“Evaluation of aqueous and alcoholic extracts of stem bark of

Spathodea campanulata for antiulcer activity”

SYNOPSIS FOR

M. PHARM DISSERTATION

SUBMITTED TO

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES

BENGALOORE, KARNATAKA

SUBMITTED BY

RADIKA D.S

I M. PHARM

DEPARTMENT OF PHARMACOLOGY

P.E.S COLLEGE OF PHARMACY

BENGALOORU– 560050

(2008-2010)

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES

BENGALOORE, KARNATAKA.

ANNEXURE-II

PROFORMA FOR REGISTRATION OF SUBJECT FOR

DISSERTATION

1. / NAME AND ADDRESS OF THE CANDIDATE. / RADIKA D.S
LOCAL ADDRESS
P.E.S. COLLEGE OF PHARMACY
50 feet road, Hanumanthnagar,
B.S.K 1st stage, Bangalore – 560 050
PERMANENT ADDRESS
D/O Srinivas D.N
Doddadasarahalli
Shidlaghatta(t)
Chikbalapur (d)
2. / NAME OF THE INSTITUTION. / P.E.S. COLLEGE OF PHARMACY
50 feet road, Hanumanthnagar,
B.S.K 1st stage, Bangalore – 560 050
3. / COURSE OF STUDY AND SUBJECT. / M.PHARM.
PHARMACOLOGY.
4. / DATE OF ADMISSION TO COURSE. / 16th JUNE 2008
5. / TITLE OF THE TOPIC:
“Evaluation of aqueous and alcoholic extracts of stem bark of
Spathodea campanulata for antiulcer activity ”
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7.4 / BRIEF RESUME OF THE INTENDED WORK
NEED FOR THE STUDY:
Gastric hyperacidity and gastro duodenal ulcer is a very common global problem today. The etiopathogenesis of peptic ulcer has changed from Schwartz dictum ‘No acid (gastric juice)-No ulcer’ to ‘No mucosal damage-No ulcers’.
Peptic ulcer occurs in that part of the gastrointestinal tract which is exposed to gastric acid and pepsin, i.e.,the stomach and duodenum. The etiology of peptic ulcer is not clearly known. it results probably due to an imbalance between the aggressive (acid, pepsin and H.pylori) and the defensive (gastric mucus and bicarbonate secretion,prostaglandins,nitric oxide, innate resistant of the mucosal cells)factors.1
Hypersecretion of gastric acid is a pathological condition, which occurs due to uncontrolled secretion of hydrochloric acid from the parietal cells of the gastric mucosa through the proton pump H+ K+ ATPase. Even the normal rate of acid secretion may cause ulceration in the breached mucosa when some gastro protective factors are lost. Peptic ulcer disease is one of the most common gastrointestinal disorders, which causes a high rate of morbidity particularly for the population of non-industrialized countries.2
Several factors are implicated in the pathogenesis of gastric ulcer including: increased acid-pepsin secretion, impaired bicarbonate neutralization, impaired mucus secretion and precipitate lesions on the mucosal layer.3,4
In recent years, gastric ulcer has also been associated with infection of gastro intestinal mucosal tissue by Helicobacter pylori. About 70% of patients with peptic ulcer disease are infected by Helicobacter pylori and eradication of this microorganism seems to be curative for the disease. 5
Extracts of bark, leaves and flowers of Spathodea campanulata are used to treat malaria, HIV, diabetes mellitus, oedema, dysentery, constipation, gastrointestinal disorders, ulcers, skin diseases, wounds, fever, urethral inflammation, liver complaints and as a poison antidote. It may be effective as a malaria prophylactic and in the control of Aedes mosquitoes.6, 8 As now there is no scientific evidence for the evaluation of of antiulcer activity of s.campanulata.Hence present study is proposed to evaluate alcoholic and aqueous extracts of stem bark of spathodea campanulata for antiulcer activity.
REVIEW OF THE LITERATURE:
PLANT PROFILE
Introduction
Spathodea campanulata is a species belonging to the Bignoniaceae family, native from equatorial Africa. It is a medium-size tree (15-25mheight) characterized by red garish flowers. It is often employed in garden in tropical and subtropical areas, including South America (JOLY, 1985). This species has many uses in folk medicine for example, the flowers are employed as diuretic and anti-inflammatory, while the leaves are used against kidney diseases, urethral inflammations and as an antidote against animal poisons. The stem bark preparations are employed against ulcers, enemas, skin diseases, herpes, stomachaches and diarrhea. It is used in traditional herbal medicine for the treatment of ulcers, fever, gonorrhea and diarrhea.7,8
