International Journal of Research in Pharmaceutical and Biomedical Sciences

ISSN: 2229-3701

______________________________________________________________________Research Paper

In Vitro Antioxidant and Antidiabetic activity of Asystasia gangetica

(Chinese Violet) Linn. (Acanthaceae)
N. V. L. Suvarchala Reddy*, Sneha J. Anarthe and N. M. Raghavendra
Gokaraju Rangaraju College of pharmacy, Bachupally, Hyderabad, A. P., India
__________________________________________________________________________________________
ABSTRACT
The present investigation evaluates the in vitro antioxidant and in vitro antidiabetic activity of methanolic
extract of leaves of Asystasia gangetica in various models. Besides, total phenolic was tested using Folin
Ciocalteau reagent. The DPPH radical scavenging of methanolic extract has IC50 value 179.67 g/ml and
reducing power of the extract was studied according to the reaction of Fe+3 to Fe+2. The reducing power of the
extract increased with the increasing amount of the concentration. The methanolic extract showed
concentration dependant -glucosidase (IC50 - 325g/ml) and -amylase (IC50 -3.75 g/ml) inhibitory activity.
Hence -glucosidase and -amylase enzyme inhibition may be the possible mechanism for the plant to exert
antidiabetic activity and considered as a potential candidate for the management of type-II diabetes mellitus.
The in vitro studies clearly indicate that the methanol extract of leaves of Asystasia gangetica has significant in
vitro antioxidant and -glucosidase and -amylase enzymes inhibitory activity.
Key words: Asystasia gangetica, in vitro antidiabetic activity and in vitro antioxidant activity
INTRODUCTION
Asystasia gangetica (L).T. (Chinese violet) is a
rapidly growing perennial shrubby herb mainly
distributed in north India, which grows to 10 m height,
at an altitude 300 m1 neutralized in some waste areas2.
Leaves are opposite petioles, flowers are pale purple
blue to violet or lime white in colour, capsules are
2.5-3.5 cm long with wide base and the seeds are 5
mm in diameter. It is mainly used in mild
hypoglycaemia, anticancer against epidermoid
carcinoma of nasopharynx. The juice of the plant is
also used as an anthelmintic3. It is used in swelling
and rheumatism, as a remedy for gonorrhea and ear
disease. It is used as folk remedy for the treatment of
diabetes mellitus in parts of south India4. It is
evaluated for anti-asthmatic activity5. Asystasia
gangetica reported to contain biologically active
substances such as carbohydrates, proteins, alkaloids,
tannins, steroidal aglycones, saponins, flavonoids and
triterpenoids. Study was undertaken to investigate the
in vitro antioxidant and antidiabetic activity of leaves
of Asystasia gangetica.
EXPERIMENTAL
Plant material and preparation of extract
Leaves of Asystasia gangetica were collected
_____________________________
*Address for correspondence:
E-mail: sneha.pharma@yahoo.co.in

Vol. 1 (2) Oct Dec 2010

from Siruvani forest Coimbatore district. The plant

was authenticated by Dr. P. Venu joint director,
botanical survey of India, Tamil nadu (voucher
specimen no BSI/SC/ 5/23/05-06/Tech-538). The airdried leaves of Asystasia gangetica were pulverized
and the powdered material was extracted with
methanol (70 %) by cold maceration. The extract was
concentrated on a rotary vacuum evaporator, which
gave a greenish-brown yield (3.65% w/w). The
proximate phytochemical analysis of methanol
extracts shows presence of flavonoids, proteins and
carbohydrates, alkaloids, tannins, saponins6.
Chemicals used
Acarbose (Bicon Ltd), -glucosidase (SRL),
maltose (Loba cheme), Glucose assay reagent
(Agappe Diagnostics), -amylase (SRL), and potato
starch (S.D. Fine-Chem).
DETERMINATION OF DPPH RADICAL
SCAVENGING ACTIVITY 7
1 ml different conc. of extract solution and
standard were taken in different vials. To this 5 ml of
methanolic solution of DPPH was added, shaken well
and mixture was incubated at 37 C for 20 min.
Measure the absorbance against methanol as blank at
517 nm. Take absorbance of the DPPH as control,
Percent antiradical activity can be calculated by using
following formula
Control Abs- sample A
% Antiradical activity =
100
Control Abs.

