IN VITRO MORPHOGENIC RESPONSE OF

DIFFERENT PARTS OF CARDIOSPERMUM

HALICACABUM LINN. (BALLOON VINE) –
PLANT WITH TREMENDOUS THERAPEUTIC
POTENTIALS

BY
Dr. ICHHA PURAK & Dr. ANITA MEHTA
DEPARTMENT OF BOTANY, RANCHI WOMEN’S COLLEGE,
RANCHI
Email : ichhapurak@yahoo.com , purak.ichha@gmail.com &
anitamehta.rwc@gmail.com
 ABSTRACT
Cardiospermum halicacabum Linn. belonging to family Sapindaceae is commonly
known as Balloon Vine due to inflated membranous balloon like tri-celled ridged fruits
having black seeds with prominent white heart shaped scar. It is recognized as a medicinal
plant of repute in Ayurvedic and Homoeopathic mode of treatment under different names
such as Jyotishmati, Kanphuti, Lataphatkari, Indravalli, Heart pea, Love- in-a -puff, Heart
seed, Winter cherry etc. The plant possesses active ingredients viz. alkaloids, flavonoids,
proanthocyanide, cyanolipids , glycosides, saponins, tri-terpenes, steroids ec . It is used to
cure rheumatism, diarrhoea, chronic bronchitis, nervous diseases, stiffness of limbs, snake
bite, pulmonary troubles, itchy skin, problems in menstrual cycle, gastric ulcer, earache ,
eyesore and piles.

Standardization of protocol was undertaken for achieving callus mass from different parts
of Cardiospermum halicacabum. Different explants responded differently to chemicals and
media they had been subjected.MS Medium supplemented with low concentration of 2,4
-D had a tendency to regenerate into shoot and complete plantlet but high concentration
induced callus formation as well as proliferation . Green embryogenic callus from leaf
explants was observed in media containing 2,4-D. Callus tissue is good source of genetic
and karyotypic variability so variants can be regenerated from these genetically variable
cells. Callus culture is useful for obtaining commercially important secondary
metabolities. Several biochemical assays can be performed from callus culture.
 INTRODUCTION
BRIEF MORPHOLOGICAL
DESCRIPTION

Cardiospermum halicacabum Linn. belonging to family Sapindaceae is commonly

known as Balloon Vine due to inflated balloon like tri-celled ridged fruits having black
seeds with prominent white heart shaped scar. Cardiospermum is widely distributed in
tropical and sub-tropical Africa and Asia.

In India it is widely distributed in tropical and subtropical plains.The plant climbs with
tendrils and needs some form of support. Leaves are deeply cut trifoliate, up to 4
inches long, with highly lobed leaflets. petioles 3-4 cm; leaflets subsessile; blades thinly
papery, margin sparsely serrate, apex acuminate .
Flower stalk has a pair of axial spiral tendrils at the base . The
bisexual flowers are white in colour and possess 4 unequal sepals, 4
narrowly elliptical white petals ,8 stamens ( 4 larger and 4 shorter),
slightly longer than petals; single carpel with 3 lobed ovary and tri-fid
stigma.

Each locule of ovary has one black seed with white heart shaped scar.
Capsules green in the beginning turns brown on ripening Disc of two
glands present below ovary. The fruit appears Balloon like due to
inflated membranous seed cover.
 MEDICINAL IMPORTANCE
OF CARDIOSPERMUM

It is recognized as a medicinal plant of repute in Ayurvedic and Homoeopathic mode

of treatment under different names such as Jyotishmati, Kanphuti, Lataphatkari,
Indravalli, Heart pea, Love- in-a -puff, Heart seed, Winter cherry etc.

It is used to cure rheumatism, diarrhoea, chronic bronchitis, nervous diseases,

stiffness of limbs, snake bite, pulmonary troubles, itchy skin, problems in menstrual
cycle, gastric ulcer, earache , eyesore and piles etc. Datta et al (2010 ).

