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REVIEW ARTICLE ON ANTIMICROBIAL ACTIVITY OF THESPESIA POPULNIA SEEDS

 

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About Author:
Sharad kumar awasthi

Shri Ram Institute Of Technology,
Jabalpur (M.P.) 482002.
Sharad20n@gmail.com

ABSTRACT:
Thespesia populnia seeds were dried and then grinded to obtain fine powder. Ethanolic and aqueous extracts and etheral extract of thespesia seeds were evaluated for antibacterial activity against E.coli, B.subtilis with the comparison control (Ciprofloxacin and Amoxicillin) for bacteria. The extract showed good inhibitory activity on the microbes.


Reference Id: PHARMATUTOR-ART-1561

INTRODUCTION

1.1 THESPESIA POPULNEA: Commonly known as the Portia Tree, is species of flowering plant in the mallow family, Malvaceae. It is a small tree or arborescent shrub that has a pantropical distribution, found on coasts around the world. However, the Portia Tree is probably native only to the Old World, and may have originated in India. It is possibly indigenous to the Hawaiian Islands and elsewhere in the Pacific, but may have been spread by early Polynesians for its useful wood and bast fibres. The Portia Tree reaches a height of 6–10 m (20–33 ft) tall and a trunk diameter of 20–30 cm (7.9–12 in). It grows at elevations from sea level to 275 m (902 ft in areas that receive 500–1,600 mm (20–63 in) of annual rainfall. The Portia Tree is able to grow in the wide range of soil types that may be present in coastal environments, including soils derived from quartz (sand), limestone, and basalt; it favors neutral soils (pH of 6-7.4).

SCIENTIFIC CLASSIFICATION


1.2 ANTIMICROBIAL ACTIVITY
Drug substances that either suppress or influence the growth of micro organisms are generally analysed by microbial method .Two procedures are generally employed in microbial assay.The increased prevalence of antibiotic-resistant bacteria due to the extensive use of antibiotics may render the current antimicrobial agents inefficient to control some bacterial diseases (Tanaka et al., 2006). Herbal medicine is frequently a part of a larger therapeutic system such as traditional and folk medicine. It is necessary to evaluate, in a scientific base, the potential use of folk medicine for the treatment of infectious diseases produced by common pathogens. Medicinal plants might represent an alternative treatment in non-severe cases of infectious diseases. They can also be a possible source for new potent antibiotics to which pathogen strains are not resistant. The search and use of drugs and dietary supplements derived from plants have been accelerated in recent years. Ethnopharmacologist, botanist, microbiologist and natural product chemist are combing the medicinal flora for biological substances that could be developed for the treatment of infectious diseases. Several medicinal plants have been extensively studied in order to find more effective and less toxic compounds. [3]

1.3 BOTANICAL DESCRIPTION :
Preferred scientific name : Thespesia populnea (L.) Sol. ex Correa

Family : Malvaceae (mallow family)

Non-preferred scientific names : Hibiscus populneus L. (1753),  Thespesia macrophylla Blume (1825), Hibiscus populneoides Roxb. (1832)

1.3.1 MORPHOLOGY:


Size and form :  Milohas a short, straight or crooked trunk and a dense crown with crowded lower horizontal branches. Height averages 6–10 m (20–33 ft) with a crown as wide as or wider than the tree is tall. Average size trees have bole diameters of 20–30 cm (8–12 in), but exceptional trees have reached 18 m (60 ft) in height with boles 60 cm (24 in) in diameter. The bark is grey and smooth to highly fissured and dark brown in larger trees.[4]

Flowers
Flowers are a typical hibiscus shape in appearance: bellshaped, 4–7 cm (1.5–2.5 in) in length, with five overlapping, broad, rounded petals. Color is pale yellow with a maroon spot at the base of each petal and with star-shaped hairs on outer surface.

Leaves
The alternate leaves are glossy green above and paler green below.

Fruit
Milofruits are brittle, dry, woody or papery seed capsules, rounded and flattened, containing five cells and several seeds.

Seeds


The brown, hairy seeds are about 1 cm (0.4 in) long and 0.6 cm (0.2 in) broad. Seeds are blown short distances by wind but are more likely to be dispersed by water. Both the lightweight fruits and seeds can float from one island to another on ocean currents. There are between 3500 and 6700 seeds/kg (1600–3045 seeds/lb)[1][2][3]

1.3.2 CHEMICAL CONSTSTITUENT :
Phytochemicals such as flavonoids, steroids, terpenoids, tannins, Saponins, alkaloids, phenols, carbohydrates  and proteins, glycosides, reducing sugars, anthocyanins, anthraquinons , leucoanthocyanins and emodins  are present in Thespesia populnea. Populneol isolated and identified as monobenzyl ether of y-resacetophenone; gossypol isolated from flowers: its methylation yielded three isomeric optically active heramethyl ethers which were characterized. Kaempferol, quercetin, rutin, kaempferol-3- rutinoside, kaempferol-3-glucoside, kaempferol-5-glucoside and quercetin-3-glucoside isolated from flowers; calycopterin isolated from heartwood.

