ISSN 0975-2366

DOI:https://doi.org/10.31838/ijpr/2020.12.04.263
Research Article

Antioxidant and Sunscreen Activities from Blackberry

Extract in vitro using Spectrophotometry UV-Visible
NENI SRI GUNARTI1*, ANGGUN HARI KUSUMAWATI2, MAYA ARFANIA3, FARHAMZAH4,
IIN LIDIA PUTAMA MURSAL5
1-5
Universitas Buana Perjuangan Karawang.
*Corresponding Author
Email ID: 1neni.gunarti@ubpkarawang.ac.id, 2anggunhari@ubpkarawang.ac.id,
3
maya.arfania@ubpkarawang.ac.id, 4farhamzah@ubpkarawang.ac.id, 5iin.lidia@ubpkarawang.ac.id
Received: 06.04.20, Revised: 25.05.20, Accepted: 19.06.20

ABSTRACT
This research was conducted to determine the antioxidant and sunscreen activity of blackberry extract
(Rubusfruticosus). Antioxidant activity was tested using the 1.1 Diphenyl-2- picrylhidrazyl (DPPH) method
using UV-Visible spectrophotometry. Sunscreen activity is also carried out in vitro using UV-Visible
spectrophotometry. The results showed that Blackberry ethanol extract had weak antioxidant activity with
IC50 407.5175 ppm and erythema% sunscreen at concentrations starting from 0.75% including total Block,%
pigmentation at concentrations starting from 0.25% including total Block and SPF values above 30 (ultra) at
concentrations starting from 0.75%. This shows that blackberry extract has weak antioxidant activity but has
potential as a natural sunscreen ingredient.
Keywords: EkstrakEtanol Blackberry, Antioksidan, Tabir Surya.

INTRODUCTION directly or processed into jam, jelly, syrup and

Blackberry fruit (Rubus sp.) Contains anthocyanin wine. Blackberry fruit is known to have various
and phenolic compounds which have high pharmacological activities such as anticancer,
antioxidant activity (Verma et al. 2014). Research antimicrobial, antioxidant, antidysentery,
on ethanol extract of Strawberry Leaves shows the antidiabetic and antidiarrheal (Koczka,
antioxidant activity of the extract by reducing Stefanovits-bányai, and Prokaj 2018).
DPPH free radicals while also having sunscreen Anthocyanin is the dominant flavonoid compound
activity (Kusuma and Sukmawati 2016). This found in blackberries, this compound is a red-
shows the correlation between antioxidants and purple-black pigment-forming compound.
sunscreen activity. This statement is reinforced by Anthocyanins of Blackberry fruit that have been
research on antioxidant activity and sunscreen on identified include cyanidin3-glucoside, cyanidin3-
the extract of Musa acuminate L. fruit skin which galactoside, cyanidin3-xyloside, cyanidin 3-
shows the highest sunscreen activity found in dioxalyl-glucoside, cyanidin 3-routineoside,
ethanol extract which has the highest antioxidant cyanidin 3-sophoroside, cyanidin 3-xyloside,
activity (Alhabsyi and Suryanto 2014). cyanidin 3-dioxalyl-glucoside, cyanidin 3-
Antioxidants can protect the skin from the routineoside, cyanidin 3-sophoroside, cyanidin 3-
negative effects of free radicals that can cause xyloside, cyanidin 3-dioxalyl-glucoside, cyanidin
skin diseases. Types of antioxidants that can be 3-routineoside, cyanidin 3-sophoroside, cyanidin
beneficial to the skin are vitamin A, vitamin E, 3-xyloside, cyanidin 3-dioxalyl-glucoside,
carotenoids, beta-carotene, lycopene, cyanidin 3-routineoside, cyanidin 3-sophoroside,
polyphenols, flavonoids and lutein (Kusuma and cyanidin 3-xyloside, cyanidin 3-dioxalyl-
Sukmawati 2016). Based on this background glucoside, cyanidin 3-routineoside arabinoside,
Antioxidant and Sunscreen Activities from perlargonidin3-glucoside, cyanidin 3- (3-malonyl)
Blackberry Extract in vitro using glucoside, and cyanidin 3- (6-malonyl) glucoside.
Spectrophotometry UV-Visible. Cyanidin 3-dioxaloylglucoside,
azwitterionicanthocyanin is a unique compound
LITERATURE REVIEW owned by Blackberry (Kaume, Howard, and
Blackberry (Rubusfruticosus) is a plant of the Devareddy 2012).
Rosaceaefamily which contains high phenolic and Antioxidants are substances that are in low
vitamin compounds, a source of minerals and concentrations when compared to the substrate to
fiber. Fresh blackberries are usually consumed be oxidized, significantly slowing down or