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International Journal of Research in Pharmaceutical and Biomedical Sciences

REDUCING POWER ASSAY8

1ml of different concentrations of extract solution
was mixed with 2.5 ml phosphate buffer and 2.5 ml
of potassium ferricyanide. The mixture was incubated
at 50 C for 20 minutes 2.5 ml of TCA was added to
the mixture, which was then centrifuged at 3000 rpm
for 10 minutes 2.5 ml of upper layer solution was
taken and mixed with 2.5 ml distilled water and 0.5
ml of ferric chloride solution and the absorbance was
measured at 700 nm. Increased absorbance of the
reaction mixture indicated increased reducing power.
IN VITRO INHIBITION -GLUCOSIDASE9
The enzyme - glucosidase inhibitory activity is
determined by incubating solution (0.1 ml) of an
enzyme preparation with 0.2 M Tris buffer, pH 8.0
(1.0ml) containing various concentrations of extract
at 37 C for 60 minutes by using glucose as working
standard. The reaction mixture is heated for two
minutes in boiling water bath to stop the reaction. The
amount of liberated glucose is measured by glucose
oxidation method. (Prashanth D, 2001) (Assay
condition 37C0.1C, pH-8.0; O.D at 540 nm).
(Enzyme activity of control Enzyme activity of extract)
% inhibition =------------------------------------------------------ 100
Enzyme activity of control
IN VITRO INHIBITION OF - AMYLASE10
A starch solution (0.1% w/v) was obtained by
stirring 0.1g of potato starch in 100ml of 16 mM of
sodium acetate buffer. The enzyme solution was
prepared by mixing 27.5mg of -amylase in 100 ml
of distilled water. The colorimetric reagent is
prepared by mixing sodium potassium tartarate
solution and 3, 5 di nitro salicylic acid solution 96
mM. Both control (Acarbose) and plant extracts were
added with starch solution and left to react with amylase solution under alkaline conditions at 25 C.
The reaction was measured over 3 minutes. The
generation of maltose was quantified by the reduction
of 3, 5 dinitro salicylic acid to 3-amino-5- nitro
salicylic acid. This reaction is detectable at 540 nm.
(Temperature 25C0.1 C, pH 4.8; O.D. at 540 nm).
(Maltose) test
% Reaction = --------------------------- 100
(Maltose) control
% Inhibition = 100- % reaction SD

TOTAL PHENOLIC CONTENT11

Total phenolic content of Asystasia gangetica
extract was measured by Folin Ciocalteau reagent
method. In this method, the blue colour formed due to

Vol. 1 (2) Oct Dec 2010

ISSN: 2229-3701

the polyphenol present in the extract was measured at

760 nm using UV spectrophotometer and results were
expressed as g/100g of gallic acid equivalent.
RESULTS AND DISCUSSION
The methanolic extract demonstrates potent
antioxidant activity in different in vitro models. The
DPPH radical scavenging of methanolic extract has
IC50 value 179.6 g/ml which is compared with
ascorbic acid as a standard (Table 1). In addition to
this the methanolic extract also possesses potent
reducing power which is compared with butylated
hydroxyl anisole (Table 2). The methanolic extract
found to contain a noticeable amount of total phenol.
The total phenolic content of Asystasia gangetica was
found to be 0.902 mg/ml, which play major role in
controlling antioxidants 12 . The result of this study
shows that the methanolic extract can be used as
easily accessible source of natural antioxidants and as
a possible food supplement or in pharmaceutical
industry. The in vitro -glucosidase inhibitory studies
demonstrated that methanolic extract of Asystasia
gangetica had -glucosidase inhibitory activity. The
percentage inhibition at 0.2, 0.4, 0.6, 0.8 and 1 mg/ml
concentration showed a concentration dependant
reduction in percentage inhibition (Table 3). Thus the
highest concentration of 1 mg/ml tested showed
maximum inhibition of nearly 91.9 %. The
percentage inhibition varied from 46-91 %. The 50 %
inhibitory concentration of methanolic extract of
Asystasia gangetica was found to be 325 g/ml
which is compared with the standard acarbose having
IC50 value 0.65 g/ml.
The in vitro -amylase inhibitory studies
demonstrated that methanolic extract of Asystasia
gangetica had -amylase inhibitory activity. The
percentage inhibition at 1, 2, 3, 4 and 5 mg/ml
concentration showed a concentration dependant
reduction in percentage inhibition (Table 4). Thus the
highest concentration of 5 mg/ml tested showed
maximum inhibition of nearly 72.1%. The percentage
inhibition varied from 4-72 %. The 50 % inhibitory
concentration of methanolic extract was found to be
3.75 g/ml which is compared with standard drug
acarbose having 50 % inhibitory concentration 92
g/ml. Thus, data presented here indicate that
methanolic extract of Asystasia gangetica possesses
significant in vitro antidiabetic activity. The
mechanism by which Asystasia gangetica exerted
action may be due to its action on carbohydrate
binding regions of - glucosidase enzyme, - amylase,
endoglucanases that catalyse hydrolysis of the
internal -1, 4 glucosidic linkages in starch and other
related polysaccharides have also been targets for the
suppression of postprandial hyperglycemia. This
enzyme is responsible in hydrolyzing dietary starch
into maltose which then breaks down to glucose prior
to absorption. Since -amylases play an important
role in starch break down in human beings and
animals, the presence of such inhibitors in food stuffs

