Antioxidant Activities of Muntingia calabura, Syzygium cumini, Ocimum
basilicum, and Eleutherine bulbosa using DPPH Method
Ani Haerani
1*
, Anis Yohana Chaerunisa
1
, Anas Subarnas
2
1 Department of Pharmaceutics, Faculty of Pharmacy, Universitas Padjadjaran, Sumedang, Indonesia,
45363
2 Department of Pharmacology and Clinical Pharmacy, Faculty of Pharmacy, Universitas Padjadjaran,
Indonesia 45363
Received: 15 May 2019 /Revised: 20 May 2019/Acepted: 25 May 2019/Published: 13 Jun 2019
ABSTRACT
Antioxidants are substances that can provide endogenous protection and exogenous oxidative stress
by capturing free radicals. Many plants are efficacious as antioxidants, namely plants that contain
polyphenols, especially flavonoids, so many are formulated as natural antioxidants. Plants such as
Muntingia calabura, Syzygium cumini, Ocimum basilicum and Eleutherine bulbosa contain polyphenol
compounds, especially flavonoids which are efficacious as natural antioxidants.This research aimed to
study antioxidant activity derived from some potential plants using the DPPH method by calculating the
IC50 value of each plant extract. This research method starts from the determination process to prove
the validity of the plants used, the extraction process using the maceration method with 70% ethanol
solvent, then the antioxidant activity of extracts from each plant was carried out using the DPPH method.
This research starts from the determination process to ensure the correctness of the plants used, then the
extraction process is carried out using the maceration method with 70% ethanol solvent. After that the
antioxidant activity was determined from the four plants using the DPPH method to see the strongest
IC50 value among the four plants. IC50 is the concentration of the sample to inhibit 50% of free radicals.
The results of IC50 values fromethanol extract of Muntingia calabura leaves, Syzygium cumini leaves,
Ocimum basilicum leaves and Eleutherine bulbosa bulbs, were 18.72; 63,84; 141.59 and 173.15 ppm.
Ethanol extract of Muntingia Calabura has a smaller IC50 value of 18.72 ppm which has a very strong
and most powerful antioxidant from the ethanol extract of Syzygium cumini, Ocimum basilicum and
Eleutherine bulbosa.
Keywords: Antioxidant, Muntingia calabura, Syzygium cumini, Ocimum basilicum, Eleutherine
bulbosa, DPPH Method
1. Introduction
Antioxidants defined as molecules having
ability to inhibit the oxidation of other molecules
mostly caused by free radicals, therefore can reduces
the damages due to oxygen [1,2,3]. Antioxidants
can be catagorized as either synthetic or natural
and both are included in formulations. Synthetic
antioxidants (e.g. butylated hydroxyanisole (BHA),
butylated hydroxytoluene (BHT), and propyl
gallate) has been largely used for many purposes
since the ease of production leading to lower prices.
However, high potential health risks following a
high consumption of synthetic antioxidants has been
reported by some studies. Despite the high demand
for synthetic antioxidants by market, concerns on
natural antioxidants have been increased in the last
few years and it is expected to continue and growing.
There are some reasons to explain this trend. Mostly,
the consumer prefer organic and natural products,
since they use less additives thus it is hoped to have
less side effects than synthetic ingredients[4].
Natural antioxidants has been used in the cosmetic
industries including a great number of substances and
extracts obtained from a variety of plants, grains and
fruits, either by reducing the skin oxidative stress or
protecting the skin from oxidative degradation [4].
Plants that are efficacious as antioxidants are
plants that contain carotenoids and polyphenols,
especially flavonoids which can be formulated as
natural antioxidants in oral dosage forms such as
vitamins and topicals for skin care products.
This research aims to determine the antioxidant
activity of Muntingia calabura leaves, Syzygium
cumini leaves, Ocimum basilicum leaves and
Eleuherine bulbosa bulbs since they contain
polyphenol compounds, especially flavonoids which
known to have efficacious as natural antioxidants.
Muntingia calabura L. contains phenolic acids
and flavonoids [5]. Syzygium cumini L. contains
anthocyanin, glucoside, isokuercin, kaempferol,
myrisetin and high total phenolic compounds
[6]. Ocimum basilicum L. contains apigenin and
rosmarinic acid which can be used as anti-radical
free [7]. Eleuherine bulbosa (Mill.) Urb contains
three groups of compounds, namely naphtalen,
naphtokuinon and anthraquinone [8].
Determination of antioxidant activity in this study
*Corresponding author, https://doi.org/10.24198/idjp.v1i2.21531
e-mail : anihaerani457@gmail.com (A. Haerani) 2019 Haerani et al
Vol 1, Issue 2, 2019 (57-61)
http://journal.unpad.ac.id/idjp
A. Haerani et al / Indo J Pharm 2 (2019) 57-61
58
using the DPPH method. The were conducted by
DPPH method is used to obtain IC50 values from a
plant extract. IC50is the concentration of the sample
to inhibit 50% of free radicals. A compound is said
to have very strong antioxidant activity if the IC50
value is less than 50 ppm, strong if the IC50 value is
50-100 ppm, moderate if it is 100-150 ppm and weak
if the IC50 value is 150-200 ppm [9].