Synonym(s)
Spathodea nilotica Seem.
Vernacular names.9
Kannada : neerukayi mara
English : African tulip tree, flame of the forest, fountain tree, Nandi flame, Nile flame, squirt
tree, tulip tree, Uganda flame
French : immortel éntranger
Hindi : rugtoora
Luganda : kifabakazi
Malayalam : panchut-panchut
Sinhala : kudaella gaha, kudulu
Spanish: amapola, espatodea, mampolo, tulipán africano
Swahili : kibobakasi, kifabakazi
Tamil : patadi.
Botanical description.9
Spathodea campanulata is medium sized, reaching a height of 10-35 m, deciduous, with a round, heavy crown of dense, dark foliage, sometimes somewhat flattened; young bark
pale, grey-brown and smooth but turns grey-black, scaly and cracked vertically and horizontally with age.
The opposite imparipinnate leaves are exstipulate. Each leaf consists of 5-7 pairs of opposite leaflets and a terminal one. The leaflets are oblong-elliptic, about 1 cm long and 0.5 cm broad, entire, broadly acuminate, unequal at the base, dark green on top and light green on the underside; there are glandular swellings at the base of the lamina (usually a pair); the midrib and nerves are yellow, raised and very slightly pubescent; the venation is reticulate; the short, thick petiole is about 0.7 cm long; there are conspicuous lenticels on the rachis; rachis base is swollen.
Flowers large, red, hermaphrodite, orange inside; calyx green, about 1 cm long and split on the posterior side, ribbed and tomentellous; petals 5, each about 1.5 cm long; stamens 4 with orange filaments; style extruding with a 2-lipped stigma; flower buds curved and contain a red sap. A yellow-flowered variety has been reported.
Fruit upstanding, dark brown, cigar-shaped, woody pod, 15-25 cm long and split on the ground into 2 boat-shaped valves, releasing many flat-winged seeds; 1-4 pods usually develop from 1 flower cluster; seeds thin, flat and surrounded by a filmy wing.
Ecology and distribution.9
History of cultivationThe species is found throughout tropical Africa and is widely grown as an ornamental.
Natural HabitatS. campanulata grows naturally in Africa in secondary forests in the high forest zone and in deciduous, transition, and savannah forests.
Geographic distributionNative : Angola, Ethiopia, Ghana, Kenya, Sudan, Tanzania, Uganda, ZambiaExotic : Colombia, Costa Rica, Cuba, India, Jamaica, Puerto Rico, Sri Lanka, Zanzibar.
chemical constituents: -
Spathoside, a new cerebroside was isolated from the stem bark of Spathodea campanulata, besides known compounds (n-alkanes, linear aliphatic alcohols, sitosterol and their esters, β-sitosterol-3-O-β-D-glucopyranoside, oleanolic acid, pomolic acid, p-hydroxybenzoic acid and phenylethanol esters). Spathodic acid,steroids, saponins, ursolic acid,and tomentosolic acid.10
Uses:-
Spathodea campanulata has many medicinal uses both where it is native and introduced. Extracts of bark, leaves and flowers are used to treat malaria, HIV, diabetes mellitus, oedema, dysentery, constipation, gastrointestinal disorders, ulcers, skin diseases, wounds, fever, urethral inflammation, liver complaints and as a poison antidote. It may be effective as a malaria prophylactic.6,8
Traditional medicinal uses:-
It is used in traditional herbal medicine for the treatment of ulcers, filaria, gonorrhea, diarrhea and fever. S. campanulata was also known in Cameroon traditional medicine to have a healing activity in burn wounds.
Literature:-
Spathoside, a new cerebroside was isolated from the stem bark of Spathodea campanulata, besides known compounds. The antibacterial activity of the isolated compounds against a wide range of microorganisms was examined. They inhibited significantly the growth of some gram-positive and gram-negative bacteria.10
