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College of Veterinary Medicine

BUDDHA BELLY LEAF (Jatropha podagrica)

A Thesis Presented to the Faculty of the

College of Veterinary Medicine
Cagayan State University
Tuguegarao City


In Partial Fulfillment
Of the Requirements for the Degree


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College of Veterinary Medicine








School year:




This study was conducted to evaluate a wound healing rate of Buddha belly leaf
(Jatropha podagrica) ointment on incised wound of albino rats.
30 albino rats were randomly distributed into five treatment groups with 6
replicate each group. The Buddha belly leaf extract (1ml, 1.5ml and 2ml) was
incorporated into 10 g of a simple ointment base by melting and trituration to produce
three batches of the ointment formulation. Incision wound measuring 1 inch (2.5 cm) was
created on the dorsal pelvic region of the albino rats and the ointment was applied
topically twice a day on the wound which was measured at intervals of 3 days until
epithelialization or complete wound closure. The other group was treated with Bactroban
ointment (2%) which serve as the control and standard treatments, respectively. And the
last group were remained untreated and heals naturally.
Topical application of the leaf extract of Jatropha podagrica incorporated into an
ointment base on the incision wound in rats caused a significantly (P<0.05) higher rate of
wound healing and reduced the epithelialization period in a doserelated manner. The
result shows that the ointment formulated from extracts of Jatropha podagrica containing
the highest concentration of the sample extract (2ml/10g ointment) showed the highest
rate of wound closure when compared to the no applied ointment treated group as well as
the formulated ointment containing the lower concentrations (1ml and 1.5ml/10g of
ointment base).

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Chapter I

Topical antimicrobial therapy is one of the most important methods of wound
care. Some medicinal plants have been employed in folk medicine for wound. Some of
these plants either possess pro-wound healing activities or exhibit antimicrobial and other
related properties which are beneficial in overall wound care. Wound healing processes
are well organized biochemical and cellular events leading to the growth and regeneration
of wounded tissue in a special manner. Healing of wounds is an important biological
process involving tissue repairs and regeneration. It involves the activity of an intricate
network of blood cells, cytokines, and growth factors which ultimately leads to the
restoration to normal condition of the injured skin or tissue (Clark, 1991).
Since the earliest times, people rely on herbs for treatment of various deseases.
Herbs became an affective source of human health due to the active compounds present
within rendering them many pharmacological activities. Treatment may not heal as fast as
commercial drugs that we have now but they proved to be cheap and efficient (Longid
2014). One of the frequently used herbs is the Buddha belly plant (Jatropha podagrica).
The genus name Jatropha derives from the Greek word jatros (doctor/healer) and
trophe (food), which implies medicinal uses and the species podagrica (podagrikos) as a
liable to gout. Jatropha podagrica Hook (various names: Buddha belly, Gout plant, Coral
nut, Guatemala rhubarb, physic nut; Family Euphorbiaceae) is a shrub native to tropical

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America, but it is also found in Australia, the Hawaiian Islands, Southern Africa,
Mozambique, Zambia and warmer parts of Asia.
Jatropha podagrica is known locally in south western Nigeria as Lapalapa
funfun. Where they are used in folk medicine to treat various diseases including skin
infections, sexually transmitted diseases, jaundice and fever. Moreover, various medicinal






molluscidal and insect antifeedant have been attributed to this plant. It is also used as an
antipyretic, diuretic, choleretic, purgative, reduction of swelling, pain relief and to
detoxify snakebites and treatment of scabies.
Those qualities claimed present in Buddha belly are the reason for the researcher
to investigate a methanol extract of Jatropha podagrica (Euphorbiaceae) which was
formulated in an ointment base for pro-wound healing activity on incision wound model.

Objectives of the study

Generally, this study aims to determine the wound healing rate of a Buddha belly
To compare the wound healing rate of the different concentrations (1ml, 1.5ml
and 2ml) of Buddha belly leaf ointment with the standard drug Bactroban
ointment in incisional wound healing.

Significance Of The Study

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1) Owners/Farmers: this study will help the owners in supplementing their
knowledge concerning how to treat problems of clean cut skin/incised wound
using herbal preparation.
2) Community: to use as alternative, affordable and practical in treatments of
wounds on ointment base preparation.
3) Future researchers: this study will serve as added reference and which shall
contribute to the growing literature on drug discovery as preliminary study of the
Buddha belly leaf extract as wound healer.
This study was came up with a natural wound healer solution product.

