IOP Conference Series: Earth and Environmental Science

PAPER • OPEN ACCESS You may also like

- Toxicity of GO and rGO suspension
Antibacterial and Natural Pesticide Properties of against P. acnes: physical puncture and
oxidative stress
Nyiri (Xylocarpus granatum) Dongfang Dai, Hao Qu, Jianxin Lv et al.

- Encapsulated Lime Peel Essential Oil

(Citrus hystrix) Into Chitosan Nanoparticle:
To cite this article: Saat Egra et al 2022 IOP Conf. Ser.: Earth Environ. Sci. 1083 012020 New Entity to Enhanced Effectivity Against
Propionilbacterium Acne in Vitro
Linda Julianti Wijayadi and Taty Rusliati
Rusli

- The Effect of Temperature and Pressure

View the article online for updates and enhancements. on the Performance of a PEMFC Exposed
to Transient CO Concentrations
Mahesh Murthy, Manuel Esayian, Woo-
kum Lee et al.

This content was downloaded from IP address 139.255.15.182 on 24/10/2022 at 06:41

CABE-2021 IOP Publishing
IOP Conf. Series: Earth and Environmental Science 1083 (2022) 012020 doi:10.1088/1755-1315/1083/1/012020

Antibacterial and Natural Pesticide Properties of Nyiri

(Xylocarpus granatum)

Saat Egra1*, Hartini1, Mardhiana1, Nurjannah1, Muhammad Adiwena1, Dwi

Santoso1, Harlinda Kuspradini2, Tohru Mitsunaga3.
1
Agriculture Faculty of Universitas Borneo Tarakan, Jl. Pantai Amal Lama No. 1
Tarakan, Indonesia
2
Forestry Faculty of Mulawarman University. Jl. Ki Hajar Dewantara Kampus
Gunung Kelua. Laboratory of Forest Product Chemistry, Faculty of Forestry,
Mulawarman University, Samarinda, Kalimantan Timur, Indonesia
3
Applied Biological Sciences, Gifu University, Japan

Email : saat.egra@borneo.ac.id*

Abstract. Nyiri (Xylocarpus granatum) thrives in the mangrove areas of Indonesia, especially
Tarakan, North Kalimantan. X. granatum is one of many species found in the Tarakan
mangrove forest and belongs to the Meliaceae family. This plant has been utilized by
communities for pesticides and health care. So, the purpose of this study is to determine the
potential of X. granatum in inhibiting the growth of Ralstonia solanacaerum and
Propionibacterium acnes. This study we used well agar diffusion methodology with a
concentration of 5000 ppm, 10,000 ppm, 20,000 ppm. Positive controls in this study were
Chloramphenicol and 40% ethanol as negative control. The determine variables are the
percentage of inhibition diameters (DDH). The percentage of DDH shows that the ethanol
extract of leaves and bark X. granatum against R. solanacearum and P. acne at a concentration
of 5000 ppm was not able to inhibit, whereas the concentration of 10,000 ppm and 20,000 ppm
was able to inhibit the R. solanacearum with a percentage of 31.09 respectively % and 34.46%.
Whereas, the bark extract was able to inhibit R. solanacearum with the percentage of DDH at a
concentration of 10,000 ppm was 35.73% and 20,000 ppm was 39.94%. Furthermore, ethanol
extract of X. granatum leaves against P. acne was able to inhibit the concentration of 10,000
ppm with a value of 33.83% and 20,000 ppm with a value of 38.01%. The percentage of DDH
for X. granatum bark is able to inhibit P. acne at all concentrations, namely at concentrations
of 5000 ppm (35.34%), 10,000 ppm (39.59%) and 20,000 ppm (41.63%).

Keywords: Antimicrobial, Mangrove, P. acne, R. solanacearum, X. granatum.

1. Introduction
Mangrove is a forest area that has a lot of plant diversity which is generally located in coastal areas of
the tropics and subtropics. The diversity of mangrove forest vegetation in Indonesia that has been
recorded is as many as 202 species, dominated by four families, namely Rhizophoraceae,
Sonneratiaceae, Avicenniaceae and Meliaceae [1].
Mangrove plants contain bioactive compounds, including alkaloids, steroids, terpenoids, saponins,
and tannins as well as flavonoid and quinone compounds with various bioactivities such as
antimicrobial, antifungal, antiviral, antitumor, antileukemic, and insecticide [2]. Extracts and raw
materials from mangrove plants have been widely used by coastal communities for natural medicines.

