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RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES, BANGALORE
KARNATAKA
ANNEXURE II
PROFORMA FOR REGISTRATION OF SUBJECT FOR DISSERTATION
1.
NAME OF CANDIDATE
Dr. PAVITRA
And Address (in block
POST-GRADUATE STUDENT
letters)
DEPARTMENT OF PHARMACOLOGY
J.J.M. MEDICAL COLLEGE
DAVANGERE - 577004
KARNATAKA
J.J.M.MEDICAL COLLEGE
2.
NAME OF THE
INSTITUTION
DAVANGERE-577004.
POST GRADUATION
3.
COURSE OF STUDY &
SUBJECT
M.D. PHARMACOLOGY.
4.
DATE OF ADMISSION TO
6TH JUNE 2013
COURSE
5.
TITLE OF TOPIC
"ANTIASTHMATIC EFFECT OF
ETHANOLIC EXTRACT OF
MORINGA OLEIFERA
(DRUMSTICK) SEED KERNELS
IN AN EXPERIMENTAL ANIMAL
MODEL".
BRIEF RESUME OF THE INTENDED WORK:
6.1 Need for the study:
Asthma is a chronic inflammatory disorder of the airway that causes recurrent episodes
of wheezing, breathlessness, chest tightness, cough and is usually associated with variable degree
of bronchoconstriction and airflow limitation that is partly reversible either spontaneously or with
treatment [1].
In 2007, it was estimated that 300 million people worldwide suffer from asthma and is
expected to increase by more than 100 million by 2025[2].
The drugs currently used in treating asthma are relatively expensive and have
troublesome adverse effects, like muscle tremor and hypokalemia with β2 agonists[3,4],
theophylline has narrow therapeutic index and requires monitoring of plasma drug levels [5], fluid
retention, weight gain, osteoporosis, hypertension, diabetes, cataract, psychosis is seen with
corticosteroids [6]. Therefore there is a need for search of a drug with less side effects and better
tolerance.
Herbal alternatives with known pharmacological activities offer benefits of being cheap,
natural, having fewer side effects and have health benefits beyond what we strictly expect.
Moringa oleifera is a member of “Moringaceae” family commonly called as
‘Drumstick’ in India. It is a native of Himalayas in northwestern India, but widely cultivated in
tropical and sub-tropical areas [7]. The plant possess antimicrobial activity [8], while the seeds
reportedly have antispasmodic, anti-inflammatory and diuretic activity [9]. Seeds contain
chemical ingredients like benzyl isothiocyanate, benzylglucosinolate, moringine. The alkaloid
“Moringine” in seeds of Moringa oleifera may cause bronchodilatation which is comparable to
ephedrine and thus useful in treatment of asthma [10].
In the present study, we are evaluating anti-asthmatic potential of ethanolic extract of
seed kernels of Moringa oleifera in an experimental animal model like bronchodilatation using
histamine induced bronchospasm in guinea pigs.
6.2. Review of Literature :
Asthma is a disease of the airways that is characterized by activation of mast cells,
infiltration of eosinophils and T helper 2 (TH2) lymphocytes. Mast cell activation by allergens and
physical stimuli releases bronchoconstrictor mediators, such as histamine, leukotriene D4 and
prostaglandin D2 which cause bronchoconstriction, microvascular leakage and plasma
exudation[11].
The causes of airway narrowing in acute asthmatic attacks include contraction of airway
smooth muscle; inspissation of viscid mucus plugs in the airway lumen; and thickening of the
bronchial mucosa due to inflammation leading to edema, cellular infiltration, and hyperplasia of
secretory, vascular, and smooth muscle cells of airway. Of these causes of airway obstruction,
contraction of smooth muscle is most easily reversed by current therapy; reversal of the edema and
cellular infiltration requires sustained treatment with anti-inflammatory agents.
Short-term relief (as in acute asthma) is most effectively achieved by agents that relax
airway smooth muscle, of which β-adrenoceptor stimulants are the most effective and most widely
used. Theophylline, a methylxanthine drug, and antimuscarinic agents are also used for reversal of
airway constriction.
Long-term control (as in chronic asthma) is most effectively achieved with an antiinflammatory agent such as an inhaled corticosteroid. It can also be achieved, though less
effectively, with a leukotriene pathway antagonist or an inhibitor of mast cell degranulation, such
