Download annexure ii - Rajiv Gandhi University of Health Sciences

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES,
BANGALORE, KARNATAKA,
ANNEXURE II
PROFORMA FOR REGISTRATION OF SUBJECTS FOR
DISSERTATION
1.
Name of the Candidate
and Address
PANKTI KALARIA
a. Permanent Address
H-6, Janakpuri app.
Zanzarada Road,
Junagadh-362 001,
Gujarat.
b. Postal Address
Shree Devi College of Pharmacy,
Airport Road, Kenjar Village,
Malavoor Panchayat,
Mangalore-574 142
Karnataka.
2.
Name of the Institute
Shree Devi College of pharmacy,
Airport Road, Kenjar Village,
Malavoor Panchayat,
Mangalore-574 142
Karnataka.
3.
Course of Study and Subject
Master of Pharmacy in Pharmacology
4.
Date of Admission to Course
July 2010
5.
Title of the Topic:
“EVALUATION OF REJUVANATIVE
SCHOLARIS LINN LEAVES”
ACTIVITY OF ALSTONIA
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BRIEF RESUME OF THE INTENDED WORK:
6.
6.1 Need of study:
Immunology is the study of immunity which is the ability of body to resist
infections. It provides us with the specific mechanism to resist against a particular
pathogen or foreign substance involving production of antibodies or activation of
T-cells. Moreover, it can discriminate between self and foreign antigens and if
this ability is lost, it leads to autoimmune diseases [1].
Immunomodulators are the diverse range of metabolites which keep the body in
homeostasis condition. These stimulate, suppress or modulate the components of
immune system
[2]
. In the recent past, there has been growing interest on
exploiting the biological activities of different Ayurvedic medicinal herbs, owing
to their natural origin, cost effectiveness and lesser side effects [3]. During the last
couple of years the use of herbal products have shown promising responses to
modulate immune system compared to
modern medicines
[4]
. Some of the
traditional herbs which magnify effect of immune system include Allium
sativum, Ocimum sanctum, Aegle marmelos etc[5].
Alstonia scholaris Linn R.Br.commonly known as Devil’s tree belongs to the
family Apocynaceae. It grows throughout India, in deciduous and evergreen
forests, also in plains. Alstonia scholaris has been mentioned in Ayurveda and is
known as saptaparna. Scientific studies have established the multifarious utility
of this plant in wide array of pharmacological activities including antimicrobial,
antipyretic, antidiarrhoeal, anticancer, anthelmentic and antifertility [6]. It is useful
in malarial fevers, abdominal disorder, leprosy, asthma, skin diseases etc. It is
also used as an analgesic in case of rheumatic pain and ear ache. The bark extract
of Alstonia scholaris is reported to produce immunostimulating effect
[7]
.
However there is no report available on immunomodulatory activity of plant leaf
[6]
. Therefore present study is designed to examine the rejuvenative activity of
Alstonia scholaris Linn R.Br. leaves extract in different experimental models of
cellular and humoral immunity in animals.
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6.2 Review of literature:
Alstonia scholaris Linn belongs to family Apocyanaceae commonly known as
Devil’s tree. Alstonia scholaris is significantly known for its medicinal value
which grows up to 3 meters in height, distributed throughout the sub-Himalayan
belt, west-Bengal, Bihar, peninsular India and southwest Asia. The bark, stem,
roots and leaves have been used traditionally as folk remedies for the treatment of
many diseases [8]. The plant is known to contain alkaloids such as ditamine and
echitamine, flavonoids and phenolic acids
[6]
. The alkaloid fraction of leaves
contains three main alkaloids picrinine, vallesamine and scholaricine [9].
The plant is already reported for anthelmintic, digestive, febrifuge, antipyretic,
laxative, galactogogue, stomatic, cardiotonic properties. It is useful in fevers,
abdominal disorders, dyspepsia, pruritis, leprosy, asthma, bronchitis and debility.
Milky juice is applied on wounds, ulcers and rheumatic pain; mixed with oil and
dropped into ear it relieves ear ache. Drug is also used in case of snake bite[7].
Potent alpha glycosidase inhibitory activity was found in plant extract of Alstonia
scholaris that contributes towards the treatment of diabetes [10].
It has been reported to be used in management of hypertension [11]. Methanolic
extract of plant was found to exhibit pronounced antiplasmodial activity. The
plant is reported to have antimutagenic effect. The bark extract of Alstonia
scholaris has immunostimulating effect [7]. Alstonia scholaris was documented as
an effective herb for treatment of chronic respiratory disease and its leaf crude
extract is used for relieving tracheitis and cold symptoms [12].
6.3 Objective of study:
The objective of the proposed study is to investigate the effect of the leaf extract
of Alstonia scholaris Linn (LEAS) on different experimental models of cellular
and humoral immunity in animals.
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SPECIFIC OBJECTIVES:

To carryout extraction of leaf extract of Alstonia scholaris (LEAS) using
suitable solvent and extraction procedure.