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 Neni Sri Gunarti et al / Antioxidant and Sunscreen Activities from Blackberry Extract in Vitro Using
Spectrophotometry UV-Visible

inhibiting substrate oxidation (Halliwell and % transmisieritema =

Gutteridge, 1995). At one time, one antioxidant
molecule can react with a single free radical and % transmisipigmentasi =
can neutralize free radicals by donating one Sun Protection Factor (SPF) is a universal indicator
electron. The body naturally produces that explains the effectiveness of a UV protective
antioxidants (endogenous antioxidants) to product or substance, the higher the SPF value of
neutralize free radicals and protect the body from a sunscreen active product or substance, the
tissue damage, but this process is not 100% more effective it is at protecting the skin from the
effective if free radical production is too excessive adverse effects of UV rays. (Sharif, 2017).
and its effectiveness also decreases with age. SPF value is calculated using the Mansur method
Exogenous antioxidants obtained from food also (Yulianti, 2015):
play an important role to protect the body (1)
(Halliwell and Gutteridge, 1995). Dimana:
Sunscreen is one example of cosmetic ingredients EE = Efekeritemalspektrum
or preparations whose role is to protect the main I = Intensitascahayaspektrum
skin from the dangers of sunlight, especially Abs = Absorbansibahantabirsurya
ultraviolet (UV) (Adilah, 2017). Meanwhile, CF = Faktorkoreksi (= 10)
according to Sharif in 2017, sunscreen Nilai EE X I is constant and is shown in Table 1
preparations are cosmetics that are used for the below:
purpose of absorbing sunlight effectively,
especially in the ultraviolet wave area so that it Table 1: Normalized product function used in
can prevent the occurrence of skin disorders by SPF calculations
sunlight. Sunscreens can be made in various
dosage forms such as creams, lotions, and No. Wavelength(λ nm) EE X I
ointments. Based on its mechanism of action, the 1. 290 0,0150
active ingredient of sunscreen is divided into two, 2. 295 0,817
namely a physical blocking mechanism (reflecting 3. 300 0,2874
solar radiation) and a chemical absorbing 4. 305 0,3278
mechanism (absorbing solar radiation). 5. 310 0,1864
Mechanism of action Physical sunscreen reflects 6. 315 0,0839
ultraviolet radiation, its ability based on particle 7. 320 0,0180
size and layer thickness, can penetrate the dermis Total 1
to subcutaneous or hypodermic and is effective in
the UV-A, UV-B and visible light spectrum. While Table 2: Classification of Effectiveness of
the mechanism of action of chemical sunscreens Sunscreen Preparations Based on SPF Value
absorb ultraviolet radiation and convert it into a (Warnida and Nurhasnawati, 2017)
form of heat energy, it can absorb almost 95% of
Sunscreen Protection
UV-B radiation that can cause sunburn (erythema No. SPF value
Category
& wrinkles) (Anggraini, 2013). sunscreen
potential determined by In Vitro using data from 1. 6-10 Low protection
the transmittance measurement (% T) using a UV- 2. 15-25 Medium protection
Vis spectrophotometer. 3. 30-40 High protection
Sunscreen potential is determined based on 4. >50 Maximum protection
percent transmission of erythema (%Te), percent
transmission of pigmentation (% Tp) and The FDA requires that all sunscreens contain Sun
determination of SPF (Sun Protecting Factor) Protection Factor (SPF). The SPF range starts from
value. Determination of the percentage of 2 to more than 50, Sunscreens are recommended
erythema transmission and pigmentation was with at least SPF 15. The sunscreen SPF rating is
carried out with extract concentrations of 100, calculated by comparing the amount of time
150, 200, 250 and 300 ppm, then each needed to produce sunburn on the sunscreen
absorbance was measured using a UV-Vis protected skin with the amount of time needed to
spectrophotometer at wavelengths that could cause sunburn on unprotected skin (Lavi, 2013).
cause erythema of 292.5 - 317.5 nm and the
pigmentation is 322.5 - 372.5 nm. The category RESEARCH METHODOLOGY
of sunscreen activity is then assessed based on The tools used in this study are UV-Vis
the percentage of erythema and the percentage Spectrophotometer (Thermo Scientific 33-
of pigmentation calculated using the equation: PPPTS2017-L205-0006), rotary evaporator
(EYELA type N-1110), maserators, triangles,

