GC-MS Analysis of Bioactive Compounds in Lime Leaf Ethanol Extract (Citrus amblycarpa (Hassk.) Ochse), and Its Potential as a Traditional Medicine Agents

Abstract

Lime (Citrus amblycarpa (Hassk.) Ochse) has been widely used in traditional medicine. However, only some studies have comprehensively revealed the content of leaves, including their use as candidate medicinal ingredients. This study aimed to analyze the active compounds of lime leaves through GC-MS analysis and their benefits in traditional medicine. Experimental research with a laboratory approach is used. The implementation of research by analyzing the bioactive content of lime leaf ethanol extract using GC-MS along with a comprehensive analysis related to its benefits as a traditional medicine agent. Data analysis using the Willey 7 Library database and descriptive analysis. Using GC-MS showed that lime leaf ethanol extract contains 75 compound components dominated by 11 principal components, including citronellol, caryophyllene, hexadecanoic acid, ethyl ester, 1-heptatriacotanol, Phytol, ethyl 9,12,15-oate; methyl glycocholate, 3TMS derivative; 3,7-dimethyloct-6-en-1-yl stearate; ethyl iso-allocate; Rhopin; and tricyclo [20.8.0.0 (7,16)] triacontane, 1(22),7(16)-diepoxy-. Based on the findings of the content evaluation, the compound constituents of lime leaves have the potential to be developed as traditional remedies for both degenerative and infectious diseases. However, further research is needed precisely to identify the therapeutic effects exerted by using compounds on lime leaves.

Full text

JPPIPA 10(4) (2024)
Jurnal Penelitian Pendidikan IPA
Journal of Research in Science Education
http://jppipa.unram.ac.id/index.php/jppipa/index
___________
How to Cite:
Arsana, I. N., Juliasih, N. K. A., & Widyantari, A. S. S. (2024). GC‒MS Analysis of Bioactive Compounds in Lime Leaf Ethanol Extract (Citrus
amblycarpa (Hassk.) Ochse), and Its Potential as a Traditional Medicine Agents. Jurnal Penelitian Pendidikan IPA, 10(4), 1994–2006.
https://doi.org/10.29303/jppipa.v10i4.3735
GC‒MS Analysis of Bioactive Compounds in Lime Leaf Ethanol
Extract (Citrus amblycarpa (Hassk.) Ochse), and Its Potential as a
Traditional Medicine Agents
I Nyoman Arsana1*, Ni Ketut Ayu Juliasih1, A. A. A. Sauca Sunia Widyantari1
1 Department of Biology, Faculty of Information Technology and Science, Hindu University of Indonesia, Denpasar, Indonesia.
Received: April 27, 2023
Revised: February 11, 2024
Accepted: April 25, 2024
Published: April 30, 2024
Corresponding Author:
I Nyoman Arsana
arsanacita@gmail.com
DOI: 10.29303/jppipa.v10i4.3735
© 2024 The Authors. This open access article
is distributed under a (CC-BY License)
Abstract: Lime (Citrus amblycarpa (Hassk.) Ochse) has been widely used in
traditional medicine. However, only some studies have comprehensively
revealed the content of leaves, including their use as candidate medicinal
ingredients. This study aimed to analyze the active compounds of lime leaves
through GC‒MS analysis and their benefits in traditional medicine.
Experimental research with a laboratory approach is used. The implementation
of research by analyzing the bioactive content of lime leaf ethanol extract using
GC‒MS along with a comprehensive analysis related to its benefits as a
traditional medicine agent. Data analysis using the Willey 7 Library database
and descriptive analysis. Using GC-MS showed that lime leaf ethanol extract
contains 75 compound components dominated by 11 principal components,
including citronellol, caryophyllene, hexadecanoic acid, ethyl ester, 1-
heptatriacotanol, Phytol, ethyl 9,12,15-oate; methyl glycocholate, 3TMS
derivative; 3,7-dimethyloct-6-en-1-yl stearate; ethyl iso-allocate; Rhopin; and
tricyclo [20.8.0.0 (7,16)] triacontane, 1(22),7(16)-diepoxy-. Based on the findings
of the content evaluation, the compound constituents of lime leaves have the
potential to be developed as traditional remedies for both degenerative and
infectious diseases. However, further research is needed precisely to identify the
therapeutic effects exerted by using compounds on lime leaves.