Spathodea campanulata is used as wound-healing agent in Ashanti traditional medicine in Ghana. The methanol extract of and Spathodea campanulata bark showed some level of antimicrobial activity exhibiting selective antifungal activity against Trichophyton species. The antioxidant activities and antimicrobial activities suggest that the use of the plants in wound healing may be based on antioxidant and antiseptic effects of its constituents.11
The extracts have been shown to have anti-inflammatory and antioxidant properties due to flavonoids, triterpenoids, diterpenoids and caffeic acid derivatives from spathodea campanulata.12
Insect mortality in Spathodea campanulata Beauv. (Bignoniaceae) flowers.13
Spathodea campanulata stem bark decoction (SCD) has shown hypoglycemic activity in mice. It was separated by column chromatography into different fractions, which were evaluated for their hypoglycemic, anticomplement and anti-HIV activities. The most polar fraction exerted by far the most prominent effect in different biological models.14
The antimalarial activity of Spathodea campanulata stem bark extract on Plasmodium berghei berghei in mice.15
OBJECTIVE OF STUDY:-
Ø  Preparation of alcoholic and aqueous extracts of stem bark of Spathodea campanulata.
Ø  To investigate preliminary phytochemical constituents of alcoholic and aqueous extracts of stem bark of Spathodea campanulata.
Ø  Determination of LD50 of the aqueous extract of Spathodea campanulata as per OECD guidelines.
Ø  To establish the pharmacological profile of prepared extracts for its antiulcer activity.
MATERIAL AND METHODS:-
SOURCE OF DATA:-
Whole work is planned to generate data from laboratory studies i.e. experiments are performed as described in references. Experimental studies in journals and in text books available with college and various institutions.
IISc library, Bangalore.
PESCP library, Bangalore
RGUHS digital library(Helinet), Bangalore
Web sites: www.scienencedirect.com
www.pubmed.com
www.google.com
www.jhetchem.com
www.ijp-online.com
PLAN OF WORK:
The whole study is divided in the following phases
Phase I: Collection of plant material.
The stem bark of the tree spathodea campanulata will be collected from wild source and will be authentified. They will be subjected for alcoholic and aqueous extractions. The concentrated extract will be useful for our studies.
Phase II: Preparations of extracts.
Alcoholic extract and aqueous extract of stem bark of spathodea campanulata. Alcoholic extract is prepared by successive extraction with soxhlet apparatus. The marc obtained after alcoholic extraction was macerated with water to obtain an aqueous extract.
Phase III: Preliminary Phytochemical investigation.
Preliminary phytochemical investigation has been done in this phase as described by Khandelwal.
Phase IV: Determination of LD50 of the aqueous extract of Spathodea campanulata as per OECD guidelines.
PHASE V:
For determination of antiulcer activity the following experiments are to be performed.
All the models used in the pharmacological experiments will consist of the below 7 groups consisting of 6 animals in each group. Separate set of animals shall be used for individual test (or) models
GROUP CLASSIFICATION:
TREATMENT / DRUG / DOSE
Group-1 / Vehicle / -
Group-2 / Control / -
Group-3 / Omeprazole ( Standard ) / -
Group-4 / Alcoholic extract of Spathodea campanulata / High dose
Group-5 / Alcoholic extract of Spathodea campanulata / Low dose
Group -6 / Aqueous extract of Spathodea campanulata / High dose
Group-7 / Aqueous extract of Spathodea campanulata / Low dose
Evaluation of anti-ulcer activity will be done using following models
ANTI-ULCER ACTIVITY
1.  Pylorus ligation induced ulcer model
2.  Indomethacin induced ulcer model
3.  Cold stress induced ulcer model