Indian system of medicine also recommends Cardiospermum halicacabum leaves for

rheumatism, stiffness of limbs,chronic bronchitis and snakebite ( Chopra,1980,1981 )
Cardiospermum is an active ingredient in creams and lotions for dermatitis, eczema,
and psoriasis.
 ACTIVE INGREDIENTS

The plant possesses active ingredients viz. alkaloids, proanthocyanide,

cyanolipids , glycosides, saponins, steroids etc . Leaves contain b-
sitosterol, quebrachitol, saponin, oxalic acid and Apigenjn,
proanthocyanidin,stigmatosterol Dass(1966), Satyavati (1955). Rao et al
2006 mentioned antidiarrhoeal activity of the extracts of Cardiospermum
halicacabum due to presence of phytochemical constituents such as
sterols, tannins, flavonoids and triterpenes.
 THERAPEUTIC APPLICATIONS

Leaves are crushed and made into a tea, which cures itchy skin. Salted leaves are used
as a poultice on swellings.

Young leaves can be cooked as vegetables.

The leaf juice has been used as a treatment for earache . Nadkarni (1976),Asha et al
(1999 ),Gopalkrishnan et al (1976),Sadiqwe et al (1987),Joshi et al (1981).

It is used in nervous diseases and in dropsy.

The fresh decoction of leaves is given to asthmatic patients while its powdered form is
used to heal wounds.

Varier (1993) reported the sedative actions of plant on central nervous system probably
due to presence of hydrocynic acid.
 MODE OF APPLICATION AND
DOSE

It is used for medicinal purposes as tea,Decoction,Juice, paste,

Poultice,oil, chutney, tooth brush etc.

As Decoction of leaves/root, a teaspoon at a time should be taken

twice a day

Leaves and young shoots are edible

 CONSERVATION OF IMPORTANT
MEDICINAL PLANT

Pharmaceutical companies depend largely upon materials procured from

naturally occurring stands that are being rapidly depleted. Plant tissue culture is
an alternative method of propagation based on totipotent nature of cells. It
plays a key role in large scale multiplication of medicinally important plant
species (Pattnaik and Chand 1996, Rout er al., 1999, 2000,Ahmad and
Anis,2007 ) For enhancement of secondary metabolite content through genetic
transformation or by manipulating the constituents of growth media ,an efficient
in-vitro regeneration protocol is necessary. In-vitro mass propagation obviousky
has great potential to fulfill the immediate requirement of plant based
pharmaceutical industries.
.The regeneration of plants from callus culture has proved very useful
commercially. Keeping these things in mind the present investigation was
undertaken. Micropropagation plays a distinct role in conservation of
species, especially those having pharmacological value. Most of the
pharmaceutical plants are collected from the wild and very few from the
cultivated stands. In many cases, the entire plant is severed during
harvesting. To overpass this insufficiency and for safeguarding the species
from extinction, it is urgent to develop protocols which are cost effective and
be utilized for scaling-up the same (Bajaj et al. 1988)
Conventionally the plant is propagated through seeds but is not feasible due to low

germination rate ( 35-40 % ) , low viability and delayed rooting by seedlings Kirtikar

and Basu (1935 ) There are few earlier reports on regeneration of Cardiospermum

halicacabum Babber et al (2001),Thomas and Maseena (2006), Girish et al,( 2008 )

and Jahan and Anis (2009 ) In the present investigation an attempt has been made to

study micropropagtion of this species. The aim of this Study was to initiate and

maintain the calli produced from tissue of various plant parts ( leaf,stem

internode,node,flower,seed ).
 MATERIAL AND METHODS

Culture medium
MS ( Murashig and Skoog,1962 ) basal medium was used in the present investigation .
Concentrated stock solution of major and minor salts, iron source, vitamins and amino
acids were prepared separately and stored under refrigeration. Required volume of stock
solution was pipetted out during media preparation and 0.3% sucrose was added. The
chemicals used for preparing media were of analytical grade from Loba, Merck and
Sigma. Medium was homogenized by boiling and continuous stirring with pH of medium
adjusted upto 5.8 before adding 0.8% agar by using 0.1 N NaOH or 0.1N HCL.Desired
concentration of growth regulators was added and mixed properly. 15-20 ml medium was
poured into culture tubes ( 15x2.5cm ) which were washed thoroughly rinsed in distilled
water and oven dried.Sterilizarion of medium was done for 15- 20 minutes at pressure 1.1
kg/cm2 at temperature 121ºC .Tubes containing sterilized medium were left in tilted
position to prepare slants in air conditioned room.
Preparation of Explant(Culture material )

Some seeds of Cardiospermum halicacabum were soaked in water after removing the black

covering by scraping to facilitate in-vitro germination.Shoots bearing leaves were

collected from plants growing in garden of P P Compound area of Ranchi. Different

vegetative parts such as stem node,internode,leaf, flower etc which were taken as expant

after thoroughly washing with tap water and were treated with Cetavelon (1:100)

solution for about 5 minutes and again washed with running tap water.The explants were

then surface sterilized in 0.2% of HgCl2 by immersing for 2-3 minutes and then were

rinsed in double distilled water. These sterilized stem and leaves were then cut into

pieces of about 1.0-1.5cm and made ready for inoculation.

