Journal of Pharmacognosy and Phytochemistry 2020; 9(2): 805-809

E-ISSN: 2278-4136
P-ISSN: 2349-8234
www.phytojournal.com Pharmacognostic and phytochemical evaluation of
JPP 2020; 9(2): 805-809
Received: 04-01-2020 Blepharis repens
Accepted: 06-02-2020

Sawan K Rathod Sawan K Rathod, Vinod M Thakare and Hemant S Kanhere

Dadasaheb Balpande College of
Pharmacy, Nagpur,
Maharashtra, India
Abstract
Blepharis repens (Vahl.) Roth, (Synonym–Acanthus repens Vahl, Blepharis molluginif oliapers)
belonging to family Acanthaceae, commonly known as Hadsan (in Marathi). The plant has natural
Vinod.M. Thakare
distribution indeciduous forest. There are about 130 species distributed throughout tropical and
Dadasaheb Balpande College of
Pharmacy, Nagpur, subtropical region of the world. In Maharashtra this genusis represented by two species: Blephari srepens
Maharashtra, India (Vahl) Roth. And B. Madras petansis (L.) Roth. Mostly all parts are medicinally used. In present study
Pharmacognostic and Phytochemical Evaluation were carried out.
Hemant S. Kanhere
Dadasaheb Balpande College of Keywords: Pharmacognostic, phytochemical, Blepharis repens
Pharmacy, Nagpur,
Maharashtra, India Introduction
Acanthaceae is one of the large pantropical family of about 229 genera and 3450 species.
Family Acanthaceae have been reviewed by several authors from embryological point of view,
in order to solve various taxonomic problems [1].
Plant-derived substances have recently become of great interest owing to their versatile
applications. Medicinal plants are the richest bio-resource of drugs of traditional systems of
medicine, modern medicines, nutraceuticals, food supplements, folk medicines,
pharmaceutical intermediates and chemical entities for synthetic drugs [2].
Plants have played a significant role in maintaining human health and improving the quality of
human life for thousands of years and have served humans well as valuable components of
medicine, seasonings, beverages, cosmetics and dyes. Blepharis repens (Vahl.) Roth,
(Synonym –Acanthus repens Vahl, Blepharis molluginif oliapers) belonging to family
Acanthaceae, commonly known as Hadsan (in Marathi) the decoction of leaves used in
treatment of old persistent fever and the paste is used for fractured bones [3].
Blepharisrepens (Vahl.) Roth, is a plant of medicinal importance and no pharmacog-nostical
studies are available on this species, so the present investigations was undertaken. Which is a
traditional medicinal herb of the family Acanthaceae. This plant is used traditionally to treat
bone fractures, skin diseases, urinary discharges and allergies. Whole plant is highly
medicinal, used against chronic fever and as a diuretic, aphrodisiac and expectorant, and also
for urinary discharges. Stem powder is also consumed to curebone fracture. The plant has
natural distribution indeciduous forest. There are about 130 species distributed throughout
tropical and subtropical region of the world. In Maharashtra this genusis represented by two
species: Blephari srepens (Vahl) Roth. andB. Madras petansis (L.) Roth [4].
The plant Blepharis repens a rare medicinal species, belongs to the Family Acanthaceae. This
leaves may contain cystoliths, calcium and carbonate. Flat branches (phyllodes) are heated and
tied in case of joint-ache. The phyllodes are jointed like the knees. Leaves are roasted and then
extract is obtained. This extract is drunk as a remedy against flatulence. Roots are employed as
antidote on snake – bite. Fruits are roasted and applied on swellings. Stem powder is
consumed to cure bone fracture. The availability of the plant is very low. The importance of
the shoot part will emergence for the in vitro approaches [5].