1.3.3 MEDICINAL USES:
·        According to Ayurveda, Paraspipal is astringent, acrid, cooling, depurative, vulnerary, alternative and useful in skin related troubles, leprosy, diseases of blood and urinary system, diarrhea, dysentery, Cholera, diabetes, ascites etc.
·        In Fiji, a decoction of the leaves has been used in treating coughs and headaches.
·        An infusion of the bark has been used to treat intestinal diseases. In Tonga, a drink made from the leaves and bark is used to treat fevers in teething children.
·        Various parts of the plants have high tannin contents and plant extracts have been shown to have anti-bacterial and anti-viral activity.
·        The leaves are applied locally in swollen joints for their anti-inflammatory effects and also for skin diseases, hepatitis, jaundice, ulcers, wounds, psoriasis, scabies, urinary tract infections, diabetes, cholera, cough, asthma and guneaworm infections .

2. LITERATURE REVIEW:
1) M. Vasudevan  et.al, reported  Pharmacological actions of Thespesia populnea relevant to Alzheimer's disease.
2) A SARAVANAKUMARet.al, EVALUATE ANTIBACTERIAL ACTIVITY, PHENOL AND FLAVONOID CONTENTS OF THESPESIA POPULNEA FLOWER EXTRACTS.
3) Sompong Boonsri et.al, reported Cytotoxic and Antibacterial Sesquiterpenes from Thespesia populnea.
4) Annie, S et.al,reported Antihepatotoxic Activity of Thespesia populnea.
5) Pankajamani R. et al, Isolate  gossypol from Thespesia.
6) Vasudevan, M. et.al,, Antinociceptive and anti-inflammatory     effects of Thespesia populneabark extract.
7) Krishnamoorthy, P. et.al, reported Effect of the extract of Thespesia populnea  leaves on mice testis. 
8) Ilavarasan, R. et.al, reported Antioxidant activity of Thespesia populnea bark extracts against carbon tetrachloride-induced liver injury in rats.
9) Nagappa, A.N.et.al, reported  Wound healing activity of the aqueous extract of Thespesia populnea fruit. 
10) Kavimani, S.et.al, reported Anti-steroidogenic activity of floral extract of Thespesia populnea Corr. in mouse ovary.
11) Satyanarayana T. et.al, reportedAntihyperglycemic and hypoglycemic effect of Thespesia populnea fruit in normal and alloxan-induced diabetes in rabbits.

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4. MATERIAL & METHOD

4.1 COLLECTION OF HERBS:
Collection of herb is generally influenced by the availability of the herbs as well as the part of the plant to be collected. The herbs was purchased from the local market from jabalpur district of M.P. in september 2011.

4.2 HARVESTING
The seeds were washed with running water and crushed using pestle-mortar.
4.3EXTRACTION AND PHYTOCHEMICAL SCREENING :

4.3.1. EXTRACTION:
(a)        PLANT MATERIAL:       Seeds of Thespesia populnea
(b)       CHEMICALS:         Ethanol, petroleum ether,water
(c)        PREPARATION OF EXTRACT: The seeds of Thespesia populnea after crushed and extracted using Soxhlet Apparatus with above given  solvent.

EXTRACTIVE VALUE

EXTRACT

WEIGHT OF HERB

EXTRACTIVE VALUE

Ethanolic extract

50gm

1.25gm

Etheral extract

50gm

4.0ml

Aquous extract

50gm

0.5mg

4.3.2  PRELIMNARY PHYTOCHEMICAL INVESTIGATION:
Identification Of The Plant Constituents By Phytochemical Tests:  
Ethanolic extract is subjected to various preliminary phytochemical analysis to test for the presence or absence of various phytoconstituents by the following tests.

1. Test for alkaloids:   To the extract dilute hydrochloric acid will be added and filtered. The filtrate will be treated with various alkaloid reagents.
a) Mayer’s test:   
The filtrate will be treated with Mayer’s reagent: appearance of cream colour indicates the presence of alkaloids.
b) Dragendroff’s test:  
The filtrate will be treated with Dragendroffs reagent: appearance of reddish brown precipitate indicates the presence of alkaloids.
c) Hager’s test:    
The filtrate when treated with Hager’s reagent, appearance of yellow colour precipitate indicates the presence of alkaloids.