 INOCULATION

The explants were in-oculated into test tubes containing sterilized media under Laminar

Air Flow cabinet.Transfer area was sterilized with UV light and by swabbing the foor

surface with 95% ethyl alcohol.Inoculation tools were flamed before transferring the

explants. Each experiment with 10 culture tubes was repeated 4 times and were

observed every day for their morphological response.

 RESULT AND DISCUSSION

In the present investigation different explants responded differently to chemicals and

media they had been subjected. All types of explants showed initial hypertrophy. Low

concentration of 2,4-D ( 0.05 mg /l to 0.5 mg/l ) had a tendency to regenerate into

shoot or complete plantlet.But high concentrations of 2,4-D (2.5-5.0mg/l) induced

callus formation as well as its proliferation.

Green embryogenic callus was also observed from leaf explants in medium

containing 2.5mg/l 2,4-D. (Fig . 1 ) after 4 weeks of inoculation. White fragile

callus was observed in 1.5 months old culture obtained from leaf explants.( Fig. 2 )

in medium supplemented with 2.5mg/l 2,4-D.These calli were about 1.5 cm in

diameter. Nodulated and compact calli were induced in 1.5 months old stem

culture in medium containing 2.0-2.5 mg/l 2,4-D.(Fig 3 & 4 )

1 2

3 4
Fig. 1 1.5 months old leaf culture on MS Medium
containing 2,4-D ( 2.5 mg/l ) showing green
embryogenic callus.

Fig. 2 1.5 months old leaf culture on MS Medium

containing 2,4-D ( 2.5 mg/l ) showing white
Fragile callus.

Fig.3 45 days old stem culture on MS medium containing

2,4-D ( 2.0 mg/l ) showing nodulated and compact
callus

Fig. 4 1.5 months old stem culture on MS Medium

supplemented with 2.5 mg/l 2,4-D
showing compact and nodulated callus
Callus tissue is good source of genetic and karyotypic variability so variants can be

regenerated from these genetically variable cells. Callus culture is useful for obtaining

commercially important secondary metabolities. Several biochemical assays can be

performed from callus culture.

Such callus formation had been reported previously by Girish et al (2008) who

employed MS medium supplemented with auxins viz. 2,4-D,NAA IAA and cytokinins

viz. BAP and Kn alone in different concentrations..

A protocol for rapid micropropagation of C. halicacabum through plant

regeneration from leaf and nodal explant derived calli has been developed by

Thomas and Maseena (2006). Nodal and leaf explants were cultured on MS

medium supplemented with 0.5-9 μM 2,4-D. Highesr frequency of callus formation

was recorded with 5 μM 2,4-D.These calli on subculturing with kinetin and IAA

resulted shoot regeneration.Rooting of shoots was recorded when subjected to MS

medium supplemeted with 2.5 μM IBA

Babber et al (2001) were able to produce calli from different explants of

Cardiospermum halicacabum as cotyledon,hypocotyls,leaf,internode and node in MS

medium supplemented with BAP and NAA.Maximum number of shoots were produced

on subculturing calli on MS and BAP ( 17.8 μM ).

Recently Jahan and Anis (2009) developed a simple,rapid and efficient protocol for

microprogation of this plant via axillary bud multiplication by using nodal segments and

employing BA(Benzyladenine ), Kn (Kinetin), TDZ(Thidiazuron ) and 2-iP ( 2 –

isopentenyladenine. Multiple shoots differentiated directly without callus mediation

within 4 weeks.
As Cardiospermum halicacabum, a noxious weed owes tremendous therapeutic

potentials so is being exploited very frequently from its wild and native habitats. As it is

not a cultivated plant may face danger of extinction. Keeping this scenario in view it is

advisable to develop protocols for rapid in- vitro regeneration of this species through

callus as well as multiple shoot formation.

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