The Effect of Processing Methods on Antioxidant Activity of Loloh Tempuyung (Sonchus arvensis L.)

Abstract

The effect of different extracts from dry and fresh leaves by two extraction methods on the total polyphenols, tannin contents and the antioxidant activity of Sonchus arvensis L. that used to make loloh tempuyung was studied. The antioxidant activity of these extracts was investigated by using DPPH radical scavenging. The results showed that the extracts exhibited different level of total polyphenols and antioxidant activity. The antioxidant activity of the extract from fresh leaves by boiled water were significantly higher than the extract from dry leaves and by other extraction methods. In conclusion, all extract from both fresh and dry leaves that used to make loloh tempuyung as potential sources of new antioxidant agents.

Full text

The Effect of Processing Methods on Antioxidant Activity of
Loloh Tempuyung (Sonchus arvensis L.)
Wita Kusumawati IGA1), Darmawijaya I Putu2), Yogeswara I.B.A1)
1 Nutrition Science, Dhyana Pura University, Bali, Indonesia
E-mail: wita_kusumawati@yahoo.co.id
2Fisioteraphy Dhyana Pura University, Bali, Indonesia
Abstract— The effect of different extracts from dry and fresh leaves by two extraction methods on the total polyphenols, tannin contents and
the antioxidant activity of Sonchus arvensis L. that used to make loloh tempuyung was studied. The antioxidant activity of these extracts was
investigated by using DPPH radical scavenging. The results showed that the extracts exhibited different level of total polyphenols and
antioxidant activity. The antioxidant activity of the extract from fresh leaves by boiled water were significantly higher than the extract from
dry leaves and by other extraction methods. In conclusion, all extract from both fresh and dry leaves that used to make loloh tempuyung as
potential sources of new antioxidant agents.
Keywords— loloh tempuyung, antioxidant activity, processing methods, DPPH
I. INTRODUCTION
Flavonoids are a group of compounds present in
fruits, vegetables, nuts, seeds and beverages of plant
origin, which have been reported to exhibit anti
carcinogenic, anti inflammatory and antimicrobial
activities. These benefits are explained by its potent
antioxidant effect as metal chelator or free-radical
scavenging activities observed in vitro. [1,2,3,4,5,6,7].
Balinese people have been used plant extract as a
medicine based on the concept of usada taru pramana [8].
Tempuyung is one of plants that can be used as a herbal
medicine. The plants have been used by the Balinese
people and its believe to treat several diseases. The used
of herbal medicine in traditional society its because the
adverse effect of synthetic chemical drugs [9,10].
Tempuyung (Sonchus arvensis L.) plants can grow
wild and very easy to bred [11,12,13]. Tempuyung
leaves is less favored when consumed directly so they
processed tempuyung leaves in form of beverage or loloh
(in Balinese). Balinese people usually consumed loloh
tempuyung to treat several diseases such as asthma,
antidepressan, cough, kidney stone [13] and
antidiarrhorea [12]. As reported [14] that the formulation
of tempuyung leaves powder and water as a solvent
extraction (1 g : 100 mL) exhibit antioxidant activity
4,43 ppm. Moreover, the addition of sweetener (brown
sugar) increased the antioxidant activity of loloh
tempuyung [15].
The part of the plants that is often used to make
loloh is the leaves. Traditional methods to make loloh
tempuyung is by boiling the leaves for 30 minutes at 100
C or by adding the mash of fresh leaves into the mineral
water [16]. The processing methods will effect the
bioactive compound during the processing of loloh
tempuyung. A study conducted by Szydlowska- Czerniak
(2012) reported that, there is a different antioxidant
content between processed foods and unprocessed foods
[17].
The aim of this research is to study the effect of
different processing methods on the antioxidant activity
of dry leaves and fresh leaves
II. MATERIALS AND METHODS
A. Plant material
Tempuyung leaves was harvest in March 2015 at Desa
Kukuh, Tabanan, Bali. Fresh leaves samples were
prepared by washing 4 kg of the leaves, rinsed and
weight the leaves as much as 25 g. For the dry leaves,
were prepared by naturally dried the leaves for seven
days without directly exposed to the sunlight. The dry
samples were weighed to 1 g of samples. Each of the
samples were extract using hot extraction methods and
cold extraction to make loloh tempuyung.