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International Journal of Research in Pharmaceutical and Biomedical Sciences

ISSN: 2229-3701

may be responsible for impaired starch digestion13, 14.

-amylase inhibitor may be of value as novel
therapeutic dietetic agents.15 In conclusion, data
presented here rationalize the methanolic extract have
potential to emerge as new remedy for treatment of
type-II diabetes mellitus.

BHA

10

0.07160.0017

BHA

30

0.34760.0014

BHA

40

0.55160.0014

BHA

50

1.010.00118

Acarbose-like drugs, that inhibit glucosidase present in the epithelium of small

intestine, have been demonstrated to decrease postprandial hyperglycemia, 16 and improve impaired
glucose metabolism without promoting insulin
secretion in NIDDM patients 17. These medications
are most useful for people who have just been
diagnosed with type-II diabetes and who have blood
glucose levels slightly above the level considered
serious for diabetes. They are also useful for people
taking sulfonylurea medication, who need an
additional medication to keep their blood glucose
level within safe range. Therefore, the retardation and
delay of carbohydrate absorption with a plant-based
-glucosidase inhibitor offers a prospective
therapeutic approach for the management of type-II
diabetes mellitus 18.

AGM

25

0.040.0011

AGM

50

0.03690.00176

AGM

100

0.03860.00033

AGM

200

0.03460.00033

Table 1: Anti-radical activity of ascorbic acid and

methanolic extract of Asystasia gangetica leaves.
S. no.

Control
(mcg)

Mean SEM

AGM- methanolic extract of Asystasia gangetica, BHA- Butylated

hydroxyl anisole as standard.

Table 3: Inhibitory activity of methanolic extract of

Asystasia gangetica and standard drug acarbose
against -glucosidase
Sample

Acarbose

%
Antiradical
activity

ASC

10

0.34110.00043

59.98

ASC

20

0.25080.000026

70.51

ASC

30

0.22730.00011

73.28

ASC

40

0.19540.00045

76.99

ASC

60

0.19490.04303

74.20

ASC

80

0.17050.04194

83.02

AGM

100

0.24710.000116

63.94

AGM

250

0.23650.000693

69.57

AGM

500

0.30660.00066

70.95

Asystasia
gangetica

(g/ml)

%
inhibition
(SD)

0.2

35.01.087

0.4

42.471.12

0.6

47.90.73

0.8

58.431.23

1.0

62.20.43

200

46.810.381

400

60.131.011

600

76.772.057

800

89.580.233

1000

91.190.806

Concentration

IC50

0.65g/ml

325 g/ml

Data expressed as mean SD, n=6.

ASC-Ascorbic acid, methanolic extract of Asystasia gangetica,

Absorbance of blank = 0.8512

Table 4: Inhibitory activity of methanolic extract of

Asystasia gangetica and standard drug acarbose
against -amylase.
Concentration
(g/ml)

% inhibition
(SD)

50

40.36.929

100

52.03.323

150

64.381.202

200

76.950.565

250

89.2651.279

Sample

Table 2: Reducing power assay of BHA and

methanolic extract of Asystasia gangetica leaves.
S. no.

Acarbose

Control
(mcg)

Mean SEM

Vol. 1 (2) Oct Dec 2010

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IC50

92
g/ml

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International Journal of Research in Pharmaceutical and Biomedical Sciences

Asystasia
gangetica

100

4.051.131

200

14.40.353

300

25.950.636

400

63.30.1414

500

72.12.545

3.75
g/ml

Data expressed as mean SD, n=6.

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