1.PYLORUS LIGATION INDUCED ULCER MODEL
Forty two rats of either sex were randomly allotted to seven groups each consists of six animals. Group I served as the vehicle control; Group II served as the pylorus ligation control (4 h); Group III served as standard control; Group IV and V –alcoholic extract; and Group VI and
VII-aqueous extract; were administered orally respectively by gavage using feeding needle.
Albino rats of either sex weighing 150-200 g were used. After 18 hours of fasting, ulcer induction was undertaken. The rats were quickly and mildly anaesthetized with ether and the abdomen was cut open through a midline incision. The pylorus was secured and ligated with silk sutures, after which the wound was closed and the animal were allowed to recover from anesthesia.
After ligation of the pylorus, drinking water was withheld and the gastric examinations were under-taken 19 hours after pylorus ligation. The animals were sacrificed with an overdose of ether and the stomachs were removed after clamping the esophagus. The gastric contents were collected through the esophagus. The gastric juice was centrifuged and volume was recorded.16,17
Total acidity of gastric juice
An aliquot of 1ml of gastric juice was taken in to a 50 ml conical flask and two drops of phenolphthalein indicator was added and titrated with 0.01N NaOH until a permanent pink color was established. The volume of 0.01N NaOH consumed was noted. The total acidity was expressed as meq/l by the following formula: N × 0.01× 36.45 × 1000; which N is volume of NaOH consumed, 40.0 is molecular weight of NaOH, 0.01 is normality of NaOH and 1000 is the factor to be respected in liter.
Macroscopic evaluation of stomach
The stomachs were opened along the greater curvature, rinsed with saline to remove gastric contents and blood clots and examined by a ×5 magnifier lens to assess the formation of ulcer. The number of ulcers was counted. Ulcer scoring was undertaken.
The scores were: 0 = no ulcer, 1= superficial ulcer, 2= deep ulcer, 3= perforation.Ulcer area was assessed by using 3 M scaled surgical transpore tapes, which was fixed on a light and transparent sheet. Each cell on the tape was 1mm2 in area, so the number of cells was counted and the ulcer
area was measured for each stomach.Ulcer index was measured by using following
formula.
UI = UN + US + UP × 10-1
which UI = ulcer index, UN = mean of ulcer number, US =mean of ulcer score, UP = ulcer probability (incidence %) for each group.16,18
2.INDOMETHACIN INDUCED ULCER MODEL
Forty two rats of either sex were randomly allotted to seven groups each consists of six animals. Group I served as the vehicle control; Group II served as the indomethacin control; Group III served as standard control; Group IV and V –alcoholic extract; and Group VI and VII-aqueous extract; were administered orally respectively by gavage using feeding needle.
Albino rats of either sex weighing 150-200 g were used.The test drugs are administered orally in 0.1% Tween 80 solution 10 min prior to oral indomethacin in a dose of 20 mg/kg (4 mg/ml dissolved in 0.1%Tween 80solution). Six hours later, the rats are sacrificed in CO2 anesthesia and their stomachs removed.
Formol-saline (2% v/v) is then injected into the totally ligated stomachs for storage overnight. The next day, the stomachs are opened along the greater curvature, then washed in warm water, and examined under a 3-fold magnifier.
The scores were: 0 = no ulcer, 1= superficial ulcer, 2= deep ulcer, 3= perforation. Ulcer area was assessed by using 3 M scaled surgical transpore tapes, which was fixed on a light and transparent sheet. Each cell on the tape was 1mm2 in area, so the number of cells was counted and the ulcer
area was measured for each stomach (15).Ulcer index was measured by using following
formula.
UI = UN + US + UP × 10-1
Where UI = ulcer index, UN = mean of ulcer number, US =mean of ulcer score, UP = ulcer probability (incidence %) for each group.18,19