A potent antioxidant and a probable natural radioprotector. Evidenc Comple Alter Med (eCAM). 2007:1-8.
9
/SIGNATURE OF THE CANDIDATE
/ (Mr. V. S. Padasalagi.)10 / REMARKS OF THE GUIDE / The above mentioned information and literature has been extensively investigated, verified and was found to be correct. The present study will be carried out under my supervision and guidance.
11 / NAME AND DESIGNATION OF GUIDE
SIGNATURE
/ Dr. P. V. HABBU, M. Pharm., Ph.DProfessor,
Post Graduate Department of Pharmacognosy,
SET’s College of Pharmacy, S. R. Nagar,
Dharwad. Karnataka – 580 002.
12
/NAME AND DESIGNATION OF CO – GUIDE
SIGNATURE / -----13 / NAME AND DESIGNATION OF HOD
SIGNATURE / Dr. P. V. HABBU, M. Pharm., Ph.D
Professor & HOD
Post Graduate Department of Pharmacognosy,
SET’s College of Pharmacy, S. R. Nagar,
Dharwad. Karnataka – 580 002.
14
/REMARKS OF PRINCIPAL
/ The above mentioned information is correct and I recommend the same for approval.15 / NAME OF THE PRINCIPAL
SIGNATURE / Dr. V. H. KULKARNI, M. Pharm., Ph.D
Principal,
SET’s College of Pharmacy,
S. R. Nagar, Dharwad. Karnataka – 580 002.