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1. CARDIOPROTECTIVE ROLE OF TERMINALIA CATAPPA.
Linn AND CISSAMPELOS PAREIRA. Linn ON DOXORUBICIN
INDUCED CARDIAC TOXOCTIY IN RATS
THESIS
Submitted to
Jawaharlal Nehru Technological University, Anantapur,
In the partial fulfillment of the requirements for
the award of the degree of
MASTER OF PHARMACY
IN
PHARMACOLOGY
By
Y.LALITHA
[Reg.No.10AF1SO108]
Under the guidance of
V. JAYA SANKAR REDDY
M.Pharm, (Ph.D)
Department of Pharmacology
KRISHNATEJA PHARMACY COLLEGE, TIRUPATHI, CHITTOOR (Dt)
SEPTEMBER -2012

2.  AIM AND OBJECTIVE OF THE STUDY
 The Goal of the present study is to investigate
• Cardiac protective activity of Terminalia catappa.L and Cissampelos
pareira.L against doxorubicin induced cardiac toxicity in rats.
 The following objectives were set to achieve the goal of the present study
• To perform the acute toxicity studies of ethanolic extracts of Terminalia
catappa.L and Cissampelos pareira.L on mice.
• To investigate the effect of various doses of Terminalia catappa.L and
Cissampelos pareira.L on serum biomarkers (CK, LDH, SGOT, SGPT) in
normal and cardio toxic rats.
• To investigate the effect of various doses of Terminalia catappa.L and
Cissampelos pareira.L on antioxidant enzymes (SOD and catalase) in
normal and cardio toxic rats.

3. PLAN OF WORK
Plant selection and acquisition
Botanical identification
Drying of plant materials
Extraction of plant materials
Phytochemical profiling
Acute toxicological studies
Screening of cardio protective activity

4. INTRODUCTION
• Cardiovascular diseases remain the principal cause of death in
both developed and the developing countries.
• Myocardial infarction is a prominent implication of the
cardiovascular disease. According to WHO 16.7 million
people die each year owing to heart attacks.

5. • Traditionally, Terminalia catappa.Linn and Cissampelos
pareira.Linn were used as cardio-tonics and pharmacologically
both shown significant antioxidant property, hence these plants
were chosen to evaluate the Cardio protective Activity.
• Doxorubicin, a successful antitumor drug also known for its
cumulative and dose dependent cardio toxicity. Hence Dox
has been chosen for the study to induce cardio toxicity.

6. MYOCARDIAL INFARCTION
• Myocardial infarction is an ischemic necrosis of a portion of the
myocardium due to sudden occlusion of a branch of coronary artery.
 CLASSIFICATION:
. ST elevation MI(STEMI)
. Non- ST elevation (non-STEMI)

7.  EPIDEMOLOGY:
• MI is a common occurrence in young man(40-60yrs) and in
women (60-85yrs).
• World wide more than three million people were suffering
with STEMI and more than four million people were suffering
with Non-STEMI.
• Blacks and whites are equally affected.
• Mortality estimates due to CVD vary widely by state, ranging
from 10% in Meghalaya to 49% in Punjab.

8.  AEITOLOGY:  SYMPTOMS:
• Hypercholesterolemia
• Pain
• Diabetes mellitus
• Difficulty in breathing
• Smoking
• Dizziness
• Stress
• Obesity • Nausea
• Insufficient exercise • Shortness of breath
• Hypertension • Mimics like MI

9.  Pathophysiology:
Role of plaque in MI

10.  DIAGNOSIS OF MYOCARDIAL INFARCTION
• The diagnosis of myocardial infarction can be made after
assessing patient's complaints and physical status.
• ECG changes, Coronary angiogram and levels of cardiac
markers help to confirm the level of infarction.

11.  MYOCARDIAL INFARCTION COMPLICATIONS
• Arrhythmias
• Heart failure
• Pericarditis
• Emboli

12. PLANT PROFILE
Terminalia catappa.L Cissampelos pareira.L

13.  TAXONOMY:  TAXONOMY:
Kingdom :Plantae Kingdom : Plantae
Division :Magnoliophyta Division : Magnoliophyta
Class :Magnoliopsida Class : Magnoliopsida
Order :Myrtales Family : Menispermaceae
Family :Combrataceae Genus : Cissampelos
Genus : Terminalia Species : Pareira
Species :catappa

14. Uses
Terminlia catappa.Linn Cissampelos pareira. Linn
 It has been used for dysentry,  It has been used for heart
rheumatoid arthritis, cough, problems,asthma,inflammation
asthma, leprosy, nausea, eye
, malaria, STD, snake bite and
problems, liver problems and
Kidney problems.
STD.
 It has been used as diuretic,
 It is used as cardiotonic, diuretic,
antiseptic and anti periodic
antipyretic, aphrodisiac and
antimicrobial agent.
property.

15. REVIEW OF LITERATURE
Terminlia catappa. l
S.NO Authors & year Activity done
1 B.V.Jawal et al.,(2012) Antioxidant potential
2 Abulmanzur et al.,(2011) Antimicrobial activity
3 A. Aimola et al., (2011) Erythropoiesis
4 Azrul et al.,(2011) Anthelmintic potential
5 Paul chidokachikezie (2011) Antipolymerization
6 Saheb L and SS Wadje.,(2011) Fungicide activity
7 Saroja M et al.,(2011) Antioxidant activity of phenolic fraction
8 H.V Annegowda et al.,(2010) Analgesic and antioxidant properties
9 Jing gao et al.,(2010) Hepatoprotective activity
Kinoshita S et al.,(2007)
10 Minakshi G Joshi et al., (2008) Antibiotic
11 Ahmed SA et al.,(2006) Antidiabetic activity
12 Fan YM et al., (2004) Phytochemical and anti-inflammatory
activity

16. Cissampelos pareira.L
S.NO Authors & year Activity done
1 Singh BK et al., (2012) Cardiac protective activity
2 Mahendra sigh Yadv Hepatoprotective activity
etal.,(2011)
3 PadminiShukla et al., (2011) In-vitro anthelmintic activity
4 Pramodine d et al., (2011) Memory enhancing activity
5 Surendran S et al., (2011) Hepatoprotective activity
6 Gupat M et al.,(2010) Antipyretic activity
7 Bafna A, Mishra S.,(2009) Immunomodulatory activity
8 Amresh G et al., (2008) Toxicological screening
9 Amresh G et al., (2007) Antinociceptive and antiarthritic
10 G. Amresh et al., (2011) Gastroprotective
11 AmreshG,Reddy GD et al., Antioxidant activity
(2007)

17. • MATERIALS:
Animals: Wistar rats(150-200gms)
Swiss albino mice(20-30gms)
S.NO Chemical Manufacture
1 Doxorubicin Celon
2 Normal saline Claris life sciences Ltd
3 CK-MB kit span diagnostics Ltd
4 SGOT kits span diagnostics Ltd
5 SGPT kits span diagnostics Ltd
6 Formalin Sd-fine chem. Limited
7 Potassium dihydrogen phosphate Merk specialities
8 Ethanol Changshuyangyuan chemicals
9 Disodium hydrogen phosphate Fisher scientific

18. • METHODS
 Identification and authentification of plants
 Collection of plant materials
 preparation of plant materials for the extraction
 Extraction procedure
 Preliminary phytochemical studies

19. ACUTE TOXICITY STUDIES
 The procedure was followed by using OECD 423 (Acute Toxic
Class Method).
 The starting dose level of Terminalia catappa.L and
Cissampelospareira.L was 2000 and 5000mg/kg body weight p.o.
 Dose was administered to overnight fasted mice`s. Food was
withheld for and further 3-4 hours after administration.
 The body weight of the mice`s before and after administration were
noted.
 the changes in the body weight were not so prominent.
 The onset and signs of toxicity was also not observed. No mortality
was observed.

20. PHARMACOLOGICALSCREENING
 GROUPING:
• Group I : Control (Normal control)
• Group II : Positive control(doxorubicin(15 mg/kg B.W, through
I.P))
• Group III: Test I (Plant Extract-250mg/kg B.W of EETC+DOX)
• Group IV : Test II (Plant Extract-500mg/kg B.W of EETC+DOX)
• Group V : Test III(plant extract-250mg/kg B.W of EECP+DOX)
• Group VI : Test IV (Plant extract-500mg/kg BW of EECP+DOX)

21. • Group VII: Test V(DOX+Plant Extract-250mg/kg B.W of
EETC)
• Group VIII: Test VI(DOX+Plant Extract-500mg/kg B.W of
EETC)
• Group IX: Test VII(DOX+ Plant extract-250mg/kg B.W of
EECP)
• Group X: Test VIII(DOX+ Plant extract-500mg/kg B.W of
EECP)

22. • Rats were divided into ten groups each group contain six animals.
• Group I serves as normal which receives normal saline(5ml/kg B.W)
for 30 days and Group II serves as positive control which receives
doxorubicin 7.5mg/kg B.W in two equal injections at 27thand 29th
day(15mg/kg B.W; I.P route)and normal saline(1ml/kg B.W).
• Group III, Group IV, Group V and Group VI serves as test which
receives plant extracts of Terminalia catappa.L and Cissampelos
pareira.L at dose of 250 and 500 mg/kg B.W, through oral route
throughout the study period and also receives doxorubicin (15 mg /kg
B.W) at 27thand 29thday.