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 Neni Sri Gunarti et al / Antioxidant and Sunscreen Activities from Blackberry Extract in Vitro Using
Spectrophotometry UV-Visible

beaker glass, waterbath, micro pipette, analytical 10 52,24766751

balance , measuring cups, measuring flasks,
vaporizer cups, drop pipettes, spatulas.
The research materials used were Blackberry fruit,
96% ethanol, methanol pa, Aluminum foil,
Aquadest, Vitamin C pa (Coated Ascorbic Acid
Type Ec, Code 0425117, Analysis 06202961),
DPPH (1,1-diphenyl-2- picrylhydrazyl) D9132 -
16.
The research procedure was carried out with the
stages of gathering raw materials of blackberries,
extracting blackberries using maceration method
with 96% ethanol solvent, then extracting the
extract using a rotary evaporator, then testing the
antioxidant activity and SPF of ethanol extracts of
blackberries.
Based on the measurement results of the
antioxidant activity of Blackberry ethanol extract
showed IC50 407.5175 ppm compared to the
antioxidant activity of vitamin C IC50 9.2744 ppm.
Linear regression curves can be seen in Figure 1.
Regression formula y = 0.128 x - 2.4475 with the
coefficient of determination (R2) 0.9762. IC50
values are weak because of > 200ppm
(Molyneux).

FINDINGS
The antioxidant activity of Blackberry ethanol
extract uses the DPPH method. The principle of
reducing free radicals with the DPPH method is
the reduction of DPPH solution in methanol to
non-vertical DPPH-H due to hydrogen donors
Fig. 1: Linear Regression Curves
from antioxidants. This process is characterized
by a change in DPPH color from purple to yellow
Determination of the sunscreen potential of
as measured by spectrophotometry at λmax 515-
Blackberry ethanol extract was done by In Vitro
517 nm.
using the UV-Vis spectrophotometry method at
DPPH free radical reduction test uses positive
concentrations of 0.25%, 0.5%, 0.75%, 1% and
control of antioxidants in the form of ascorbic
1.25% at wavelengths of 290-375 nm. The
acid. The results of the measurement of
potential of sunscreen is determined based on the
antioxidant activity with UV-Visible
calculation of the value of percent transmission of
spectrophotometry of Blackberry ethanol extract
erythema (%Te) and percent of pigmentation
can be seen from table 3 while the results of the
transmission (% Tp) and calculation of the SPF
measurement of comparative antioxidant activity
(Sun Protection Factor) value. The average
(Vitamin C) can be seen in table 4. percent transmission rate of erythema (%Te) at
concentrations of 0.25%, 0.5%, 0.75%, 1% and
Table 3: Results of IC50 Value of Antioxidant
1.25% can be seen in Table 6.
Blackberry Ethanol Extract
At ethanol extract concentrations of Blackberry
Concentration 0.75%, 1% and 1.25% showed the category of
% Inhibition IC50
(ppm) Total Block based on percent erythema
100 12.9771 transmission (Te%). The average value of percent
200 22.70992 transmission of pigmentation (%Tp) at
300 33.58779 407,5175 concentrations of 0.25%, 0.5%, 0.75%, 1% and
400 45.6584 1.25% can be seen in table 7. Based on the value
500 65.83969 of pigmentation transmission, Blackberry ethanol
extract starting from the smallest concentration
Table 4: Results IC50 Antioxidant Vitamin C p.a has shown Total Block activity. The average SPF
values at concentrations of 0.25%, 0.5%, 0.75%,
Concentration 1% and 1.25% can be seen in table 8. The
% Inhibition IC50
(ppm) highest SPF value (Ultra) was shown in Blackberry
2 9,202714165 ethanol extract starting from a concentration of
4 18,44783715 0.75%.
9,27443
6 31,50975403
8 45,7591179

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 Neni Sri Gunarti et al / Antioxidant and Sunscreen Activities from Blackberry Extract in Vitro Using
Spectrophotometry UV-Visible

Table 6: Percent Transmission Erythema (%Te)

% Transmission Erythema
Replication
0,25% 0,50% 0,75% 1% 1,25%
I 15.49 2.37 0.29 0.033 0.006
II 15.54 2.35 0.3 0.032 0.007
III 15.45 2.39 0.28 0.032 0.008
Average
15.49 2.37 0.29 0.03 0.007
%Te
Extra
Sunscreen Regular Total Total Total
Protectio
Category Suntan Block Block Blok
n

Table 7: Percent of Pigmentation Transmission (%Tp)

% Pigmentation Transmission
Replication
0,25% 0,50% 0,75% 1% 1,25%
I 38.78 13.74 4.43 1.07 0.39
II 38.85 13.72 4.58 1.07 0.4
III 38.79 13.77 4.37 1.09 0.42
Average %Tp 38.81 13.74 4.46 1.08 0.40
Sunscreen Category Total Block Total Block Total Block Total Block Total Block

Table 8: SPF Average Value

SPF Value
Replication
0,25% 0,50% 0,75% 1% 1,25%
I 2.85 9.24 34.55 167.25 328.86
II 2.9 9.35 33.56 166.29 305.73
III 2.9 9.34 33.48 184.75 341.15
SPF Average 2.88 9.31 33.86 172.76 325.25
Sunscreen
Minimal Extra Ultra Ultra Ultra
Category

DISCUSSION AND CONCLUSION
Maceration is a method of separating compounds
by immersion using organic solvents at room
temperature and in a certain time. In this study
the extraction process using maceration method is
carried out so that the classes of compounds that
cannot stand the heat are not easily oxidized, one
of them is an antioxidant compound. The
advantage of maceration method in separating
secondary metabolite compounds besides being
cheap and easy to do is during immersion of
compounds in cytoplasm which will dissolve in the
solvent according to its polarity. Extraction results
showed that from 3000 grams of fresh fruit,
436.7 grams of thick extract was obtained with
extract yield of 14.6% according to research
conducted by Kawiji et al (2015). The amount of
yield produced would depend on the polarity of
the bioactive compounds contained in the solvent
used, so that most likely the compounds
contained in blackberry fruit extracts are more
polar, this is because 96% ethanol solvent is a
polar solvent with polarity index of 5.2 (Satyajit et
al, 2012).
Quantitative antioxidant activity testing of
blackberry fruit extracts was carried out using the
DPPH method. This method is used because it has
several advantages, namely testing is simple,
easy, fast, and requires a small number of
samples in a short time. In addition, this method
has proven to be accurate and practical
(Molyneux, 2004). The mechanism of action of
antioxidants occurs in three main stages, namely
initiation, propagation, and termination. In
principle, the measurement of antioxidant activity
using the DPPH method is a change in the
intensity of the purple DPPH color which is
proportional to the concentration of the DPPH
solution. DPPH free radicals that have no electron
pair will give a purple color and the color will turn
yellow when the electrons are paired. This change
in purple intensity occurs because of the reduction
of free radicals produced by the reaction of DPPH
molecules with hydrogen atoms released by the

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Spectrophotometry UV-Visible
sample compound molecules to form diphenyl
picrilhidrazil compounds and cause DPPH color
changes from purple to yellow. This color change
results in changes in absorbance at the maximum
wavelength of DPPH using UV-Vis
spectrophotometry so that the value of free
radical scavenging activity will be expressed as
IC50 values (Molyneux, 2004). Before measuring
the absorbance of the extract with DPPH, the
maximum wavelength of DPPH was measured.
Absorbance values that are in the range range
from 0.2 to 0.8 (Skoog et al, 2007). From the
results of measurements that have been made,
the maximum wavelength obtained is 515,679
nm with an absorbance of 0.798. Antioxidant
activity test of the ethanol extract of blackberry is
done by making a series of 100-500 ppm
solution concentration. The test results obtained
by the IC50 value of strawberry methanol extract
of 407.5175 ppm. This shows that the ethanol
extract of blackberries has weak antioxidant
activity according to Molyneux (2004) because it
has an IC50 value of 250-500 ppm. The
comparison used in this research is vitamin C pa
because it has an antioxidant function, vitamin C
is included as a secondary class antioxidant which
is able to ward off free radicals, because vitamin
C has a free hydroxyl group that acts as a catcher
of free radicals and if it has a polyhydroxy group
it will increase antioxidant activity (Isnindar et al,
2011). In this test, vitamin C solution was made
100 ppm mother solution and then made serial
glow with concentrations of 2 ppm, 4 ppm, 6
ppm, 8 ppm and 10 ppm. Then added with a 2
ml DPPH solution and sufficient with methanol p.a
to 10 ml, after that it is allowed to sit in a dark
room for 30 minutes at 370C to react with the
DPPH solution. The absorbance measurements
were then performed using UV-Vis
spectrophotometry at a wavelength of 515,679
nm. The process is carried out in a room that is
protected from direct sunlight so that the sample
solution and DPPH remain stable because
sunlight can decompose the solution and affect
the measurement results. IC50 value of Vitamin C
is 9.227443 ppm with a strong antioxidant
category.
Determination of the sunscreen potential of
blackberry fruit extracts was done in vitro using a
UV-vis spectrophotometer in the wavelength
range of ultraviolet rays. The determination of
sunscreen potential is based on the percent
transmission value of erythema and pigmentation
transmission by calculating the SPF (Sun
Protecting Factor) value. Blackberry ethanol
extract has weak antioxidant activity with IC50
407.5175 ppm and% erythema sunscreen activity
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at concentrations starting from 0.75% including
total Block,% pigmentation at concentrations
starting from 0.25% including total Block and SPF
values above 30 (ultra) at concentrations ranging
from 0.75% which are able to reflect UV A and
UV B rays, and have a very long time to block UV
rays from entering the skin.
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