2. Materials and Methods
2.1 Materials
Muntingia calabura leaves, Syzygium cumini
leaves, Ocimum basilicum leaves and Eleutherine
bulbosa bulbs from Manoko plantation, Lembang,
Bandung. Determination of plants is evaluated at the
Laboratory of Taxonomy, Department of Biology,
Faculty of Mathematics and Natural Sciences,
Padjadjaran University. The chemicals used in this
study include ethanol 96% p.a (Merck), DPPH (2,2-
diphenyl-1-pycryl-hydrazil) (Sigma), ascorbic acid
(Merck). Absorbance of sample is analyzed using
UV-Vis spectrophotometer (TECAN M200Pro).
2.2 Methods
2.2.1 Material collection and plant determination.
Plant material used was obtained from the
Manoko plantation, Lembang, West Java. Plants were
determined at the Taxonomy Laboratory, Biology
Department, Faculty of Mathematics and Natural
Sciences, Padjadjaran University.
2.2.2 Extraction
The extraction of Muntingia calabura leaf,
Syzygium cumini leaf, Ocimum basilicum leaf and
Eleutherine bulbosa bulbs plant were conducted
using maceration with 70% ethanol solvent for 3
times 24 hours. Liquid extract concentrated with a
rotary evaporator then steamed above a water bath
until constant weight of the extract. The yield of the
extract can be calculated by the formula :
2.2.3. In vitro Antioxidant Activity Test of Extract
a. Sample Preparation
Ethanol extracts were prepared with a stock
solution of 1000 ppm. 1000 ppm stock solution
is diluted to concentration 0, 5, 10, 15, 20 and 25
ppm for Muntingia calabura L., 0, 20, 40, 60, 80
and 100 ppm for Syzigium cumini L., 0, 50, 100,
150, 200 and 250 ppm for Ocimum basilicum L.
and Eleutherine bulbosa (Mill.).
b. Preparation of Comparative Solutions
Ascorbic acid was prepared with a solution of
100 ppm. Diluted a stock solution standard to
concentration 0, 1, 2, 3, 4, 5 ppm [10].
c. Preparation of DPPH solution (2,2-diphenyl-1-
pycryl-hydrazyl)
DPPH was weighed and dissolved in ethanol p.a
at a concentration of 160 ppm for immediate use
and maintained in low temperatures and protected
from light exposure[10].
d. Maximum Wavelength Determination
DPPH 0.2 mL solution was dissolved with 0.8
mL ethanol with a concentration of 250 ppm,
measured at 500-530 nm wavelength to obtain an
absorbance of ± 0.2-0.8[10].
e. Determination of IC50 with DPPH Method
The ethanol extract of Muntingia calabura leaves,
Syzygium cumini leaves, Ocimum basilicum leaves
and Eleutherine bulbosa bulbs were added 1 mL
DPPH for each concentration, then sonication and
incubated for 30 minutes at room temperature.
The absorbance was measured at 517 nm. The
inhibition percentage was calculated using the
equation [11]:
3. Results
3.1. Determination
The results of plant determination show Muntingia
calabura leaves belongs tothe Family: Muntingiaceae,
Genus: Muntingia, Species: Muntingia calabura L,
Indonesian name : Kersen. Syzygium cumini leaves
included in the Family: Myrtaceae, Genus: Syzygium,
Species: Syzygium cumini(L.) Skeels., Indonesian
name : Jamblang. Ocimum basilicum leaves
included in the Family: Lamiaceae, Genus: Ocimum,
Species: Ocimum basilicum L,., Indonesian name :
Kemangi. Eleutherine bulbosa bulbs included in the
Family: Tridaceae, Genus: Eleutherine, Species:
Eleutherine bulbosa (Mill.) Urb., Indonesian name :
Bawangdayak.
3.2. Extraction
Muntingia calabura leaves extraction (250 g)
was macerated with 70% ethanol solvent resulting
leaf extract of 78,836 g (rendement = 31,53 %).
Syzygium cumini leaves, Ocimum basilicum leaves
and Eleutherine bulbosa bulbs extraction (100 g) was
macerated with 70% ethanol solvent resulting 28,49;
19,94; 13,24 g (rendement% = 28,49; 19,94; 13,24
%) (Table 2 - 5 and Figure 1 – 4)
3.3 Antioxidant Activity
In this study, antioxidant activity was tested for all
four plants, using the DPPH method. The principle
of the DPPH method is based on measurements of
capture of antioxidant capacity. DPPH is a free
A. Haerani et al / Indo J Pharm 2 (2019) 57-61
59
radical is stable in purple, can be reduced by the
presence of antioxidant molecules resulting in
changes in color from purple to yellow.The results of
testing the antioxidant activity indicate IC50 values
can be seen in table 6.