Content from this work may be used under the terms of the Creative Commons Attribution 3.0 licence. Any further distribution
of this work must maintain attribution to the author(s) and the title of the work, journal citation and DOI.
Published under licence by IOP Publishing Ltd 1
CABE-2021 IOP Publishing
IOP Conf. Series: Earth and Environmental Science 1083 (2022) 012020 doi:10.1088/1755-1315/1083/1/012020

A number of mangrove plants and associated plants are also used as traditional ingredients for
insecticides and vegetable pesticides. Vegetable pesticides are a solution to control plant pest
organisms (OPT), one of which is wilt disease caused by the bacterium Ralstonia solancearum. R.
solanacearum is a soil-borne plant pathogen that is commonly found in subtropical and tropical areas,
this bacterium naturally infects roots and reproduces in xylem tissue [3]. R solanacearum causes wilt
disease in plants, especially agricultural crops such as tomatoes, peanuts, bananas, potatoes, tobacco
and other Solanaceae tribes [4]. In addition to using the bacteria R. solanacearum in this study, the
bacteria P. acn0e that causes acne were also used.
Acne is a chronic skin disease due to abnormal sebum production in the sebaceous glands that
appears when the oil glands on the skin are overactive [5]. Acne can occur at a young or old age with
the percentage of occurrence in women as much as 27% and 34% in men. P. acnes is a bacterium that
plays a role in the formation of acne [6]. Most people treat acne by using antibiotics such as
tetracycline and clindamycin which are considered to have side effects that can cause irritation and the
emergence of bacterial resistance. Therefore, it is necessary to develop acne drugs that come from
nature that do not cause side effects if consumed properly. Thus, as an effort to reduce or inhibit the
growth of these two pathogenic bacteria, this study used X. granatum plants.
Xylocarpus granatum (Nyiri) is one of the most common species found in mangrove forests
belonging to the Meliaceae family. X. granatum has been used for generations by coastal communities
traditionally. X. granatum obtained from mangrove forests contain flavonoids, saponins, tannins and
phenols. These compounds have antibacterial and antioxidant properties that can inhibit or slow down
the damage caused by the oxidation process, and are needed by the body to protect against free radical
attack [7]. Based on this information, it is necessary to test the potency of the extract of the leaves and
bark of X. granatum in inhibiting the growth of R. solanacearum and P. acnes bacteria.

2. Materials and Methods

Samples and Materials

This research was conducted in the Laboratory of Natural Product Chemistry and Laboratory of Plant
Diseases and Pests. The raw materials for the leaves and bark of X. granatum were taken in the
Mangrove and Proboscis Monkey Conservation Area (KKMB) of Tarakan City. Other materials used
were distilled water, ethanol, P. acnes and R. solanacearum microbes, nutrient agar, chloramphenicol.
The equipment used is a spectrophotometer model Shimadzu UV Vis 1200 (Shimadzu co, Japan),
evaporator, shaker, oven, autoclave model All Americans.

Sample Preparation, Moisture Factor (MF) and Yield

Sample preparation for extraction was by washing and drying the sample leaves and bark for 3-6 days
at room temperature and then blended to get a small size and then soaked in 96% ethanol. A rotary
evaporator is used to obtain a concentrated extract of the sample. Moisture factor was obtained by
cutting the sample into several parts to facilitate the drying process in the oven at 100OC for 1 x 24
hours. To calculate and obtain the value of the moisture factor, the dry weight of the kiln is divided by
the initial weight. Then the preparation and the moisture factor will get the yield value by dividing the
weight of the extract by the weight of the fresh sample multiplied by the humidity factor multiplied by
100% [8].

Antimicrobial Assay
The main ingredients used for bacterial growth media are nutrient broth (Difco), glucose, then evenly
dissolved in distilled water and sterilized by autoclave at 121ºC for 45 minutes. Antimicrobial testing
was carried out using the diffusion method for agar wells and bacterial suspensions equivalent to 106
cells/ml (0.5 McFarland standard). Furthermore, ketepeng leaf extract was tested in several
concentrations (20000ppm, 10000ppm and 5000ppm). Chloramphenicol was used for positive control
and acetone for negative control. Antimicrobial activity was indicated by the presence of an inhibition
zone around the well containing extract, with a number greater than 8 mm [9].
.