Plant profile
Name: Blepharis repens (Vahl) Roth [3]
Common name: Hadsan (in Marathi) [3] Hariduhachaga (in kannada) [13]
Kingdom: Plantae – Plants
Corresponding Author:
Order: lamiales
Sawan K Rathod Family: Acanthaceae
Dadasaheb Balpande College of Genus: Blepharis
Pharmacy, Nagpur, Species: repens
Maharashtra, India Synonym: Acanthus repens Vahl
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Chemical constituents: May be Cystoliths, Calcium and

Carbonate [5].

Uses: treat bone fractures, skin diseases, urinary discharges

and allergies [4]. It is used in joint ache and roots are employed
as antidote on snake-bite [5]

Fig 3: Scheme of Successive Extraction.

Preliminary Test for Phytoconstituents

Test for Carbohydrates
Test for Reducing Sugars
A. Molisch’s test: To 2-3 ml aqueous extract, add few drop
of alpha- napthol solution in alcohol, shake and add conc.
sulphuric acid from side of test tube.
B. Fehling’s test: Mix 1 ml Fehling’s A and 1 ml Fehling’s
B solution, boil for one minute, Add equal volume of test
solution. Heat in boiling water bath for 5-10 min.
C. Benedict’s test: Mix equal volume of Benedict’s reagent
and test solution in test tube. Heat in boiling water bath
for 5 min.

Test for Monosaccharide

A. Barfoed’s test: Mix equal volume of Barfoed’s reagent
Fig 1: Blepharis repens (Vahl) Roth and test solution. Heat for 1-2 min. In boiling water bath
and cool.
Experimental Work
Collection of Plant Material Test for Tannins
The plants of Blepharis repens were collected from forest of A. Gelatin test: To a solution of tannin, aqueous solution of
Yavatmal district, Maharashtra, India. The collected plants Gelatin and sodium chloride were added.
was carefully examined for infected parts and were removed B. Ferric chloride: Drug solution + 5% ferric chloride.
accordingly. Only fresh parts were taken for the analysis. C. Acetic acid: Drug solution + acetic acid solution.
These plant parts were dried in the shade till all its moisture D. Bromine water: Drug solution + Bromine water.
gets evaporated.
Test for Alkaloids
Authentication of plant A. Dragendroff’s test: To 2-3 ml filtrate, add few drops
The herbarium sheet was prepared and submitted to Botany Dragendroff’s reagent.
department of Rashtrasant Tukdoji Maharaj Nagpur B. Wagner’s test: 2-3 ml filtrate with few drops Wagner’s
University, Nagpur, and authentified with preference no. reagent.
10227. C. Hager’s test: 2-3 ml filtrate with Hager’s reagent.

Test for Flavonoids

A. Lead acetate test: To small quantity of residue, add lead
acetate solution.
B. Sulphuric acid test: solution + sulphuric acid (66% or
80%).