Table 1. Phytochemical Screening Results of Etha-
nol Extracts of M. calabura leaves, S. cumini leaves,
O. basilicum leaves and E. bulbosa bulbs.
Parameter
Mc
Sc
Ob
Alkaloid
+
+
+
Flavonoid
+
+
+
Polifenol
+
+
+
Tanin
+
+
-
Saponin
+
+
+
Steroid & Terpenoid
+
+
+
Monoterpenoid &Seskui-
terpenoid
+
+
+
Kuinon
+
+
-
Note :
(+) : Identified
(-) : Not Identified
Mc : Muntingia calabura L.
Sc : Syzygium cumini L.
Ob : Ocimum basilicum L.
Eb : Eleutherine bulbosa
Based on the results of phytochemical screening
in the table 1. The four plants contain compound al-
kaloid, flavonoid, polifenol, monoterpenoid and ses-
kuiterpenoid.
Tabel 2. IC
50
of Muntingia calabura L.
Table 3. IC
50
of Syzygium cumini L.
Concentration (ppm)
Absorbance
% Inhibition
0
0,9244
0
20
0,76795
16,92449
40
0,6183
33,11337
60
0,47605
48,50173
80
0,3787
59,03288
100
0,1959
78,80788
Table 4. IC
50
of Ocimum basilicum L.
Concentration (ppm)
Absorbance
% Inhibition
0
0,8334
0
50
0,6431
22,83417
100
0,52245
37,31102
150
0,3534
57,59539
200
0,2598
68,82649
250
0,15415
81,50348
Table 5. IC
50
of Eleutherine bulbosa Merr.
Concentration (ppm)
Absorbance
% Inhibition
0
0,7756
0
50
0,6425
17,16091
100
0,4945
36,24291
150
0,40095
48,30454
200
0,3235
58,29036
250
0,2725
64,86591
Table 6. IC
50
Muntingia calabura leaves extract,
Syzygium cumini leaves, Ocimum basilicum leaves
and Eleutherine bulbosa bulbs
Plant Name
IC
50
(ppm)
Muntingia calabura
18,72
Syzygium cumini
63,84
Ocimum basilicum
141,59
Eleutherine bulbosa
173,15
4. Discussion
Phytochemical screening was carried out on
Muntingia calabura leaves extract, Syzygium cumini
leaves, Ocimum basilicum leaves and Eleutherine
bulbosa bulbs. Phytochemical screening aims to
determine and determine the secondary metabolites
contained therein[12]. The results of phytochemical
screening showed that all four plants contained al-
kaloid compounds, flavonoids, polyphenols, mono-
terpenoids and sesquiterpenoids. Compounds such as
flavonoids and polyphenols are compounds that have
Concentration (ppm)
Absorbance
% Inhibition
0
0,8199
0
5
0,69495
15,23966
10
0,58015
29,24137
15
0,4642
43,38334
20
0,3598
56,1166
25
0,3164
61,40993
A. Haerani et al / Indo J Pharm 2 (2019) 57-61
60
Figure 1. Curve calibration of Muntingia calabura
L.
Figure 2. Curve calibration of Syzygium cumini L.
Figure 3. Curve calibration of Ocimum basilicum L.
Figure 4. Curve calibration of Eleutherine bulbosa.
antioxidant activity. Based on the compound content
in it, the four plants have positive antioxidant activity
[13].
The anti oxidant activity test was carried out using
a UV-Viss pectrophotometer. This test was conducted
to determine thea bsorbance of the remaining DPPH
after adding the sample. If the DPPH solution is dis-
solved with a compound that has antioxidant activ-
ity, there will be a decrease in the value of DPPH
absorbance which is characterized by a change in
color from purple to yellow after incubation for 30
minutes.
This study uses the DPPH method to obtain IC
50
values from a plant extract. IC
50
is the concentration
of the sample to inhibit 50% of free radicals. A com-
pound is said to have very strong antioxidant activity
if the IC
50
value is less than 50 ppm, strong if the IC
50
value is 50-100 ppm, moderate if it is 100-150 ppm
and weak if the IC
50
value is 150-200 ppm (Zuhra et
al., 2008). Based on the results of the study of IC
50
Muntingia calabura leaf ethanol extract of 18.72 ppm
including very strong, the IC
50
Syzygium cumini leaf
value of 63.84 ppm was strong, IC
50
Ocimum basi-
licum leaf value was 141.59 ppm including moder-
ate and IC
50
Eleutherine bulbosa leaf value ie 173.15
ppm including weak. Based on IC
50
values produced
from the four plants, it can be seen that the ethanol
extract of Muntingia calabura has the strongest anti-
oxidant activity because it has a very high content of
phenolic compounds, especially flavonoids.
5. Conclusion
The result showed that ethanol extract of Muntingia
calabura produced an IC
50
value of 18.72 ppm so it
can be concluded that ethanol extract of Muntingia
A. Haerani et al / Indo J Pharm 2 (2019) 57-61
61
calabura leaf has the strongest and most powerful
antioxidant activity between Syzygium cumini, Oci-
mum basilicum and Eleutherine bulbosa.
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