as cromolyn or nedocromil. Finally, clinical trials have established the efficacy of treatment for
severe asthma with a humanized monoclonal antibody, omalizumab, which is specifically targeted
against IgE, the antibody responsible for allergic sensitization [12].
Kirtikar et al (1975) reported alkaloid moringine in Moringa oleifera relaxes bronchioles
and found to have antiasthmatic potential [10].
Shailaja G Mahajan et al (2007) investigated the effect of ethanolic extract of seeds of
Moringa oleifera on the potential prevention of immune mediated inflammatory responses in
toluene diisocyanate induced asthma in wistar rats wherein Moringa oleifera treated rats did not
manifest any airway abnormality[13].
Anita Mehta et al (2008) studied the effect of alcoholic extract of Moringa oleifera seed
kernels on various experimental models of bronchial asthma. Significant (P<0.05) increase in
preconvulsion time was observed due to pretreatment with Moringa oleifera when guinea pigs
were exposed either to acetylcholine or histamine aerosol confirming its antiasthmatic potential[4].
Ganatra Tejas H et al (2012) reported alcoholic extract of Moringa oleifera seed kernels
were found to be spasmolytic in acetylcholine, histamine, Barium chloride and Serotonin (5HT)
induced bronchospasm[14].
From the above literature reviews it is known that alcoholic extract of Moringa oleifera
seed kernels have potential antiasthmatic property that may be due to its bronchodilator, antiinflammatory, spasmolytic activity, but all the above studies have used crude extract of moringa
oleifera, hence the specific active ingredient responsible for bronchodilatation is not been
identified.
6.3. Objectives of study:
 To study the antiasthmatic effect of ethanolic extract of Moringa oleifera seed kernels and
to Compare its action with a standard drug ketotifen fumarate for bronchodilatation
activity.
MATERIALS AND METHODS:
7.1 Source of data:
Dunkon Hartley guinea pigs of either sex weighing 350-500g [4] which are bred in
central animal house of J. J. M. Medical College, Davangere is used. Animals will be randomly
housed at an ambient temperature (22 ± 1ºC) and humidity (55± 5ºC) with a 12 hour light and 12
hour dark cycle. Animals have access to standard pellet diet and water.
7.2 Method of collection of data (including sampling procedure, if any)
Study on Histamine induced bronchospasm in guinea pigs using histamine chamber :
Materials required
Histamine (1%) [15]
0.5% Sodium Carboxymethyl cellulose (CMC) [4].
Ketotifen fumarate [4].
Alcoholic extract of Moringa oleifera seed kernels [4].
Method:
A total of 24 animals (N=24) of either sex weighing 350-500g will be divided into 4
groups of 6 (n=6) animals each. The following schedule of treatment was administered [4]:
Group 1 (Control): Sodium Carboxymethyl cellulose (CMC), 0.5% [4], 0.5ml per 20g body
Weight [17].
Group 2 (Standard): Ketotifen fumarate 1mg/kg body weight, per oral [4].
Group 3 (Test A): Alcoholic extract of Moringa oleifera 100mg /kg body weight, per oral[4]
Group 4 (Test B): Alcoholic extract of Moringa oleifera 200 mg/kg body weight, per oral[4]
The drugs mentioned above were administered orally in 0.5% sodium
carboxymethyl Cellulose (CMC) 0.5ml per 20g body Weight [18]. (It is used as a thickening
agent, as a viscosity modifier and water retaining agent). The single dose treatments were
given one and half hour before the study. Later the animals were exposed to an aerosol of 1%
histamine and time for preconvulsion was noted for each animal.
Histamine induced bronchospasm:
This method involves use of histamine Chamber (24 x 14 x 24 cm) made of perplex
glass in which guinea pigs were exposed to (1%) histamine aerosol under constant pressure
(40mm/hg) and the preconvulsion time that is “Time interval from exposure of histamine to
appearance of signs of convulsion”
[17]
is recorded. As soon as Preconvulsant dyspnea
commences, the animals were removed from the chamber and exposed to fresh air. Animals
which could withstand exposure to histamine aerosols for 15 minutes were considered to be
protected.
Inclusion and Exclusion criteria:
Inclusion Criteria :