2
CABE-2021 IOP Publishing
IOP Conf. Series: Earth and Environmental Science 1083 (2022) 012020 doi:10.1088/1755-1315/1083/1/012020

3.Results

Moisture Factor
The humidity factor aims to determine the availability of water contained in the nyiri (X. granatum)
plant. The calculation of the humidity factor is carried out after obtaining the dry weight value of the
kiln from the sample used. The results of the calculation of the humidity factor can be seen in table 1.
Table 1. Moisture factor and yield
First Last Moisture
Sample Rendement
simplisia (g) Extracts (g)
weight(gr) weight (gr) Factor (%)
Leave 1,00 0,31 0,31 158,34 17,73 8,91
Bark 1.00 0,53 0,53 125,19 29,78 24,98

After drying for 24 hours in an oven with a temperature of 100oC, the humidity factor of the leaves
is 0.31 and the bark is 0.53. Solvent extraction is used maceration. The extraction process by
maceration technique is carried out with several times of shaking or stirring at room temperature.
Extraction using ethanol solvent. The results of the calculation of the yield on X. granatum were
8.91% of leaves and 24.98% of leaves. The higher the yield value, the more the amount of extract
produced and the high yield value was due to the weight of the simplicia used. The heavier the weight
of the simplicia used, the more weight the extract and yield produced. The antimicrobial activity of X.
granatum extract against R. solanacearum and P. acnes was determined by measuring the zone of
inhibition formed around the well. The diameter of the inhibition zone is the area around the wellbore
where no bacterial growth was found. Antibacterial test in this study was carried out with 3 repetitions
of each treatment obtained different zones of inhibition. The results of the calculation of the
percentage of inhibition can be seen in Table 2.

Table 2. Percentage of inhibition

Inhibition Presentation (%)
Samples R. solanacearum P. Acne
5000ppm 10000ppm 20000ppm 5000 ppm 10000 ppm 20000 ppm

Leave 0 31.09 34.46 0 33.83 38.01

Bark 0 35.73 39.94 35.34 39.59 41.63
Chloramphenicol 100 100
Etanol 40% 0 0

The percentage of inhibition showed that the leaf extract of X. granatum had inhibition against R.
solanacearum at concentrations of 10000 ppm and 20000 ppm with the percentage values of inhibition
were 31.09% and 34.46, respectively. The percentage of inhibition of X. granatum bark extract was
able to inhibit the growth of R. solanacearum bacteria at concentrations of 10000 ppm and 20000 ppm
with inhibition percentage values of 35.73% and 39.94%, respectively. for a concentration of 5000
ppm in both parts of the plant has no inhibitory power.
The percentage of inhibition showed that the leaf extract of X. granatum had inhibitory power
against P. acnes at concentrations of 10000 ppm and 20000 ppm with inhibition percentage values of
33.83% and 38.01%, for a concentration of 5000 ppm it had no inhibitory power. The inhibition value
of the bark extract of X. granatum against P. acne with concentrations of 5000 ppm, 10000 ppm and
20000 ppm with the percentage inhibition values of 35.34%, 39.59% and 41.63%, respectively.

3
CABE-2021 IOP Publishing
IOP Conf. Series: Earth and Environmental Science 1083 (2022) 012020 doi:10.1088/1755-1315/1083/1/012020

4. Discussions

Moisture Factor
The humidity factor is obtained from drying using high temperatures. The moisture factor of the leaves
and bark of X. granatum has different values. The leaves of X. granatum have a humidity factor of
0.31 and the bark of X. granatum has a humidity factor of 0.53. In addition to humidity, water content
provides a minimum limit on the amount of water content in the material. Water that is still remaining
in the simplicial at a certain level can be a medium for the growth of microorganisms that will affect
the active substances contained therein. Therefore, the smaller the water content contained in the
simplicia, the less damage caused by microorganisms will be. Simplicial overgrown with mold will
affect the active substances contained therein. Susiani et al. (2017) said the drying process was carried
out until the leaves were easily crushed with a drying time of 1 month [10].
Drying is the most important step in maintaining the stability of compounds in simplicial. Drying
of plant material aims to prevent the growth of molds and fungi, reduce water content, prevent
enzymatic reactions and make it easier when simplicial will be ground into powder.

Yield Percentage
The percentage yield of X. granatum leaf extract on the leaves and stems were 8.91% and 24.98%,
respectively. To obtain more active substances, it takes a long process time because this extraction
does not use the help of heat. Nurasiah (2010) report that due to the absence of other force assistance,
the maceration was only carried out by immersion so that the osmosis of the solvent into the solid was
static even though the solvent was changed using the remaceration method [11]. The results of the
antibacterial assay of the leaf and stem bark extracts of X. granatum against R. solanacearum observed
after 18 hours of incubation can be seen in Figure 1.