To determine the phytochemical constituents of LEAS.

To arrive at the therapeutic dose range after acute toxicity studies
following WHO guidelines.

To explore the influence of LEAS on the following models of immunity
evaluation in animals:
1. Mice lethality test
2. Indirect haemagglutination test
3. Cyclophosphamide induced neutropenia
4. Serum immunoglobulins
7.
MATERIALS AND METHODS:
7.1 Source of Data:
Data will be obtained from laboratory based studies by using albino rats or albino
mice of either sex weighing between 150-200 gms and 25-30 gms respectively,
maintained at room temperature, having free access to food (std pellet diet), tap
water ad libitum. These studies will be carried out using different models of
immunity such as mice lethality test, indirect haemagglutination test,
cyclophosphamide induced neutropenia, serum immunoglobulins.
7.2 Method of Collection of Data:
Chemicals and reagents will be procured from standard companies. Extraction of
Alstonia scholaris leaf extract (LEAS) using suitable solvent and extraction
procedure will be carried out and extract will be subjected to different methods of
evaluation of immunity as discussed in objectives.
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EXPERIMENTAL MODELS:
1. Acute toxicity studies:
.
The dose will be selected by the acute toxicity studies following limit test
procedure according to OPPTS guidelines12. Briefly, a test dose of 2 g/kg and 5 g/kg
will be given to the mice. Around 1/10th and 1/20th of the maximum safe dose (either
2 g/kg or 5 g/kg) will be used as low and high dose respectively.
2.Mice lethality test1[3]:
Albino mice of either sex will be treated with different doses of LEAS for 21
days. On 7th and 17th day, they will be vaccinated with formalin
inactivated potash alum adsorbed pasteurella vaccine and at the end of treatment;
all animals will be challenged with broth culture of 25LD50 of duck pasturella
organism containing 335 CFU.
Following animals groups will be used for the study (n=6):
Group
I:
No vaccination, no treatment.
Group II: Vaccination on 7th and 17th day with 0.2 ml of formalin
inactivated potash alum adsorbed pasteurella vaccine (Vaccine).
Group III: Standard (Ocimum sanctum 100 mg/kg, p.o for 21 days) + vaccination
on 7th and 17th day.
Group IV: Low dose of LEAS orally for 21 days + vaccination on 7th and 17th
day.
Group V: High dose of LEAS orally for 21 days + vaccination on 7th and 17th
day.
On 21st day, the animals will be challenged with broth culture of 25 LD50 of duck
pasturella organism containing 335 CFU in 0.2ml of nutrient broth
approximating to 1017 cells. The animals will be observed for a period of 72 hr
and the mortality ratio will be determined using the formula.
Mortality ratio: Number of animals dead
Total number of animals
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3. Indirect heamagglutination test[14]:
Rats will be pretreated with the drugs for 14 days and each rat will be immunized
with 0.5x109 sheep red blood cells (SRBCs) intraperitoneally, including control
rats. The day of immunization will be referred to as day 0. The drug treatment
will be continued for another 14 more days and blood samples will be collected
from each rat at the end of the drug treatment and the titre value will be
determined by titrating serum dilutions with SRBC (0.025x109 cells) in microtitre
plates. The plates will be incubated at room temperature for 2 hr and examined
visually
for
agglutination.
The
highest
dilution
of
serum
showing
haemagglutination will be expressed as haemagglutinating titre.
Animal groups will be made as follows (n=6):
Group I: Control, no treatment.
Group II: Standard (Ocimum sanctum 100 mg/kg, p.o for 28 days).
Group III: Low dose of LEAS orally for 28 days
Group IV: High dose of LEAS orally for 28 days.
4.Cyclophosphamide induced neutropenia [15]:
On 10th day of LEAS/standard administration, single dose of cyclophosphamide
200 mg/kg s.c will be given. The TLC and DLC will be determined before and 3
days after cyclophosphamide administration. Albino mice of either sex will be
divided into four groups as given in Indirect heamagglutination test.
5. Serum immunoglobulin(Ig) levels[16]:
Albino rats of either sex weighing between 150-200 gm will be treated with
LEAS/standard for 21 days. Six hours after the last dose of drug, blood will