Dunkon-Hartley Guinea pigs of either sex weighing between 350-500 g.

Healthy animal with normal behavior and activity.
Exclusion Criteria:

Guinea pigs <300 g or >500 g.

Pregnant female guinea pigs or those who have delivered and lactating till suckling
(around 6 weeks after delivery).

Animals previously used in other experiments were not used till 6weeks.
Statistical analysis: The end point variables are subjected to a suitable statistical analysis
after confirming normality of distribution. If needed these data will be suitably transformed (to
any of the following, log10, aresin, square root etc., depending on the condition).Then any of the
following such as‘t’ test, ANOVA, ANCOVA or MANOVA will be used to draw the inferences.
7.3 Does the study require any investigations or interventions to be conducted on patients
or other humans or animals? If so, please describe briefly.
NA.
7.4 Has ethical clearance been obtained from your institution in case of 7.3?
Yes.
List of references:
1
Kumar V, Abbas AK, Fausto N, Aster JC. Lung; In Robbins and Cortan pathological
basis of disease, (Ed’s) 8th Ed, Gurgaon (India). Saunders Elsevier; 2010; p. 688-737.
2
Global surveillance, prevention and control of chronic respiratory diseases: a
comprehensive approach, WorldHealthOrganization [Internet] 2007.
[Updated2007][Accessedon20.10.2013.[Availablefrom:www.who.int/gard/publications/
GARD%20Book%202007.pdf
3 Haalboom JRE, Deenstra A, Stuyvenberg A. Hypokalemia induced by inhalation
Of Fenoterol. Lancet 1st edition, 1985; p 1125-1127.
4. Nelson HS, Adrenergic therapy of bronchial asthma. J. Allergy Clin. Immunol. 1986;
77:771–785.
5. Nasser SS, Rees PJ. Theophylline; Current thoughts on the risks and benefits of its
Use in asthma. Drug Safety, 1993 Jan; 8(1):12–18.
6. Dajani BM, Sliman NA, Shubair KS, Hamzeh YS. Bronchospasm caused by
intravenous hydrocortisone sodium succinate (Solu-Cortef”) in aspirin-sensitive
Asthmatics. J. Allergy Clin. Immunol.68, 1981; p 201-204.
7. Sharma V, Paliwal R, Pracheta, Sharma S. Phytochemical analysis and evaluation of
antioxidant activities of hydro-ethanolic extracts of Moringa oleifera lam. Pods. J of
Pharm Res 2011; 4(2): 554-557.
8. Caceres A, Cebreva O, Morales O, Miollinedo P, Mendia Pharmacological properties of
M.oleifera 1: Preliminary screening for antimicrobial activity. J. Ethnopharmacol. 33,
1991; p 213-216.
9. Caceres A, Saravia A, Rizzo S, Zabala L, De-Leon E, Nave F. Pharmacological
properties of M. oleifera 2: Screening for antispasmodic, anti-inflammatory and diuretic
activity. J. Ethnopharmacol. 36, 1992; p 233-237.
10. Kirtikar KR, Basu BD. Indian medicinal plants. New Cannaught Place,
Dehradun, 2nd edition, Vol.1, 1975; 676-683.
11. Brunton LL, Chamber BA, Knollmann BC. Goodman and Gillman’s the
Pharmacological basis of therapeutics. 12th Edition, New Delhi, Mc Graw hill
Companies; 2011.
12. Katzung BG, Masters SB, Trevor AJ. Drugs used in asthma: Basic & Clinical
Pharmacology, 11th edition, China: Mc Graw-Hill Companies; 2009. Pages 339-340.
13. Mehta A, Mahajan G S, Inhibitory action of ethanolic extract of seeds of Moringa
oleifera seed lam in systemic and local anaphylaxis, Journal of
Immunotoxicology,2007, vol 4, no 4, Pages 287-294.
14. Haranath PSRK, Shyamalakumari S. Experimental study on the mode of action
Of Tylophora asthmatica in bronchial asthma. Indian J. Med. Res. 63, 1975; 661670.
15. Patil SD, Ahale SV, Surana SJ. Evaluation of antiasthmatic and antianaphylactic activity
of balanites aegyptiaca (delile), (balanitaceae). Asian journal of pharmaceutical and
clinical research. 2011; 4(1); 52-55.
16. Bennett RN, Mellon FA, Foidl N, Pratt JH, Du pont MS, Perkins L and
Kroon.PA. Profiling glucosinolates and phenolics in vegetative and reproductive
tissues of the multipurpose trees Moringa oleifera L. and Moringa stenopetala L,
J Agric Food Chem., 2003; 51(12), 3546-3553.
17. Ghosh MN, Some standard techniques In, Fundamentals of Experimental Pharmacology,
3rd Ed, Kolkata, Hilton and Company; 2005, Page 17.
SIGNATURE OF
CANDIDATE
REMARKS OF GUIDE
NAME AND DESIGNATION
OF (In Block Letters)
11.1 Guide
All the facilities for this study are available in
our department and this work can be
undertaken.
Dr. GEETHA M MD
PROFESSOR
DEPARTMENT OF PHARMACOLOGY
J.J.M. MEDICAL COLLEGE
DAVANGERE – 577004.
11.2 Signature
11.3 Co-Guide (If any)
11.4 Signature
11.5 Head of the Department
11.6 Signature
12.1 REMARKS OF THE
CHAIRMAN AND
PRINCIPAL
12.2 Signature.
Dr. SOMASHEKAR.H.S. M.D.
PROFESSOR AND H.O.D.
DEPARTMENT OF PHARMACOLOGY
J.J.M. MEDICAL COLLEGE
DAVANGERE – 577004.