(a) (b)
Figure 1. The results of observations of X. granatum extract against the bacterium R. solanacearum
Description (a) X. granatum leaf extract, (b) X. granatum bark extract

In this study, chloramphenicol was used as a positive control because chloramphenicol is a broad-
spectrum antibiotic that can inhibit Gram positive and negative bacteria [12]. The mechanism of
chloramphenicol in inhibiting bacteria by joining the ribosomal subunits so as to prevent the joining of
amino acids into proteins so that the synthesis of amino acids is disrupted or even does not take place.
This results in bacterial cell death. Antibiotics with the mechanism of interfering with protein
synthesis have high antibacterial activity. In this study, crude extract of X. granatum was used as an
antibacterial. Crude extract contains various other compounds that can influence to inhibit bacterial
growth, namely triterpenoids, steroids, saponins and tannins and flavonoids [13].
Positive control (Chloramphenicol) on R. solanacerum had 100% inhibition and negative control
(Ethanol) had no inhibition. The percentage inhibition of R. solanacearum indicated that the extract of
the leaves and bark of X. granatum had antibacterial activity. Leaf and bark extract of X. granatum
with a concentration of 20000 ppm had the highest value in inhibiting R. solanacearum, namely
34.46% and 35.73%. The lower the concentration of the extract tested, the smaller the diameter of the

4
CABE-2021 IOP Publishing
IOP Conf. Series: Earth and Environmental Science 1083 (2022) 012020 doi:10.1088/1755-1315/1083/1/012020

inhibition formed and the greater the concentration of the starfruit leaf extract, the greater the
inhibition zone [14]. Tanendri (2017) revealed that the difference in the concentration of the extract
used caused the clear zone which showed different inhibition. It is stated that the higher the
concentration of guava leaf extract, the more concentrated the solution and the greater the amount of
antimicrobial substances contained in it [15].
This study showed that the extract of the leaves and bark of X. granatum also had antibacterial
activity that was able to inhibit the growth of P. acnes bacteria. The positive control
(Chloramphenicol) on P. acnes also had 100% inhibition and the negative control (Ethanol) had no
inhibition. Leaf extract with a concentration of 5000 ppm on P. acnes had no inhibitory power but the
bark extract with a concentration of 5000 ppm had an inhibitory power of 35.34%, leaves and bark of
X. granatum with a concentration of 20000 ppm had the highest value in inhibiting P. acnes are
38.01% and 41.63%.
The results of the antibacterial assay of the leaf and stem bark extracts of X. granatum against P.
acnes observed after 18 hours of incubation can be seen in Figure 2.

(a) (b)

Figure 2. Observation results of X. granatum extract against P. acne bacteria. Description; (a.) X.
granatum leaf extract, (b.) X. granatum bark extract

Antibacterial compounds are substances that can inhibit the growth of bacteria and can be used for the
treatment of infections in humans, animals, and plants. According to Murhadi (2002), antibacterial
compounds derived from plants are mostly known as secondary metabolites from the phenolic group,
terpenes in essential oils and alkaloids. Most of these secondary metabolites are biosynthesized from
primary metabolites such as amino acids, acetyl Co-A and intermediate metabolites [16]. In this study,
X. granatum extract was able to inhibit the growth of R. solanacearum and P. acnes bacteria. It is
suspected that the extract of X. granatum contains secondary metabolites. Such as flavonoid
compounds, saponins, tannins, and phenols [7].
Flavonoids are the largest group of phenolic compounds so that flavonoids have effective
properties in inhibiting the growth of bacteria, viruses and fungi [17]. The mechanism is through
damage to the cytoplasmic membrane, H+ ions from phenolic compounds and their derivatives or
commonly called flavonoids will attack polar groups or phosphate groups, this will cause the
cytoplasmic membrane to leak and bacterial growth will be inhibited. The mechanism of saponins
works as an antimicrobial by disrupting the stability of the bacterial cell membrane, causing cell
bacteriolysis. This causes the cytoplasm to leak so that intracellular compounds leave the cell resulting
in cell death [18].
Tannins are a group of flavonoids. Tannin compounds have antibacterial and antiviral activity. The
antibacterial properties of tannins are caused by the pyrogalol groups and galloil groups, while the
inhibitory properties against viruses are determined by the tertiary structure of the compound catechol
or pyrogalol groups with their galloil groups [19].
Terpenoid compounds have chemical compounds which are natural ingredients used as drugs.
Terpenoids are fat soluble and are found in the cytoplasm of plant cells. The mechanism of terpenoids