Keywords: Citrus amblycarpa; Ethanol extract; GC-MS; Lime leaf; Traditional
medicine
Introduction
Citrus amblycarpa (Hassk.) Ochse, often known as
lime leaves or kaffir lime, is a West Java-endemic
endemic shrub. It is a hybrid citrus lime species with the
formula Citrus micrantha and Citrus reticulata (Ollitrault
et al., 2020; Soetjipto & Martono, 2017; Talon et al., 2020).
Citrus amblycarpa (Hassk.) Ochse, sometimes known as
the lime leaf, is a citrus plant native to Southeast Asia. It
is planted extensively in Indonesia, Thailand, Malaysia,
and other tropical nations. This plant's spines are
extraordinarily sharp and rigid. The fruit is tiny, has a
thick skin, and tastes exceptionally sour. Like other
limes, the lime has dark green, rectangular leaves, white,
fragrant blooms, and green fruits with a rough peel and
juicy flesh.
In Indonesia, lime leaf is frequently utilized as an
aromatic and flavor-enhancing fruit. Fruits and lime
leaves are utilized more frequently as flavor enhancers
in cooking (Budiarto et al., 2022; Budiarto & Sholikin,
2022). Lime leaves are frequently used as seasoning in
traditional Southeast Asian cuisine. Due to their
chemical composition, lime leaves have potential
benefits in traditional medicine and their culinary
purpose (Budiarto et al., 2021b; Lubinska-Szczygieł et
al., 2018; Panakkal et al., 2021). The bioactive

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
1995
components found in lime leaves include essential oils,
flavonoids, alkaloids, and coumarins. Several volatile
chemicals, including citronellal, linalool, limonene, and
-pinene, make up the essential oil of lime leaves. These
chemicals are responsible for lime leaves' distinctive
scent and flavor. The essential oil content of lime leaves
varies by cultivar, harvesting period, and processing
technique (Budiarto et al., 2021b; Dertyasasa & Tunjung,
2017; Hien et al., 2020; Siti et al., 2022; Suresh et al., 2021).
Lime leaves have been used in traditional medicine
to cure various diseases for millennia. In Indonesia, lime
leaves are frequently employed to cure fever, cough, and
the common cold. Additionally, lime leaves cure
digestive issues such as bloating, gas, and nausea. It is
believed that lime leaves have a cooling impact on the
body, which makes them helpful in treating fever and
other inflammatory disorders (Dertyasasa & Tunjung,
2017; Harmayani et al., 2019; Siti et al., 2022). In
traditional medicine, lime leaves have been used to treat
various diseases for millennia. In Indonesia, lime leaves
are extensively employed to treat fever, cough, and cold.
The leaves of the lime tree are also used to treat digestive
issues such as bloating, flatulence, and nausea.
According to popular belief, lime leaves have a
cooling impact on the body, making them beneficial for
treating fever and other inflammatory diseases (Putra et
al., 2018). Several studies have demonstrated that the
active ingredient lime has biopesticide potential against
mosquitoes (Abdullah & Othman, 2018; Budiarto et al.,
2021a; Kasman et al., 2021; Kasman, 2020; Adnyana et al.,
2021), potentially overcoming hypertension
(Kusumawati et al., 2021), and as an antidiabetic agent
by inhibiting the activity of the enzyme α-glucosidase
(Tambunan et al., 2020). There is limited information on
the health benefits and side effects of lime leaf
consumption. However, the related plant kaffir lime
leaves have been found to have several health benefits,
including reducing stress, improving oral health,
fighting bacteria, promoting healthy skin and hair,
aiding digestion, and relieving piles (Anuchapreeda et
al., 2020; Lim, 2012; Suresh et al., 2021; Wulandari et al.,
2019).
Despite the potential pharmacological properties of
lime leaves, there is a need for more research, mainly
using gas chromatography‒mass spectrometry (GCMS).
GCMS is a powerful analytical technique that can
identify and quantify lime leaf volatile and nonvolatile
compounds. GCMS can provide a more comprehensive
understanding of the chemical composition and
potential pharmacological properties of lime leaves. This
study aimed to analyze the active compounds of lime
leaves through GCMS analysis and their benefits in
traditional medicine. Hopefully, this research can
contribute ideas related to compounds in lime leaves
with potential as candidates for traditional medicine. A
comprehensive analysis of the content is sought to
provide an overview of the importance of using lime
leaves as candidates for medicinal ingredients.
Method
This experimental research with a laboratory
approach was used in this study (Darwin et al., 2021).