Scope And Delimitation Of The Study

This study focuses only on the wound healing rate of Jatropha podagrica leaf
extract application on albino rats. In this manner, it is cheaper compare to commercial

Time and Place of the Study

The Buddha belly leaves were gathered at Sacpil,Conner, Apayao. The extraction
process and the experimental part was performed and conducted at Cagayan Valley
Herbal Processing Plant from August to October 2015.

Definition of terms:
Antimicrobial: inhibiting the growth of microorganisms.

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Disease: an illness that affects a person, animals, or plants that prevents the body from
working normally.
Epithelialization: healing by the growth of epithelium over a denuded surface.
Excision: the act of cutting out.
Extract: to remove something by pulling it out or cutting it out.
Herb: a plant or part of a plant that is used as medicine.
Ointment: a smooth substance that is rubbed on the skin to help heal a wound or to
reduce pain or discomfort.
Regeneration: the process of regenerating.
Restoration: the process of returning something into its original condition or by repairing
Wound: an injury that is caused when a knife, bullet, etc., cuts or breaks the skin.

Chapter II

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Buddha belly (Jatropha podagrica)

Origin and Description
The genus name Jatropha derives from the Greek word jatros (doctor/healer) and
trophe (food), which implies medicinal uses and the species podagrica (podagrikos) as a
liable to gout (Kumar and Sharma, 2008).
Jatropha podagrica Hook (various names: Buddha belly, Gout plant, Coral nut,
Guatemala rhubarb, physic nut; Family Euphorbiaceae) is a shrub native to tropical
America, but it is also found in Australia, the Hawaiian Islands, Southern Africa,
Mozambique, Zambia and warmer parts of Asia.
Jatropha podagrica is one of the approximately 175 species of succulent plants of
the genus, Jatropha. A popular ornamental plant that typically grows between 0.5-1
meters in height, taller when ground-planted in garden than being potted. Exotic, yet
weird looking in a captivating way, with a stout and knobby stem that develops from a
swollen gout-like, bottle-like, or belly-like base or caudex, hence attributing to a few of
the plants common names. The trunk is greyish in colour with bristled scars. Foliage is
medium to dark green, sometimes slightly two-toned, with large peltate leaves, up to 30
cm across. They are orbicular with 3-5 distinct and deep lobes, bluntly tipped and held on
long petioles that fans out from the top tip of the succulent stem, forming a luxuriant

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green crown. As showy as the fabulous foliage are its clusters of small and brightly
coloured orange-red flowers pedicels. The inflorescence is cymose and coral-like in
manner, held terminally on along and slender peduncle that emerges from terminal stem
as with the leaf petioles. Jatropha podagrica flowers all year round in favourable
environment, especially in the warmer months of the year. Roundish seed pods or fruits
are set freely after flowering, start off a lovely fresh green and turn blakish brown when
matured. When ripened, they burst open to scatter smooth and glossy brown seeds several
meters away. Seeds will self-sow easily on suitable soil.
The genus Jatropha is one of the important sources for biologically active
phytochemicals. The important biological activities and toxicities of several purified
compounds from Jatropha species have been well established (Nowshin N. Rumzhum et
al. 2012).
Jatropha species are used in traditional medicine for various diseases including
skin infections, sexually transmitted diseases such as gonorrhoea, jaundice and fever.
Different parts of the plant are also used for antipyretic, diuretic, choleretic and purgative
effects. Various medicinal and pesticidal properties, including antimicrobial, antitumour
and insect antifeedant activities, have also been attributed to this plant (Hossain Sohrab et
al. 2012).
It was also mentioned at Wiley InterScience that the roots have shown
antimicrobial, antibacterial and antifungal activity.

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In previous phytochemical investigations, a methanol crude extract of a stem bark
of J. podagrica was isolated japodic acid, erytrinasinate, n-hexacosane, -amyrin, lupeol
palmitate, quercetin, apigenin, vitexin, isovitexin, rutin, tetramethylpyrazine, podacycline
A, podacycline B and 3-acetylaleuritolic acid. Another study also reported the isolation of
six compounds fraxidin, fraxetin, scoparone, 3acetylaleuritolic acid -sitosterol and
sitosterone from J. podagrica. In addition, different parts of the plant have been
investigated chemically and many compounds including flavonoids, steroids, alkaloids
and diterpenoids have been isolated from this plant that yields with antimicrobial, anticancerous, nematicidal, antibiotic, anti-inflammatory etc. (Muhammad Abdullah AlMansur et al. 2012).

Traditional Use:
Traditionally this plant is considered as being sweet, bitter and cold. It has been
used as analgesic, antipyretic, ant-inflammatory and detoxicant (Globinmed).
In Indonesia the whole plant is used to treat fever. The Africans however make
use of the seed oil as part of the ingredients in a compound medicine to treat fever. It has
been used in the treatment of malaria. The whole plant is used as haemostatic, and also
applied on hematoma to aid in rapid reduction of swelling. The plant is known to have
diuretic properties and is used to treat haematuria. It is also used to treat gonorrhea. In
Indonesia and china, the plant has been used to treat snake bites 10-15g of the plant is
pounded and then immersed in rice wine (Globinmed).

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In Brazil, the seeds are used to expel intestinal worms, and the leaves are burnt to
fumigate houses against bed bugs. These plants are used in natural remedies, particularly
in homeopathic medicine. The latex contains an alkaloid Jatrophine, which is thought to
have anti-cancerous properties. It is also used as an external application for skin diseases
and rheumatism. A juice extracted from the leaves is used in some countries as an
external application for haemorrhoids. The roots are also considered to be an antidote for
snakebite (Julius T. Mwine and Patrick Van Damme, 2011).
Similarly, China used the said plant for reduction of swelling, pain relief and to
detoxify snakebites, while in Ghana and Nigeria, it is used as an antipyretic, diuretic,
choleretic and purgative. In Mexico and southwestern United States, it is used to tan
leather and produce a red dye.
In popular medicine the latex of Jatropha podagrica is used for the infected
wounds, skin infections and scabies (Kosasi et al. 1989; Kroes et al. 1996).


Antimicrobial activity of the stem and stem bark
According to study, the initial stages, the stem and stem bark extracts in three
different solvent viz, chloroform, methanol and hexane were evaluated for antimicrobial
activity against clinical isolates of S. aureus, E. coli and Candida albicans. Stem and
stem bark extract prepared screened for their capacities to inhibit the growth of these
clinical isolates that were quite resistant to antibiotics. Stem bark extracts of J. podagrica
show broad spectrum antimicrobial activity. The antifungal activity of the fluconazole

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and the stem bark were found to be nearly similar since the fluconazole is the drug of
choice for Candida albicans (Bhushan Bhaskarwar et al. 2008).
Antibacterial activity of the roots
Similarly, Aiyelaagbe (2000), his study of hexane, chloroform and methanol
extracts of the rootwood and rootbarks of Jatropha podagrica were proved for their
antimicrobial activity against 18 organisms. All the extracts exhibited some broad
spectrum antibacterial activity, at a concentration of 20 mg/mL. The hexane extracts were
generally more active than the chloroform and methanol extracts. The hexane extract of
the yellow rootbark was the most active of all the extracts and its activity was comparable
to that of gentamycin but better with regard to the control of S. aureus and B. cereus.
Furtheremore, three of the extracts, hexane extract of the yellow rootbark and
hexane and methanol extracts of the rootwood showed moderate antifungal activity
against the yeast fungus, Candida albicans.

Antifeedant activity of Jatropha podagrica roots

Another investigation done by Hassanali, Ahmed (1998). Hexane and MeOH
extracts of root bark of J. podagrica (collected from Nigeria) were screened for activity
against third instar larvae of Chilo partellus. Both extracts exhibited potent activity; the
hexane extract was the most active (87.6% feeding deterrence at a dose of 100 g/leaf

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However, Japodic acid, a novel aliphatic acid from J. podagrica showed mild
insect growth inhibition activity against Helicoverpa zea (37% growth reduction at 100
ppm). Fraxidin and erythrinasinate exhibited antibacterial activity against Bacillus
subtilis while japodic acid was inactive in the antibacterial assays conducted (Olapeju O.
Aiyelaagbe and James B. Gloer. 2008).
Neuromuscular and cardiovascular activity
A study believed that an amide alkaloid isolated from the stem of Jatropha
podagrica neuromuscular blocking and hypotensive effect. The neuromuscular effect was
found to be similar to those of d-tubocurarine i.e. blocking actions at the cholinergic and
adrenergic neuro-effector junctions, the neuro-muscular junction and at the ganglia. The
hypotensive effect is proven to be of a direct vascular nature and it was found that it
blocked the extracellular entry of calcium through calcium channel and also inhibit thr
release of intracellular stored calcium in the vascular smooth muscle. This indicates that
it is a true calcium antagonist. Data showed that tetramethylpyrazine from the Jatropha
podagrica extract was hypotensive and had a direct vascular effect, it not only blocked
the entry of extracellular calcium through calcium channels but also inhibited the released
of intracellular stored calcium in the vascular smooth muscle cell. It was a true calcium
antagonist (Ojewole and Odebiyi 1980).

Schistosomicidal Activity
An organization known as Global Information Hub on Integrated Medicine, also
said that Tetramethylpyrazine (TMP) isolated from the stem bark of Jatropha podagrica

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is both miracidial and cercariacidal but not ovicidal at a minimal concentrations of 100
ug/ml. sublethal concentrations (125-800 u/ml) of TMP arrested the development of
miracidia to cercariae in the bilharzia-transmitting snail (Bulinus globosus).

Cytotoxic activity
It was also shown that the fraxidin, acetylaleuritolic acid and g-sitosterol isolated
from roots and stem showed cytotoxic activities against HeLa cell lines with IC 50 of
39.9ug/L, 35.7ug/L and 15.9ug/L respectively (Globinmed).

A previous study from the University of Rouen, Mont St. Aignan entitled
Assessment of cyto-protective, antiproliferative and antioxidant potential of a
medicinal plant Jatropha podagrica that aimed to evaluate antiproliferative,
antioxidant activity and in vitro protective effect of hydro-alcoholic extract of Jatropha
podagrica. Total phenolics and flavonoids content was higher in seeds than leaves
extract. One of their new journalist obtained a quote from the research states that: The
anti-proliferative effect on two tumoral celllines A549 and PC12 was assessed by MTT
assay. The cytotoxicity of the studied extract was evaluated on young cerebellar granule
cells. The investigation of the action mode off. Podagrica extract in protection of DNA
damages, protein carbonylation and lipid peroxidation caused by hydroxyl radicals
generated in Fentons reaction exhibits an effective protection at low concentrations. In
the presence of 100 p,M of copper sulfate, Jatropha hydro-alcoholic extract showed no
significant enhancing in the hydroxyl radicals generation and DNA fragmentation in

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According to their news editors, the research concluded: These results suggest
that extract of J. podagrica still acts as antioxidant even in the presence of free metal
ions, but with less efficiency, the J. podagrica extract which showed significant antitumor
activity against the A549 and PC12 cells deserves further research into the
chemoprevention and treatment of cancer.

In anesthesize cats it produced depressor effects, reduced the heart rate and
blocked neuromuscular transmission and appeared to have spasmolytic activity on
smooth muscles (Ojewole, 1980). Further studies confirmed blockage of adrenergic and
cholinergic transmission by tetramethylpyrazine. The compound depressed and abolished
the electrically evoked contractions of the chick oesophagus, rabbit duodenum and guinea
pig vas deferens in vitro. It is also inhibited the electrically induced contraction of the rat
isolated hemi-diaphragm and of the cats nictitating membrane in vivo. Apart from its
possible central effects, and those on the cardiac muscle and blood vessels, it could be
suggested, from the results obtained in the study, that the hypotensive effect in
experimental animals is likely to be contributed to by, or mediated via, its local anesthetic
(membrane stabilizing) activity. Through this action, the drug probably acts to block
sympathetic and parasymphatetic neurons and ganglia (Ojewole, 1981).
Furthermore, tetramethylpyrazine has a number of other pharmacological actions.
A main central effect was found to be tranquillization and sedation (Ojewole and Odebiji,
In china, tetramethylpyrazine originating from plants is used in the treatment of
occlusive cerebral vessel diseases such as cerebral embolism (Xiao, 1983).

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Wound is a bodily injury caused by physical means, with disruption of the normal
continuity of structures. Wound Healing is the restoration of integrity to injured tissues by
replacement of dead tissue with viable tissue. The process starts immediately after an
injury and may continue for months or years, and is essentially the same for all types of
wounds. Variation in wound healing are the result of differences in location, severity of
the wound, and the extent of injury to the tissues. Other factors affecting wound healing
are the age, nutritional status and general state of health of the animal and its body
reserves and resources for the regeneration of tissue (Baillieres Comprehensive
Veterinary Dictionary, 1998).
Wound healing are conveniently classified into any of three types, healing by first
intention, healing by second intention and healing by third intention, depending on the
nature of the edges of the healed wounds. Whereas the edges of wounds healed by first
intention are smoothly closed that no scar is left, wounds healed by second intention
involve formation of granulation tissues which fill up the gaps between the wound edges
and are associated with significant loss of tissue, leaving little scars. Wounds healed by
third intention are usually those wounds left for three to five days until granulation bed
falls before they are sutured resulting in extensive scars formation (Thomas, 1997). Four
distinct stages of wound healing have also been identified- Inflammatory Phase in which
the bodys defenses are aimed at limiting the amount of damage and preventing further
injury. At the initial time of tissue disruption, platelets release coagulation factors and
cytokines to initiate the healing process. Within the first day following tissue injury,
neutrophils attach to surrounding vessel walls and then move through the vessel walls to

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migrate to the wound site. Debridement or Destruction Phase, Proliferation Phase is
characterized by movement of wound macrophages into the site of injury, which in turn
attract fibroblasts. The fibroblasts then repair the site by producing new connective tissue
matrix. and Remodeling/Maturation Phase this is the period of scar contracture with
collagen cross-linking, shrinking, and a loss of edema. It is characterized by an
equilibrium between collagen synthesis and collagen degradation in an effort to
reestablish the connective tissue matrix.


1. Epithelization sealed by clot formation, epithelial cell migration across the
defect, and keratinocytesdetachment, migration, proliferation, differentiation,
2. Contraction Inward movement of the edges of the injured tissue. Begins between
days 8 and 10 after injury. Fibroblast and extracellular matrix control the process.
3. Connective tissue matrix deposition components include Collagen, Elastin,
Fibronectin, Laminin, Proteoglycans, and Hyaluronic acid.

Synthesis of Collagen
1. Combination of amino acid to form chains
2. Chains associate to form molecules
3. Molecules associate to form fibrils
4. Fibrils aggregate into fibers or bundles

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Wound healing processes are known to be influenced by among other factors by
infections, nutritional status, drugs and hormones, type and sites of wound, and wasting
diseases like diabetes (Karl et al, 1995). In fokelore medicine, medicinal plants have been
used widely in facilitating wound healing with high degree of successes.

Laboratory Animals
The Laboratory rats like its wild counterpart is an extremely intelligent animal,
probably more so than the other rodents commonly used for biomedical research.
Scientists and researchers rely on mice and rats for several reasons. One is convenience:
rodents are small, easily housed and maintained, and adapt well to new surroundings.
They also reproduce quickly and have a short lifespan of two to three years. Rats are also
relatively inexpensive and can be bought in large quantities from commercial producers
that breed rodents specifically for research. The rodents are also generally mild-tempered
and docile, making them easy for researchers to handle (Robert W. Kemp).
Another reason rodents are used as models in medical testing is that their genetic,
biological and behavior characteristics closely resemble those of humans, and many
symptoms of human conditions can be replicated in mice and rats. "Rats and mice are
mammals that share many processes with humans and are appropriate for use to answer
many research questions," said Jenny Haliski, a representative for the National Institutes
of Health (NIH) Office of Laboratory Animal Welfare.

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Herbal Ointment
Ointment is a semisolid preparation for external application to the body. Some
contain oils, fats, or beeswax along with beneficial herbs. They are used to protect, heal,
and moisturize the skin.

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Chapter III

The materials used in this study are: basin, knife, chopping board, blender,
beakers, weighing balance, whatman no.2 filter paper, graduated cylinder, spatula,
stirring rod, water bath, surgical blade, cotton, tissue, forceps, syringe, and caliper, as
well as the Albino rats and the Buddha belly leaves (Jatropha podagrica).

The drug that was used for the comparison of commercial drug from the Buddha
belly leaves extract is Bactroban (2%).

T1 = 1ml/10g of ointment base
T2 = 1.5ml/10g of ointment base
T3 = 2ml/10g of ointment base
T4 = Bactroban ointment 2% (Positive control)
T5 = No Ointment applied (Negative control)

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Collection of plant materials and extract preparation
The fresh leaves were obtained from Sacpil, Conner, Apayao. And were
identified and authenticated at the National Museum, Manila.
The leaves were washed, chopped into smaller pieces and sun-dried. The dried
leaves were grounded into powder with the aid of a blender. Thereafter, 100 g of the leaf
powder added with 100ml of distilled water was macerated with 300 ml of methanol for
72 h and agitated intermittently. The extract was filtered using Whatmans No. 2 filter
paper and the filtrate was concentrated to dryness in vacuo using a water bath for 72hrs to
remove the methanol.

Preparation of Jatropha Extract Ointments

All the materials used were under aseptic condition. 30g of petroleum jelly (white
petrolatum) were weighed into a beaker and then melted in a thermostatic water bath.
Five batches of the ointments were prepared and use for the study. Batches 1-3 contained
the three extracts of varying concentration (1ml, 1.5ml and 2ml per 10g of the ointment
base). Batch 4 was then formulated to contain a commercial bactroban (2%), used as a
standard drug treatment. Batch 5 was remained untreated.
The required quantities of the extract were weighed, added to the molten ointment
base and then homogenize by trituration. The ointments were stored in the refrigerator
until they were used.

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Experimental animals
Thirty (30) albino rats (250-300 g) were obtained from Cartimar Pasay, Manila
and place in five groups (n = 5) for the studies. They were allowed to acclimatize in the
research laboratory of Cagayan Valley Herbal Plant Processing (CVHPP) for 7 days
prior to the commencement of the study and fed with standard livestock pellets. The
animals were allowed unrestricted access to clean drinking water.

Induction of Wounds
The predetermine area for wound induction at the back of the animal were
prepared for surgery by removing the dorsal fur or hairs with a pair of scissors and
scalpel blade. The animals were held in standard crouching position, and the mobile skin
were gently stretched and held by fingers. The induction of a tranquilizer drug
Acepromazine maleate (.1ml/250g of rat) were done by IM injection and the induction of
localized anesthesia were also done by subcutaneous injection of a lidocaine solution
(0.3ml/2%) at the dorsal part of the animal and around the area under investigation to
render area painless. After five(5) to eight(8) minutes of introducing local anesthesia, the
rat were then subjected for incisional wound (Celestino et al, 2014).
The anticipated area of the wound created were outlined on the back of the
animals. A longitudinal paralumbar incision, 1 inch in length and .2 cm in
depth were incised through the skin using toothed forceps, a surgical blade
and pointed scissors. After achieving complete homeostasis by blotting the wound with
cotton swab soak in warm saline, the animals were placed singly in individual cages.

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The wounds of the animals were treated topically depending on the group. Groups
1, 2 and 3 were treated with ointments formulated from extracts at various concentrations
(1ml, 1.5ml and 2ml of extracts per 10g of ointment base). Group 4 were treated with the
standard bactroban ointment (batch 4-Positive control), while group 5 serves as the
negative control leaved untreated (batch5) which heals naturally. T1, T2, and T3
wounded animals were treated topically with Jatropha podagrica ointment every 8 hours
starting from the first day. The respective therapeutic treatment is administered topically
to the animals of respective groups until complete epithelialization starting from the day
of operation. The animals were subsequently returned to their cages and transported to
the holding rooms. Bedding in the cages were changed every 3days and cages were kept
clean to avoid contamination. Application of standard healing agent and the prepared
ointment were done twice a day. The wound length were measured with a caliper every 3
days until epithelialization and complete wound closure were observed. Wound
contraction were calculated as a percentage of original wound size (Esimone et al 2008).

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1) To compare the mean wound area and to determine the percentage of
2) To promote wound healing in a shortest time possible in 21 days.

Statistical analysis
The date were analyzed using one way analysis of variance (ANOVA and data
subjected to LSD post hoc test. Differences in mean between paired observations were
accepted as significant at P 0.05. Also, the data will be analyzed using tables, means, and


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Ahmed , Hassanali et al. Antifeedant Activity of Jatropha Podagrica Roots. Retrieved

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