B. Extraction procedure
B.1. Extraction with boiling water
Each of the samples (fresh and dry leaves) were extract
by adding 100 ml of water and let it to boil for 10
minutes. Then, the samples were filtered using filtered
paper. The water extract of the leaves is used to analyze
the antioxidant activity of loloh tempuyung.
B.2. Extraction without boiling water
The extraction methods without boiling is a traditional
methods to make loloh tempuyung by Balinese people.
Each of the samples were added 100 ml of warm water
(70 C) and were squeezed to obtain the filtrate. The
filtrate were filtered using filtered paper and proceed to
analyze the antioxidant activity of loloh tempuyung.
C. Deterrnination of total phenolics contents
The total phenolics content were determined using the
Folin-Ciocalteu reagent by [18] with some modification.
Briefly 0.1 mL of each extract was mixed with 0.5 mL of
Folin-Ciocalteu reagent during 6 min at 25⁰C. The
reaction was then neutralized with 2.5 mL of 5% sodium
carbonate, and the mixture was futher incubated at 25⁰C
for 30 min. The absorbance was then measured at 760
nm with UV-vis spectrophotometer. A gallic acid
standard curve with a linear range (0-100 mg/L) was
prepared from a freshly made gallic acid stock solution.
Results were expressed as mg of gallic acid equivalents
GAE/g extract on dry weight basis.
D. Antioxidant activity
The antioxidant activity were determined using the
DPPH radical-scavenging activity method by [19]. A
volume of 1 mL each extract was added to 3 mL of
DPPH solution (0.004% in methanol) as free radical
source. The mixtures were incubated for 30 min in the
dark at room temperature. Scavenging capacity was
measured spectrophotometrically by monitoring the
decrease in absorbance at 517 nm. Gallic acid was used
as a positive control.
E. Statistical analysis
The results were statistically analyzed by ANOVA to
determine any significant differences between the
average value obtained by the different extracts sample
and processing extracts method, at 95% confidence.
III. RESULTS AND DISCUSSION
A. Extraction
Extraction of fresh leaves or tempuyung leaf powder
performed to obtain bioactive components contained in
the leaves tempuyung. Solvent extraction using water on
the basis of the selection is H2O present high polarity
index, dielectric constant and cohesive energy, compared
with other solvents which provides strong bounding
between water molecules and polar compounds from the
solute, causing their dissolution, easy to used, efficient
and can be applied to various types of analytes [20, 21].
Another reason for the selection of water as a solvent is
Balinese people use water as a solvent to make
tempuyung loloh [14].
B. Total Phenolic Contents
The total phenolic contents (TPC) values were quantified
based on the linear equation obtained from gallic acid
standard calibration curve. Thus, TPC value were
expressed as gallic acid equivalent (mg GAE/g samples).
The total phenolic contents of the different extract from
fresh leaves and dried leaves to make loloh tempuyung
exhibited different level of total polyphenols was shown
in Figure 1. The total phenolic contents of the extracts as
estimated by Folin-Cioucalteu reagent method. Results
of ANOVA analysis indicated that there was significant
difference (p < 0.05) between fresh and powdered forms
in all samples studied.
Loloh tempuyung that make from dried leaves had the
highest TPC compared to its fresh forms. The finding
was in agreement with the study of [22] whereby they
reports that there was a significant increase of TPC in
turmeric leaf, pandan leaf and torch ginger flower
through freeze-drying process as compared with its fresh
forms which was probably due to the discharge of

phenolic compounds from the disintegration of cellular
constituents after undergoing freeze-drying process.
Apart from that, non-phenolic substances such as water,
fat, sugars, proteins and pigments may interrupt the
evaluation of TPC. Therefore, [23] reported that upon
extraction procedure to remove these compounds
significant high amount of TPC was obtained in the
medical plants studied. This may explain for the
significantly higher TPC of powdered forms as compared
with fresh forms in the present study.
Fig 1. Total Phenolic Content of loloh tempuyung from
fresh leaves and dried leaves that made by boiled water
and blanch water
Total phenolics content of blanch water loloh
tempuyung was found to be higher that of the other for
both processing methods. The activity of all phenolic
acids decreased with increasing temperature. This result
is in agreement with the reports of [24] whereby they
reports that the total phenols, individual and total caffeic
acid derivatives contents of E. purpurea flowers extracts
was enhanced by elevated extraction temperatures ranged
from 25 to 65 ⁰C. As [20] reported phenolics content of
O.ficus-indica flowers extract is not only affected by the
solvent polarity but also by the extraction process. The
capacity of water to dissolve residual substances such as
organic acids and sugars, that could interfere in the
phenolics quantification [25].
C. Antioxidant activities
The antioxidant activities of the loloh tempuyung was
evaluated by the DPPH radical scavenging assay which
is largely reported to screening antioxidant capacity due
to their simplicity, stability and rapidity. DPPH radical is
scavenged by antioxidants compounds present in the
extracts by the proton donation to form reduce DPPH
which can be quantified by its decrease of absorbance.
The higher the inhibition rate is, the greater the hydrogen
donating ability, thus the higher antioxidant activities
[20, 26, 27, 28]. The antioxidant activity of the different
extract from fresh leaves and dried leaves to make loloh
tempuyung exhibited different level of total polyphenols
was shown in Figure 2.
Fig 2. Antioxidan activities of loloh tempuyung from
fresh leaves and dried leaves that made by boiled water
and blanch water
The DPPH radical scavenging activities of the loloh
tempuyung obtained using the boiled water process are
generally higher compared to those obtained using the
blanch water. As [29] reported that beside the radical
scavenging activity, the total phenolics and flavonoids
contents in the C. sulcate peel and edible fruits extracts
obtained by the soxhlet extraction method were higher
than those in the extracts obtained by ultrasound and
high pressure water extraction. Even though, some of
heat sensitive compounds may be decomposed in the
soxhlet technique [30]. However, thermostable
compounds cannot be dehydrolyzed thanks to their
stability. It can be concluded that tempuyung leaf
extracts are rich in thermostable compounds which have
antioxidant properties and they are better extracted using
boiled water processed.

With regard to the TPC of the different studied loloh
tempuyung, the obtained results showed that a high
content of phenolic compounds does not correspond
automatically to a high antioxidant activity. Loloh
tempuyung that make from dried leaves with the highest
TPC (13.15 mg GAE/g) did not show the highest
antioxidant activity. This may be also due to synergy
phenomena, which occur within each extract.
IV. CONCLUSIONS
The extracts of tempuyung leaves exhibited different
level of total polyphenols and antioxidant activity. The
antioxidant activity of the extract from fresh leaves by
boiled water were significantly higher than the extract
from dry leaves and by other extraction methods. In
conclusion, all extract from both fresh and dry leaves that
used to make loloh tempuyung as potential sources of
new antioxidant agents.
ACKNOWLEDGMENT
This research was funded by the Ministry of Education
and Culture via Hibah Penelitian Dosen Pemula.
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