23. • Group VII, Group VIII, Group IX and Group X also serves as test
and receives doxorubicin(15 mg/kg B.W) first and then receives test
extracts at an dose of 250 and 500mg/kg B.W, through oral route
throughout the study period.
• After the study period animals were sacrificed by collecting blood
through cardiac puncture and collected blood was centrifuged and
serum was collected and biochemical analysis were performed for
serum levels CK-MB, LDH, SGOT, SGPT, SOD and Catalase.
• The hearts were isolated and placed in 10% formalin and
histopathological studies were performed.

24. RESULTS
 Phytochemical analysis:
S.NO COMPOUNDS ETHANOLIC EXTRACT OF ETAHANOLIC EXTRACT OF
TERMINALIA CATAPPA CISSEMPELOS PAREIRA
1 Carbohydrates + +
2 Fixed oils + +
3 Alkaloids + +
4 Glycosides + +
5 Steroids + +
6 Flavonoids + +
7 Tannins + +
8 Phenolic compounds + +
9 Amino acids - -
10 Saponins + +
+VE: Presence of compound
-VE : Absence of compound

25.  Acute toxicological study:
• From acute toxicity studies their was no mortality at a dose of
5000mg/kg B.W.

26.  Pharmacological screening:
Effect of ethanolic extracts of Terminalia catappa. L and
Cissampelos pareira. L on serum biomarkers in normal and
doxorubic in exposed rats
S.NO Groups CK-MB LDH SGOT SGPT
1 Group I 126.20±2.584 116.79±2.412 68.6±4.188 46.213±0.7026
2 Group II 246.52±6.609 234.76±4.889 161.98±0.7525 92.82±1.898
3 Group III 181.00±2.369*** 143.22±4.770*** 140.23±1.074*** 81.35±0.7794***
4 Group IV 152.80±7.061* 136.65±3.484*** 116.68±1.145*** 68.68±0.8520***
5 Group V 171.49±2.513*** 137.22±4.019*** 130.80+4.414*** 73.67±1.433***
6 Group VI 156.10±2.768** 120.49±5.220*** 87.53±6.7332*** 57.23±0.9472***
7 Group VII 223.28±8.677*** 198.25±3.146*** 178.51±2.785** 78.32±0.8053***
8 Group VIII 192.84±5.893*** 178.52±2.276*** 146.14±1.837* 69.21±1.686***
9 Group IX 178.12±3.503*** 147.22±2.742*** 126.61±4.753*** 68.88±1.879***
10 Group X 149.70±2.876* 133.86±2.883*** 88.248±3.664*** 51.84±1.501***

27. Effect of ethanolic extracts of EETC and EECP on antioxidant levels in doxorubicin treated rats hearts
S.NO GROUPS SOD CATALASE
1 Group I 7.1±0.03890 11.24±0.1472
2 Group II 3.747±0.2449 5.07±0.1451
3 Group III 5.168±0.08392*** 7.75±0.2808***
4 Group IV 5.597±0.2800*** 9.12±0.1183***
5 Group V 4.980±0.2493** 8.30±0.2516***
6 Group VI 6.112±0.1657*** 10.250±0.07638***
7 Group VII 4.942±0.1241** 6.74±0.1700***
8 Group VIII 4.92±0.2554** 7.64±0.1148***
9 Group IX 4.97±0.1179** 8.54±0.2683***
10 Group X 5.900±0.3246*** 10.81±0.2629***
All values were expressed as mean+ SEM; n= 6 observation, values were significant when compared
with positive control, * P<0.05,** P<0.01,*** P<0.001(one way ANOVA followed by Turkey`s test)

28. Effect of Terminalia catappa.L and Cissampelos pareira.L on serum CK,
LDH, SGOT & SGPT levels in doxorubicin induced cardiac toxic rat

29. Histopathlogical studies

34. CONCLUSION
• The present study suggest that the Terminalia catappa.L and
Cissampelos pareira.L could be used as an antioxidant during or
after doxorubicin therapy plants shows a greater protective effect in
preventive as compared with curative model in rats.
• Further studies should be required for the detection, isolation and
structural elucidation of the chemical constituents and further
investigation should be required to find exact mechanism of action
for cardioprotective activity of Terminalia catappa.L and
Cissampelos pareira.L plants.