Test for Saponin Glycosides

A. Foam test: Shake the drug powder vigorously with
water.

Tests for Steroids

A. Salkowaski reaction: To 2 ml of extract, add 2 ml
chloroform and 2 ml conc. H2SO4. Shake well.

Test for Triterpenoids

A. Test solution + 5 ml conc. Sulphuric acid from side of test
Fig 2: Authentification of Blepharis repens (Vahl) Roth.
tube [11]
Extraction
Determination of Extractive Value, Ash Value & Loss on
Ethanolic extract
Drying
About 200g of powder of Blepharis repens (Vahl) Roth. Were
Extractive Value
dried in shade under normal environmental condition and
Ethanol-soluble extractive value
such powder drug was extract into maceration method and
Macerate 5 g of the air-dried drug, coarsely powdered, with
successive extraction was carried out with ethanol solvent.
100 ml of ethanol of the specified strength in a close flask for
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24 hour, shaking frequently during the first 6 hour and filtrate, evaporate to dryness and ignite at low temperature.
allowing to stand for 18 Hour. Thereafter, filter rapidly taking Calculate the percentage of ash with reference to the air dried
precaution against loss of ethanol, evaporate 25 ml of the drug.
filtrate to dryness in a tarred flat-bottomed shallow dish, dry
at 105 °C and weigh. Calculate the percentage of ethanol- Sulphated ash
soluble extractive with reference to the air-dried drug. A silica crucible was heated to redness for 10 minutes,
allowed to cool in desiccators and weighed. 1 g of substance
Water – soluble extractive value was accurately weighed and transferred to the crucible. It was
Add 5 g to 50 ml of water at 80 °C in a stoppered flask. Shake ignited gently at first, until the substance was thoroughly
well and allow to stand for 10 minutes, cool, add 2 g of charred. Then the residue was cooled and moistened with 1
kieselguhr and filter. Transfer 5 ml of the filtrate to a tarred ml concentrated sulfuric acid, heated gently until white fumes
evaporating dish, 7.5 cm in diameter, evaporate the solvent on are no longer evolved and ignited at 800° ± 25 °C until all
a water-bath, continue drying for 30 minute, finally dry in a black particles have disappeared.
steam oven for 2 hour and weigh the residue. Calculate the The ignition was conducted in a place protected from air
percentage of water-soluble extractive with reference to the currents. The crucible was allowed to cool, and a few drops of
air-dried drug. concentrated sulfuric acid were added and heated. Ignited as
before, allowed to cool, and weighed. The operation was
Petroleum ether - soluble extractive value repeated until two successive weighing does not differ by
Macerate 5 g of the air-dried drug, coarsely powdered, with more than 0.5 mg. calculate the percentage of Sulphated ash
100 ml of petroleum ether of the specified strength in a close with reference to the air dried drug [12].
flask for 24 hour, shaking frequently during the first 6 hour
and allowing to stand for 18 Hour. Thereafter, filter rapidly Loss on Drying (LOD) by Gravimetric Method
talking precaution against loss of petroleum ether, evaporate Weigh about 1.5 g of the powdered drug into a weighed flat
25 ml of the filtrate to dryness in a tarred flat-bottomed and thin porcelain dish. Dry in the oven at 100˚c or 105˚c,
shallow dish, dry at 105 °C and weigh. Calculate the until two consecutive weighing do not differ by more than 0.5
percentage of petroleum ether-soluble extractive with mg. Cool in a desiccator and weigh. The loss in weight is
reference to the air-dried drug. usually recorded as moisture.

Ethyl acetate - soluble extractive value Results and Discussion

Macerate 5 g of the air-dried drug, coarsely powdered, with Extractive Value: The percentage (%) yield obtained after
100 ml of ethyl acetate of the specified strength in a close successive extraction of Blepharis repens (Vahl.) Roth, was
flask for 24 hour, shaking frequently during the first 6 hour found to be:
and allowing to stand for 18 hour. Thereafter, filter rapidly
talking precaution against loss of ethyl acetate, evaporate 25 Extractive Value
ml of the filtrate to dryness in a tarred flat-bottomed shallow Extracts%yield Extracts%yield w/w
dish, dry at 105 °C and weigh. Calculate the percentage of Sr.no. Extractives
w/w (Test) (Reference)
ethyl acetate-soluble extractive with reference to the air-dried Alcohol soluble
drug. 1 7.5% 5.3%
extract
Water soluble
2 5% 4.5%
Ash Value extract
Total ash Petroleum ether
3 4.5% 1.24%
Take about 2g, accurately weighed, of the ground drug in a soluble extract
tarred platinum or silica dish previously ignited and weighed. Ethyl acetate
4 4% 2.1%
Scatter the ground drug in a fine even layer on the bottom of soluble extract
The Extracts% yield of test was found to be greater than Extracts%
the dish. Incarnated by gradually increasing the heat-not
yield of reference [7].
exceeding dull red heat- until free from carbon, cool and
weigh.
Ash Value: The% ash value of Blepharis repens (Vahl) Roth,
If a carbon free ash cannot be obtained in this way, exhaust
was found to be:
the charred mass with hot water, collect the residue on an ash
less filter paper, increate the residue and filter paper, add the

Ash Value
Sr.no. Ash value % Ash value
1 Total Ash 8.5%
2 Sulfonated Ash 12%

Loss on drying (LOD): The% LOD of Blepharis repens (Vahl) Roth, was found to be:

Loss on drying (LOD)

Sr.no. Loss on drying (LOD) % LOD
1 By Gravimetric Method 83.34%

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Chemical Tests for Alcohol Soluble Extract

Test for Carbohydrates
Test for Reducing Sugars

Test for Reducing Sugars

Sr.no Test Observstion Inference
Molisch’s test:-To 2-3 ml aqueous extract, add few drop of alpha-napthol solution in Violet ring is formed at the Test was
A
alcohol, shake and add conc. sulphuric acid from side of test tube. junction of two liquids. positive
Fehling’s test: Mix 1 ml Fehling’s A and 1 ml Fehling’s B solution, boil for one minute, First yellow, then brick red ppt. Test was
B
Add equal volume of test solution. Heat in boiling water bath for 5-10 min. is observed. positive
Benedict’s test: Mix equal volume of Benedict’s reagent and test solution in test tube. Heat Test was
C solution appears green
in boiling water bath for 5 min. positive

Test for Monosaccharide

Sr.no Test Observation Inference

Barfoed’s test: Mix equal volume of Barfoed’s reagent and test solution. Heat for 1-2 min. In Red ppt was not Test was
A
boiling water bath and cool. observed. negative

Test for Tannins

Sr.no Test Observation Inference
Gelatin test: To a solution of tannin, aqueous solution of Gelatin
A White buff coloured ppt was observed. Test was positive
and sodium chloride were added.
B Ferric chloride: Drug solution + 5% ferric chloride. Deep blue-black colour was not observed. Test was negative
C Acetic acid: Drug solution + acetic acid solution. Red coloured solution was not observed. Test was negative
D Bromine water: Drug solution + Bromine water. Decolouration of Bromine water was not observed. Test was negative

Test for Alkaloids

Sr.no Test Observation Inference
A Dragendroff’s test: To 2-3 ml filtrate, add few drops Dragendroff’s reagent. Orange brown ppt. was found. Test was positive
B Wagner’s test: 2-3 ml filtrate with few drops Wagner’s reagent. Reddish brown ppt. was observed Test was positive
C Hager’s test: 2-3 ml filtrate with Hager’s reagent. Yellow ppt. was observed. Test was positive

Test for Flavonoids

Sr.no Test Observstion Inference
A Lead acetate test: To small quantity of residue, add lead acetate solution. Yellow colored ppt was observed. Test was positive
B Sulphuric acid test: solution + sulphuric acid (66% or 80%). Deep yellow solution was not observed. Test was negative

Test for Saponin Glycosides

Sr.no Test Observation inference
A Foam test: Shake the drug powder vigorously with water. Persistent foam was observed. Test was positive

Test for Steroids

Sr.no Test observation Inference

Salkowaski reaction: To 2 ml of extract, add 2 ml chloroform and 2 ml Chloroform layer and acid layer does not show Test was
A
conc. H2SO4. Shake well. any florescence. positive

Test for Triterpenoids

Sr.no Test observation Inference

A Test solution + 5 mL conc. Sulphuric acid from side of test tube. Greenish blue colour was observed. Test was positive

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Chemical Test for Various Extractives

Sr.no Tests Petroleum ether Water Ethyl acetate
Carbohydrates
a) Molisch’s test + + +
1 b) Fehling’s test + + -
c) Benedict’s test - + -
d) Barfoed’s test - - -
Tannins
a) Gelatin test + + +
2 b)Ferric chloride - - -
c)Acetic acid - - -
d)Bromine water + - -
Alkaloids
a) Dragendroff’s test + + -
3
b) Wagner’s test - + +
c) Hager’s test - + -
Flavonoids
4 a) Lead acetate test + + +
b) Sulphuric acid test + + +
Steriods
5
a) Salkowaski reaction + + +
Saponin glycosides
6
a) Foam test + + -
7 Triterpenoids + + -

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