2) Test for carbohydrates and reducing sugar :  The small quantities of the filtrate will be dissolved in 4ml of distilled water and filtered. The filtrate will be subjected to
a) Molisch’s test: 
A small portion of the filtrate will be treated with Molisch’s reagent and sulphuric acid. Formation of a violet ring indicates the presence of carbohydrates.
b) Fehling’s test: 
The extract will be treated with Fehling’s reagent A and B. The appearance of reddish brown colour precipitate indicates the presence of reducing sugar.
c) Benedict’s test: 
The extract will be treated with Benedict’s reagent; appearance of reddish orange colour precipitate indicates the presence of reducing sugar.
d) Barfoed’s test: 
The extract will be treated with barfoed’s reagent and heated. Appearance of reddish orange colour precipitate indicates the presence of non reducing sugars.

3) Test for proteins:
a) Biuret test: 
The extract will be treated with copper sulphate solution, followed by addition of sodium hydroxide solution; appearance of violet colour indicates the presence of proteins.

b) Millon’s test:  The extract will be treated with Millon’s reagent; appearance of pink colour indicates the presence of proteins.

4) Test for tannins:  The extract will be treated with 10% lead acetate solution; appearance of white precipitate indicates the presence of tannins.

5) Test for phenolic compounds:
a) The extract will be treated with neutral ferric chloride solution; appearance of violet colour indicates the presence of phenolic compounds.
b) The extract will be treated with 10% sodium chloride solution; appearance of cream colour indicates the presence of phenolic compounds.

6) Test for flavonoids:
a) 5ml of extract will be hydrolyzed with 10%sulphuric acid and cooled. Then, it will be extracting with diethyl ether and divided in to three portions in three separate test tubes. 1ml of diluted sodium carbonate, 1ml of 0.1N sodium hydroxide, and 1ml of strong ammonia solution will be added to the first, second and third test tubes respectively. In each test tube. Development of yellow colour demonstrated the presence of flavonoids.

b)Shinoda’s test:   The extract will be dissolved in alcohol, to which few magnesium turnings will beaded followed by concentrated HCL drop wise and heated, and appearance of magenta colour shows the presence of flavonoids.

7) Test for glycosides :  When a pinch the extract was treated with glacial acetic acid and few drops of ferric chloride solution, followed by the addition of conc. Sulphuric acid, formation of ring at the junction of two liquids indicates the presence of glycosides.

8) Test for saponins
Foam test : 
About 1 ml of the extract was diluted to 20 ml of with distilled water and shaken well in a test tube. The formation of foam in the upper part of test tube indicates the presence of saponins.

4.4 ANTIMICROBIAL ACTIVITY

1.  CYLINDER PLATE METHOD(CUP):  This is based on measurement of the diameter of microbial growth inhibition surrounding the cylinders containing various dilutions of the test compound which are placed on the surface of a solid nutrient medium previously inoculated with a culture of suitable microbe.Inhibition produced by the test compound is compared with that produced by known concentration of a refrence standard.

PROCEDURE:
1.         Two bacteria, (gram +ve i.e.B.subtilis, gram-ve i.e E. Coli) were used in the present study to determine the antibacterial activity of the crude extracts by agar diffusion method (cup plate method).
2.       In the agar diffusion method, nutrient agar for bacteria used as culture media and cavity were aseptically made over the culture plates using borer (9mm internal diameter).
3.       The cavities were filled with extracts, standards and control The plates were incubated at 37 0 C for 24 hrs.
4.      The activities were determined by measuring the diameter of the zone in mm.
5.      The experiment was replicated two times to confirm the reproducible results.
6.       Solvent used as negative control in each time. Standard Ciprofloxacin (100mcg/ 0.1ml), Amoxicillin (100mcg/ 0.1ml) for bacteria were used as positive control for comparison of the activities. 

5.RESULT :

S.NO

NAME

AQUOUS EXTRACT

PETROLIUM ETHER EXTRACT

ETHANOLIC EXTRACT

1

FLAVONOIDS

+

+

+

2

TANNINS

+

_

_

3

GLYCOSIDES

+

+

+

4

SAPONINS

+

_

_

5

PHENOLS

_

_

_

6

ALKALOIDS

+

+

+

7

REDUCING SUGAR

+

_

_

8

CARBOHYDRATE

+

_

+

9

PROTEIN

+

+

+

OBSERVATION TABLE

S.No

Name of drug

Conc. in mcg/disc

E.coli

mm

B.subtilis

1

Ciprofloxacin

100

41

40

2

Amoxicilin

100

21

27

3

Ethanolic extract

300

23

14

4

Aquous extract

300

14

12

5

Petroleum ether

300

20

16

CONCLUSION:
Ethanolic and aqueous extracts of thespesia seeds were evaluated for antibacterial activity against E.coli, B.subtilis with the comparison control (Ciprofloxacin and Amoxicillin) for bacteria. The extract showed good inhibitory activity on the microbes.

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