collected and the serum will be used for estimation of immunoglobulin levels
spectrophotometrically.
Animals are divided into four groups of six each as
discussed in Indirect heamagglutination test.
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7.3 Does the study require any investigation or interventions to be conducted
on patients or the human or animals? If so please describe briefly:
YES
Study requires investigation on animals. The effects of the drug will be
studied on various parameters using rats and mice as experimental animals.
7.4 Has ethical clearance been obtained from your institute
Ethical Committee approval letter is enclosed
8.
List of References:
1. Sharma HL, Sharma KK. Immunomodulation and immunotherapy,
Principles of Pharmacology. Paras Publication, 2007;1:902-920.
2. Smith A, O’Byrne A, Brandt BV, Bianchi I. Introduction to
Bioregulatory medicine, Thieme Publication 2009:22.
3. Chopra RN, Nayal SL, Chopra IC. Glossary of Indian Medicinal Plants,
CSIR, New Delhi, 2009:185-210.
4. http:// www.banyanbotanicals.com, retrieved on 3/12/2010, 11:20 pm.
5. Dr.Ansari SH. Essential of Pharmacognosy. Birla publication 2004:705708.
6. Kulkarni MP, Juvekar AR. Effect of Alstonia scholaris Linn on stress and
cognition in mice. Indian J Exp Biol, 2009;47:47-52.
7. Arulmozhi S, Mazumder PM, Narayan LS, Thakurdesai PA. Invitro
antioxidant and free radical scavenging activity of fractions from
Alstonia scholaris Linn R. Br. International Journal of Pharm Tech
Research 2010;2:18-25.
8. Thomas PS, Kanarijia A, Ghosh D, Duggar R, Katiyar CK. Alstonoside,
a secoiridoid glucoside from Alstonia scholaris. Indian J Chem 2008;
47:1298-1302.
9. Shanq JH, Caj XH, Feng T, Luo XD. Pharmacological evaluation of
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Alstonia scholaris Linn R.Br.: antiinflammatory and analgesic activity.J
Ethanopharmacol 2010;129(2):174-181.
10. Anurakkun NJ, Bhandari MR, Kawabata J. Alpha-glucosidase inhibitors
from Devil’s tree (Alstonia scholaris). Food chem 2007;103:1319-1323.
11. Bhogayata K, Sharma PP, Patel BR. A clinical evaluation of saptaparna
(Alstonia scholaris Linn R.Br.). Ayu 2009;30:318-322.
12. Shanq JH, Caj XH, Feng T, Luo XD. Pharmacological evaluation of
Alstonia scholaris Linn R.Br.: antitussive, antiasthmatic and expectorant
activities. J Ethnopharmacol 2010,129(3):293-298.
13. Ramanatha KR, Lakshmanarayana R, Gopal T. Potency test of Duck
pasturella vaccine in mice. Mysore J Agri Scie 1995;29:155-157.
14. Takuo S, Richard BR, Keith RR. Indirect Hemagglutination test that uses
glutardehyde-fixed Sheep Erythrocytes sensitized with extracts, antigens
for detection of pasturella antibody. J Clin Microbial 1982;15(5):752756.
15. Heppner GH, Calabresi P. Selective suppression of humoral immunity by
antineoplastic drugs. Annu Rev Pharmacol Toxicol 1976;16:367-379.
16. Mullen PA. Zinc sulphate turbidity test as an aid to diagnosis. Veter Ann
1975;15:451-455.
9.
SIGNATURE OF THE CANDIDATE:
10.
REMARKS OF THE GUIDE:
“EVALUATION OF REJUVENATIVE ACTIVITY OF ALSTONIA
SCHOLARIS LINN LEAVES EXTRACT IN ANIMALS” to be carried out by
Miss Pankti Kalaria of M. Pharm has been discussed and worked out under my
directions and supervision as an official guide. The project work envisaged is of
great importance in the field of herbal research. The work can be carried out in
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pharmacology laboratory of Shree Devi College of Pharmacy for which facilities
are available. Hence the project is viable and is recommended for clearance and
approval.
11.
Name and Designation of
11.1 Guide
Mr. Talha Jawad
Asst. Professor,
Dept. of Pharmacology
Shree Devi College of Pharmacy,
Mangalore-574 142
11.2 Signature
11.3 Head of the Department
Dr. Jagadish V. Kamath
Dept. of Pharmacology
Shree Devi College of Pharmacy,
Mangalore-574 142
11.4 Signature
12.
12.1 Remarks of the Principal
The Programme and the Research work that is undertaking by Miss Pankti
Kalaria have potential implication in the field of Pharmacology. The work can be
carried in the Research Laboratories of Pharmacology Department at Shree Devi
college of Pharmacy. Hence the Project is recommended and requested for
clearance and approval.
12.2 Signature
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