5
CABE-2021 IOP Publishing
IOP Conf. Series: Earth and Environmental Science 1083 (2022) 012020 doi:10.1088/1755-1315/1083/1/012020

as antibacterial is that terpenoids will form polymeric bonds that can damage porins. Through
damaged porin which is the entrance and exit of compounds, it can reduce the permeability of the
bacterial cell wall which will result in the bacterial cell being deficient in nutrients, so that bacterial
growth is inhibited or dead [20].
In addition to some secondary metabolites, the type of Gram bacteria also affects the formation of
the inhibition zone. Antibacterial assay of extracts of leaves and bark of X. granatum against R.
solanacearum showed positive results but produced a small clear zone. This happens because the
bacteria R. solanacearum is a group of Gram-negative bacteria that is classified as weak, characterized
by the formation of a small diameter of the barrier. The structure of the cell wall of Gram-negative
bacteria is more complex than the structure of the cell wall of Gram-positive bacteria. Gram negative
bacteria have several layers, namely an outer layer, a middle layer and an inner layer. Thus, this
bacterium R. solanacearum has a relatively complex cell wall structure and makes it more difficult for
antibacterial compounds to enter cells and find targets to work.
In contrast to the antibacterial assay against R. solanacearum, the antibacterial assay against P.
acne had positive results and produced a large clear zone. The possibility because P. acnes is a Gram-
positive bacterium that is easy to inhibit. Gram positive bacteria only have one layer and are more
sensitive to antibacterial components, making it easier for antibacterial compounds to enter cells and
find targets to work.

Conclusion
This research has proved the Niyiri (X. granatum) leaf and bark extracts possessed strong antibacterial
activity against R. solanacearum bacteria. The most effective in this study was at a concentration of
10,000 ppm. The results provide the basic scientific data, for further research better do the isolation
and identification compounds.

Acknowledgments
The authors thank to the Institute for Research and Community Service (LPPM), University of Borneo
Tarakan for research support and members of the Natural Chemistry Laboratory, Faculty of
Agriculture, Universitas Borneo Tarakan, North Kalimantan. This research also supported by
Laboratory of Forest Product Chemistry, Forestry Faculty, Universitas Mulawarman.

References

[1] Bengen DG. 2001. Bogor (ID): Pusat Kajian Sumberdaya Pesisir dan Lautan IPB.
[2] Soetarno S. 2000. Acta Pharmaceutica Indonesia. 12 (4): 84- 103.
[3] Yabuuchi E et al. 1995. Transfer Of Two Burkholderia And An Alcaligenes Species To
Ralstonia Gen-Nov-Proposal Of R. Comb-Nov. Microbiol. Immnunol. 39(11):879-904.
[4] Persley, G. J. In: Bacterial Wilt Disease in Asia and the South Pacific. Proceedings of
international workshop held at PCARRD, Los Banos, Philippines. 1985. p. 71-75.
[5] Kumar BHA, YN Sachidanand. 2001. Indian Journal Of Dermatology 46(3): 138-141.
[6] Harahap M. 2000. Ilmu Penyakit Kulit. Jakarta: Hipokrates.
[7] Hendrawan, Ita Z, Bagus FP 2015. Jurnal Ilmu Perikanan Tropis 20 (2): 15-22.
[8] Egra S et al. 2019. Biofarmasi J Nat Pro Biochem. Vol 16 (1): 17-23
[9] Yanti, K H., Putri AS. Egra S. 2019. BIODIVERSITAS. Vol. 20 (7): 2039-2042.
[10] Susiani EF. Borneo Journal of Pharmascientech, Vol 01 (2): 2548 – 3897.
[11] Nurasiah E S. 2010. [Skripsi]. Institut Pertanian Bogor, Bogor.
[12] Pelczar Michael J, Chan Ecs. 2008. Dasar-Dasar Mikrobiologi Jilid II. Jakarta: Ui Press.
[13] Dewi N, Puspawati, NM, Swantara IMD, Asih I, Rita WS. 2014. Indonesian E-Journal Of
Applied Chemistry 2(1): 7-16.
[14] Sari D, Suryani A. 2014. Jurnal Fakultas Ekonomi Dan Bisnis Universitas Udayana, Bali.
[15] Tanendri Arrizqiyani. 2017. Jurnal Kesehatan Bakti Tunas Husada 17 (2).
[16] Murhadi. 2002. Institut Pertanian Bogor, Bogor.
[17] Anggraini N, Saputra O. 2016. Jurnal Kedokteran 5 (1): 76-80

6
CABE-2021 IOP Publishing
IOP Conf. Series: Earth and Environmental Science 1083 (2022) 012020 doi:10.1088/1755-1315/1083/1/012020

[18] Kusumawati, Eko Et Al. 2015. Universitas Mulawarman, Samarinda.

[19] Harismah K. 2002.Universitas Muhammadiyah Surakarta, Surakarta.
[20] Cowan. 1999. Clinical Microbiology Reviews 12 (4) : 564-582.