This research analyzes the content and benefits of lime
leaves (Citrus amblycarpa (Hassk.) Ochse). The study was
conducted for one month in December 2021. Lime leaf
samples were collected from Denpasar Bali, and then
lime leaf ethanol extract was made at the Laboratory of
the Biology Study Program, Hindu University of
Indonesia. This research flow is presented in Figure 1.
Figure 1. The flow of research
The lime extract was obtained through a maceration
method with a 96% ethanol solvent (Astiti & Ramona,
2021). The simplicial setup process utilizes lime leaves
that are thoroughly washed, cut into small pieces,
blended, dried for 24 h, and filtered using a flour sieve.
After five days of drying, the material is transformed
into dry Simplicia. The lime leaf Simplicia was
macerated in 96% ethanol for 48 hours before filtration
using filter paper. The filtrate was then concentrated in
a rotary evaporator vacuum at 45 °C to produce a
viscous extract. The active component was examined
using GCMS (Shimadzu GC-210 Plus). An HP-5MS UI
capillary column (30.0 m × 0.25 mm × 0.25 m) was used
in the GC MS instruments. Helium was used as the
carrier gas at a flow rate of 1 mL per minute. The GC
temperature was set as follows: the injector temperature
was 230 °C, the column beginning temperature was 60
°C, the temperature rise rate was 10 °C/min, and the
oven final temperature was 280 °C.

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
1996
The compound was identified by matching it
against the Willey7 Library database. An integrated
research and testing lab performed the GCMS analysis
for the study at Gadjah Mada University in Yogyakarta.
The gathered data are then descriptively examined, and
literature searches are conducted about using chemicals
derived from lime leaves as candidates for traditional
medicine in Bali. Images and tales are utilized to present
the findings.
Result and Discussion
Gas Chromatography‒Mass Spectrometry Evaluation of Lime
Leaf Ethanol Extract
The Gas chromatography‒mass spectrometry (GC‒
MS) analysis approach was qualitatively carried out to
identify active compounds by looking at peak areas and
retention times. Based on the results of chromatograms,
there are 75 components of compounds (Figure 2),
including 11 principal components, most of which are
essential oils. Additionally, there are components of
active compounds that have the potential to act as
antioxidants, anticancer agents, anti-inflammatory
agents, and flavor boosters for food. Fragmentation of
this principal component can be seen from the peak on
the mass spectrum of lime extract in Table 1 and Figure
3. Each of the compounds in the compound
chromatogram in Table 1 is a significant component of
lime leaf ethanol extract, each with potential benefits.
The main components are terpenoids. Among these
components, some belong to the essential oil category.
This can be seen in the lime leaf ethanol extract, which
resembles a deep black paste after evaporation.
Figure 2. Chromatogram of lime leaf ethanol extract, peak numbers to the chemicals listed in table 1
Table 1. Main Components of Lime Leaf Ethanol Extract
Top
Retention Time
Chemical Components
Area
BM
Molecular Formula
2
10.33
Citronellol
12.03
156
C10H20O
6
12.98
Caryophyllene
2.04
204
C15H24
28
19.63
Hexadecanoic acid, ethyl ester
2.66
284
C18H36O2
29
19.87
1-Heptatriacotanol
4.58
536
C37H76O
34
21.06
Phytol
7.50
296
C20H40O
37
21.40
Ethyl 9,12,15-octadecatrienoate
3.83
306
C20H34O2
56
24.57
Methyl glycocholate, 3TMS derivative
3.59
695
C36H69NO6Si3
62
26.45
3,7-dimethyl oct-6-en-1-yl stearate
11.45
422
C28H54O2
67
27.95
Ethyl iso-allocholate
3.02
436
C26H44O5
68
28.82
Rhodopin
5.36
554
C40H58O
69
28.97
Tricyclo[20.8.0.0(7,16)]triacontane, 1(22),7(16)-diepoxy-
9.24
444
C30H52O2

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
1997
Figure 3. GC‒MS spectra citronellol mass (a); Caryophyllene (b); Hexadecanoic acid, ethyl ester (c); 1-Heptatriacotanol (d);
Phytol (e); Ethyl 9,12,15-octadecatrienoate (f); Methyl glycocholate, 3TMS derivative (g); 3,7-dimethyloct-6-en-1-yl stearate (h);
Ethyl iso-allocholate (i); Rhodopin (j); Tricyclo[20.8.0.0(7,16)]triacontane, 1(22),7(16)-diepoxy- (k).

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
1998
The Composition of Lime Leaf Ethanol Extract and Its
Traditional Medicinal Benefits
Citronellol
Citronellol is the chemical that gives lime leaves
their characteristic scent. This chemical has a retention
time of 10.33 minutes and a relative peak of 12.03% on
chromatograms. Citronellol belongs to the terpenoid
(monoterpenoid) class, is volatile and is present in
numerous essential oils, particularly leaf components.
The terpenoid chemicals are antibacterial, antifungal,
antiviral, anti-inflammatory, antioxidant, anticancer,
and anti-inflammatory. This category of compounds is
also insoluble in water. By reducing oxidative stress,
citronellol lowers endothelial dysfunction and slows the
course of atherosclerosis in mice and is considered a
possible treatment for atherosclerosis in human studies
(Lu, 2019). Citronellol has been shown to have anticancer
properties by causing apoptosis in the triple-negative
breast cancer cell line MDA-MB-231 by inhibiting the
anti-apoptotic protein Bcl-2, which activates the pro-
apoptotic protein Bax and caspase-dependent apoptosis.
Activation of the Caspase-3 protein. As a result, the pace
of proliferation of cancer cells and their capacity to travel
and survive will be slowed (Ho, 2020). In addition,
citronellol was discovered to be harmful to MCF-7 breast
cancer cells and to inhibit proliferation by causing
apoptosis in Huh7 hepatocellular cells. However, the
precise mechanism by which citronellol slows tumor cell
development remains unknown and deserves further
study (Juliasih & Adnyana, 2023; Yu et al., 2019).
Furthermore, Lopez-Romero et al. (2015) stated that
citronellol is the most effective chemical against
Escherichia coli and Staphylococcus aureus, followed by
citronellal, carveol, and carvone, according to the study.
After exposure to citronellol compounds, changes in
hydrophobicity, surface charge, and membrane integrity
with subsequent E. coli and S. aureus leakage were
detected. Thus, citronellol has the potential to be
conventionally developed as an antibacterial and
antimicrobial agent. Felim et al. (2021) stated that Citrus
amblycarpa (Hassk.) Ochse is highly connected with
numerous pharmacological effects, according to
previous studies. The safety of Ochse seed ethanol
extract (CASEE) at 12.5 and 75 g/mL was demonstrated.
CASEE at a concentration of 75 g/mL exhibited a mean
reaction of 312.75 to TNF-, 155.19 to IL1-, and 524.41 to
PGE-2, whereas CASEE at a concentration of 12.5 g/mL
exhibited a mean response of 450.08 to TNF-, 317.26 to
IL1-, and 729.66 to PGE-2. The study concluded that
CASEE is a promising anti-inflammatory agent due to its
efficacy and safety.
Citronellol is a naturally occurring acyclic
monoterpene alcohol found in the essential oil of
numerous plants, such as lime leaves (Citrus amblycarpa
(Hassk.) Ochse). It is often used in perfumes and
aromatherapy due to its flowery fragrance. Citronellol
has been utilized in traditional medicine for its anti-
inflammatory, analgesic, and antibacterial effects
(Santos et al., 2019). The medical benefits of lime leaves,
which are high in citronellol, include treating fever,
cough, and respiratory infections. According to previous
studies, citronellol has diverse pharmacological
activities, including anti-inflammatory, analgesic,
antipyretic, and anticancer activities. It is believed that
these features result from its capacity to decrease the
activity of cyclooxygenase (COX) enzymes, which are
responsible for the production of prostaglandins, which
are mediators of inflammation and pain (Desai et al.,
2018). The concentration of citronellol in lime leaves
varies by species and extraction technique. However, it
is typically in relatively high concentrations, making it a
plentiful supply of this vitally crucial active chemical.
Caryophyllene or (-)-β-Caryophyllene
Caryophyllene, or (-)-β-caryophyllene, is a natural,
whispery sesquiterpene compound in many essential
oils. This compound is seen on chromatograms with a
retention time of 12.98 minutes and a relative peak of
2.04%. Caryophyllene is a natural sesquiterpene in many
plants, including lime leaves (Citrus amblycarpa (Hassk.)
Ochse). It is often used in perfumes and flavorings
because of its spicy, woody scent. Caryophyllene has
been utilized for its anti-inflammatory, analgesic, and
anxiolytic effects in traditional medicine (Johnson et al.,
2020). Caryophyllene has been associated with several
positive pharmacological actions, including antioxidant,
anti-inflammatory, anticancer, cardioprotective,
hepatoprotective, gastroprotective, nephroprotective,
antibacterial, and immunomodulatory properties
(Machado, 2018). Caryophyllene has also been reported
to improve wound healing in a variety of ways
(Koyama, 2019; Parisotto-Peterle, 2020) and can be
considered a potential candidate for hyperoxaluria-
induced renal complication therapy (Xu & Yan, 2021),
premature lipid accumulation in preadipocyte 3T3-L1,
and improved glucose absorption in myotube C2C12
(Geddo, 2019).
The medical benefits of lime leaves, which are high
in caryophyllene, include treating fever, cough, and
respiratory infections. According to previous studies,
caryophyllene has numerous pharmacological activities,
including anti-inflammatory, analgesic, and antioxidant
characteristics. It is believed that these features are a
result of its capacity to interact with the
endocannabinoid system (ECS), a complex cell-signaling
system that regulates a variety of physiological and
cognitive processes, including pain, mood, and appetite
(Gallego-Landin et al., 2021; Stasiulewicz et al., 2020;
Zou & Kumar, 2018). Caryophyllene is exceptional

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
1999
among terpenes because it can interact with the CB2
receptor of the ECS, primarily expressed in immune cells
and tissues. This connection is believed to activate the
CB2 receptor, which controls the release of different
cytokines and other pain and inflammation mediators.
In addition to its effects on the ECS, caryophyllene
contains antibacterial characteristics that may be useful
in treating different illnesses (Handayani et al., 2019;
Lopez-Romero et al., 2015; Mulyani, 2014; Shetty et al.,
2016).
Hexadecanoic Acid, Ethyl Ester
Hexadecanoic acid, ethyl ester, is found in lime
leaves (Citrus amblycarpa (Hassk.) Ochse) as a fatty acid
ethyl ester. The chemical compound hexadecanoic acid,
ethyl ester, is found in the lime leaves of Citrus
amblycarpa (Hassk.) ochse (Adorjan & Buchbauer, 2010).
With a molecular weight of 284.4772, it is also known as
palmitic acid ethyl ester. The chemical is a form of fatty
acid ester found in other plants (Budiarto et al., 2017;
Kim et al., 2020). While research on the medical effects of
hexadecanoic acid ethyl ester is limited, studies have
demonstrated that other fatty acids contained in lime
leaves, such as palmitic acid, provide various health
benefits (Liu et al., 2022). Stearate acid is widely used as
a dietary supplement, cosmetics, and other industrial
products, such as plastics, soaps, and softened rubber
(Astiti & Ramona, 2021). The active component
hexadecanoic acid, ethyl ester, is responsible for the
larvicidal action against Aedes aegypti in the extract of
ethyl acetate leaves of Cassia uniflora Mill non-Spreng
(Toro, 2019). Hexadecanoic acid, ethyl ester, is also the
main component in Moringa oleifera leaf hydroalcoholic
extract and has antihelminthic activity against Trichuris
sp. and Ostertagia sp. (Pedraza-Hernández et al., 2021).
Hexadecanoic acid, a saturated fatty acid that is
abundant in lime leaves, has been discovered to possess
anti-inflammatory, antioxidant, and antidiabetic
characteristics (Adorjan & Buchbauer, 2010; Buathong &
Duangsrisai, 2023; Kasman et al., 2020; Liu et al., 2022).
It is hypothesized that these features result from its
ability to influence multiple signaling pathways,
including the NF-B pathway, a critical regulator of
inflammation (Cuevas-Cianca et al., 2023; Liu et al., 2021;
Rinkenbaugh & Baldwin, 2016). While there is no
specific information on the benefits of hexadecanoic
acid, ethyl ester in lime leaves, it probably possesses the
same qualities as other fatty acids in the plant. It may
also possess qualities that have not yet been identified.
1-Heptatriacotanol
1-Heptatriacotanol is a type of fatty alcohol found
in the leaves of Citrus amblycarpa (Hassk.) ochse, also
known as kaffir lime or makrut lime. The compound had
a retention duration of 19.87 minutes and a maximal
relative area of 4.58 percent. This chemical has been used
in traditional medicine to treat fever, coughs, and sore
throats, among other ailments. In traditional medicine,
1-heptatriacotanol is commonly consumed as a tea
brewed from the kaffir lime plant's leaves. Some
individuals also take it topically to treat skin diseases
such as eczema and psoriasis. Some research suggests
that 1-heptatriacotanol may possess anti-inflammatory
and antibacterial characteristics, although its mechanism
of action has yet to be entirely understood. It is believed
to function by limiting the generation of inflammatory
chemicals and preventing the growth of microbes (Dewi
et al., 2022).
1-Heptatriacotanol possesses antioxidant,
anticancer, and anti-inflammatory effects. Additionally,
1-heptatriacotanol is included in the ethanol extract of
Cnidoscolus aconitifolius leaves. It has been shown to
lower total cholesterol, LDL cholesterol, and
triglycerides while elevating HDL. It can treat coronary
heart disease (Ezinne et al., 2020). One of the essential
components of Pterocarpus cambodianus is the
antihypercholesterolemic compound 1-heptatriacotanol.
Additionally, obtained from Caralluma retrospections
exudate, 1-heptatriacotanol may have antibacterial
effects (Makeen, 2020); it is also derived from Aloe
fleurentiniorum exudate and is utilized in traditional
medicine (Moni et al., 2021). It has been identified in
Nigella sativa as an antioxidant, anticancer, and anti-
inflammatory agent (Hadi, 2016).
Phytol
Phytol is a plant-based acyclic hydrogenated
diterpene alcohol that is a component of chlorophyll.
Phytol compounds were observed at a retention time of
21.06 with a relative peak area of 7.50%. It is one of the
plant's many phytochemicals that provide various
health advantages. Phytol is a valuable essential oil used
as a fragrance with potential pharmacological and
biotechnology applications. Phytol and its derivatives
have been demonstrated to have pharmacological
applications in the central nervous system, including
anxiolytic and neuroprotective properties (Islam et al.,
2018). In recent years, Phytol's possible health
advantages have garnered increased attention. Studies
indicate that it possesses antioxidant, anti-inflammatory,
antidiabetic, and analgesic activities and may also have
insect-repellent capabilities (Agustina et al., 2022;
Tambunan et al., 2020).
In addition, it may possess antibacterial properties,
making it practical for combating bacterial and other
microbial illnesses (Islam et al., 2018). Phytol's
antioxidant, anti-inflammatory, and analgesic effects
have been demonstrated (Hassan et al., 2021). It also has
insect-repellent potential. According to previous
studies, phytol may help reduce inflammation in the

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
2000
body, which is considered to play a role in various
diseases, including arthritis and cardiovascular disease.
Phytol also has analgesic properties, suggesting that it
may aid in alleviating pain. This could make it a possible
alternative to conventional painkillers. In addition, some
research suggests that Phytol may have antibacterial
characteristics, indicating that it can combat diseases
caused by bacteria and other microbes (Dewi et al., 2022;
Islam et al., 2018). Phytol is also reported to have the
ability to act as an antimalarial (Saxena, 2018). Phytol
suppresses P. berghei and can correct some pathological
changes induced by P. berghei (Usman, 2021). Phytol
decreases clinical symptoms in experimental
autoimmune encephalomyelitis by regulating NADPH
oxidase two expression (Blum et al., 2018). Phytol has
been found to have anticancer capabilities by regulating
molecular marker expression in human embryonic
carcinoma NCCIT cells (Soltanian, 2020).
Ethyl 9,12,15-Octadecatrienoate
The chemical compound ethyl 9,12,15-
octadecatrienoate belongs to the family of fatty acid
esters. It is also known as ethyl linoleate and is typically
found in linseed and soybean, among other plant oils.
Ethyl 9,12,15-octadecatrienoate, also called ethyl
linoleate, is a long chain of ethyl ester fatty acids made
when the carboxy group of linolenic acid combines with
the hydroxy group of ethanol. Ethyl 9,12,15-
octadecatrienoate is a polyunsaturated fatty acid with
multiple double bonds in its carbon chain. Specifically,
it contains three double bonds at positions 9, 12, and 15
in the carbon chain. Ethyl 9,12,15-octadecatrienoate, a
polyunsaturated fatty acid, has demonstrated anti-
inflammatory and neuroprotective characteristics,
which may lessen the risk of chronic diseases such as
heart disease, stroke, and Alzheimer's disease (Devassy
et al., 2016; Ganesh & Mohankumar, 2017; Giacobbe et
al., 2020; