In-vitro Antioxidant Activity of Aqueous Extract of Physalis minima Linn.
Ashok
Kumar Gupta1, Shivalinge Gowda K.P.1*, Umashankar R.N.1,
Nandeesh R.2 and Sreedhar S.1
Sree Siddaganga
College of Pharmacy, B. H. Road, Tumkur-572 102, Karnataka, India.
ABSTRACT:
The antioxidant
activity is increasing as there is realization of that formation of reactive
oxygen species (ROS) and reactive nitrogen species (RNS) have been linked in
pathogenesis of several human diseases. Physalis
minima L. are
commonly known as Country gooseberry, which is widely used in the indigenous system
of medicine for the treatment of diuretic, fever and dropsy. The leaves aqueous
extract of the plant was studied for its invitro antioxidant activity by two
methods viz DPPH radical scavenging assay and Nitric oxide scavenging assay.
Its free radical scavenging activity was estimated by IC50 value and
the values at various concentrations 5 to 25mg/ ml. At 25mg/ ml DPPH radical
scavenging assay and Nitric oxide assay showed maximum inhibition 92.30% and
75.41% These results clearly indicate that aqueous extract of leaves of Physalis minima is effective in scavenging free radicals and has the
potential to be a powerful antioxidant.
KEYWORDS: Physalis minima., DPPH., Antioxidant activity, Nitric oxide.
INTRODUCTION
Recently, there
has been a surge in research on the potential role of antioxidants in the
treatment of atherosclerosis, heart failure, liver dysfunction,
neurodegenerative disorders, cancer, and diabetes mellitus1. Free radicals are reactive oxygen species (ROS)
generated in the body during normal metabolic functions or introduced from
environment. These species causes cellular damage by reacting with the various
biomolecules of body such as membrane lipids, nucleic acid, proteins and
enzymes. This damage is the major contributor of many disorders like Cancer,
Hepatopathy, Cardiovascular diseases, inflammation, diabetes mellitus, renal
failure and brain dysfunction. Body has itself antioxidant system which reacts
with reactive species and neutralizes them. This natural antioxidant system
include enzymes like catalase, superoxide dismutase and glutathione which
protect the body from free radical species and prevent oxidative stress.
Synthetic antioxidants like Butylated hydroxyl toluene and Butylated hydroxyl
anisole are carcinogenic in nature. So need of natural antioxidant arises2. In ayurvedic system of
medicine Physalis minima Linn. (Country gooseberry)
is an annual, herbaceous, upright plant that grows to a height of 15-45 cm. it
may be seen growing on the borders of cultivated fields and wastelands. The
plant distributed more or less throughout India, Baluchistan, Ceylon-
Afghanistan, tropical Africa and Australia3.
In India Physalis minima available in
the month of July-September. The whole plant of Physalis minima used in the various medicinal uses like the fruit
is considered tonic, diuretic, dropsy, urinary disease, purgative, gout4 and poultice of fruits
used for headaches, intestinal pains and used to cure spleen disorders.
Poultice of leaves oiled and heated, applied
to ulcer and gonorrhea5.
The roots are
used as vermifuge, fever, and decoction of roots also used for diabetes6. The drug is also used in
snake-poison, inflammation, and Scorpion- sting7. The other uses of Physalis
minima are laxative, expectorant or promoting the discharge of phlegm,
painful discharge of urine, ulcer, cough and bronchitis3. From the literature review Physalis minima possesses many Pharmacological activities like
Anti-inflammatory, analgesic and antipyretic6, Antifertility7,
Antimicrobial activity8
and new leishmanicidal9
has already been reported.
MATERIALS AND METHODS:
Plant material:
The fresh plant of Physalis minima was collected from the
outskirts of Tumkur, Karnataka, India. It was identified and authenticated by
Dr. Siddappa Botanist; a voucher specimen has been deposited at the department
of Pharmacology, Sree Siddaganga College of Pharmacy- Tumkur. Leaves were
separated from the plant and washed thoroughly several times with running tap
water the leaves were dried under shade, powder by a grinder and was stored in
an airtight container.
Extraction and preparation of test drug:
The powdered was weighed and
200 g of powder was subjected for aqueous extraction by using Soxhlet apparatus
by maintaining the constant temperature for 32-48 h. And 200g 0f powder of Physalis minima leaves were extracted
separately with chloroform (1.5 L) at respective boiling point of extract. The
extract was filtered using whatman filter paper and concentrated to dryness
under reduced pressure. The extract was stored in refrigerator.
In vitro antioxidant activities:
DPPH (Free radical) scavenging activity:
Free radical (DPPH) scavenging activity of Physalis
minima extract (5, 10, 15, 20 and 25mg/ ml) was estimated. Methanolic
solution of DPPH (0.3mM) produced a violet coloured which is stable at room
temperature. 1ml of DPPH solution was added to 2.5 ml of sample solution
containing different concentration of the test solution and allowed to react at
room temperature. After 30 min of reaction time, the absorbance was measured at
518 nm against appropriate blank and control. Ascorbic acid was used as
standard antioxidant1.
Nitric oxide (NO) scavenging activity:-
Sodium nitroprusside in aqueous solution at
physiological pH spontaneously generates nitric oxide which interacts with
oxygen to produce nitrite ions that can be estimated using Griess reagent2. The reaction mixture
(3ml) containing sodium nitropruside (10mM) in phosphate buffer saline and the Physalis minima extract (5, 10, 15, 20
and 25mg/ ml) was incubated at 25°C temperature for 150min. after incubation
1.5ml of the reaction mixture was removed and 1.5ml of the Griss reagent (1%
sulphanilamide, 2% orthophosphoric acid and 0.1% Napthylethyline diamine
hydrochloride) was added. The absorbance of the chromophore formed was read at
546nm. Ascorbic acid was used as standard antioxidant1.
Statistical analysis:
The results are presented as Mean ± SEM. Statistical analyses were carried out by
using with one-way ANOVA followed by Dunnett’s test at level of significance
P<0.05. All data were shown as the mean ± SEM. Statistical analysis was
performed using prism-5 statistical software (USA).
RESULTS
AND DISCUSSION:
Table-1: Effect of aqueous extract of Physalis
minima on DPPH free radical scavenging activity. (All results are expressed
as mean ± SEM; n=4)
|
Drug |
Dose mg/ml |
DPPH free radical scavenging (measured at 518 nm) |
% of Inhibition |
|
Control |
- |
1.820 |
|
|
AEPM |
5mg |
0.237 ± 0.0014 |
86.97% |
|
|
10mg |
0.214 ± 0.0009 |
88.24% |
|
|
15mg |
0.151 ± 0.0027 |
91.70% |
|
|
20mg |
0.143 ± 0.0009 |
92.14% |
|
|
25mg |
0.140 ± 0.0006 |
92.30% |
|
IC50 |
|
3.57 |
|
|
Ascorbic acid |
5mg |
0.074 ± 0.0017 |
95.93% |
|
|
10mg |
0.065 ± 0.0008 |
96.42% |
|
|
15mg |
0.063 ± 0.0018 |
96.53% |
|
|
20mg |
0.054 ± 0.0014 |
97.03% |
|
|
25mg |
0.043 ± 0.0011 |
97.63% |
|
IC50 |
|
3.12 |
|
Values are
expressed as mean ± SEM; n=10. Data analysed by One way ANOVA followed by Dunnett’s
test Significant relative to a= P < 0.05, b= P < 0.01, c= P
< 0.001
Table-2: Effect of aqueous extract of Physalis
minima on nitric oxide scavenging activity. (All results are expressed as mean
± SEM;; n=4)
|
Drug |
Dose mg/ml |
Nitric oxide scavenging (measured at 546nm) |
% of
Inhibition |
|
Control |
NA |
1.131 |
NA |
|
AEPM |
5mg |
--- |
--- |
|
|
10mg |
0.285 ± 0.0019 |
74.80% |
|
|
15mg |
0.284 ± 0.0009 |
74.88% |
|
|
20mg |
0.281 ± 0.0010 |
75.15% |
|
|
25mg |
0.278 ± 0.0009 |
75.41% |
|
Ascorbic acid |
5mg |
--- |
--- |
|
|
10mg |
0.089 ± 0.0008 |
92.13% |
|
|
15mg |
0.087 ± 0.0004 |
92.30% |
|
|
20mg |
0.068 ± 0.0004 |
93.98% |
|
|
25mg |
0.058 ± 0.0008 |
94.87% |
Values are
expressed as mean ± SEM; n=10. Data analysed by One way ANOVA followed by
Dunnett’s test Significant relative to a= P < 0.05, b= P <
0.01, c= P < 0.001
DPPH (Free radical) scavenging activity:
The 1, 1-Diphenyl-2-picrylhydrazyl (DPPH) radical
scavenging assay is the most widely used method for screening antioxidant
activity, since it can accommodate many samples in a short period and detect
active ingredients at low concentration.
The potential decrease in the concentration of DPPH radical is due to the
scavenging ability of AEPM and ascorbic acid showed the significant free
radical scavenging activity. In this method color change purple to yellow
color. The aqueous extract of leaves of Physalis
minima showed DPPH radical scavenging activity in a different
concentration. (Table-1)
Nitric oxide scavenging
activity:
AEPM showed moderately good nitric oxide scavenging
activity between 5 to 25 mg/ml. The percentages of inhibitions were increased
with increasing concentration of the extracts. Sodium nitro preside is known to
decompose in aqueous solution at physiological pH (7.2) producing nitric oxide
under aerobic conditions, nitric oxide reacts with oxygen to produce stable
products (nitrate and nitrite). This
leads to reduction of nitrite concentration in the assay media. (Table-2)
CONCLUSION:
On the basis of the results in the present study, it is
concluded that aqueous extract of Physalis
minima, which contains large amounts of tannins, exhibits high antioxidant
and free radical scavenging activities. These In-vitro assays indicate that
this plant extract is a significant source of natural antioxidant, which might
be helpful in preventing the progress of various oxidative stresses. However,
the components responsible for the antioxidative activity are currently
unclear. Therefore, further investigations need to be carried out to isolate
and identify the antioxidant compounds present in the plant extract.
ACKNOWLEDGMENTS:
Authors are grateful to Dr. S. Badami Director and
principal of Sree Siddaganga College of Pharmacy- Tumkur for providing
imminence guidance and required facility to complete this study.
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Received on 19.05.2010
Accepted on 12.06.2010
© A&V Publication all right reserved
Research J. Pharmacology and
Pharmacodynamics. 2(5): Sept.-Oct. 2010, 332-334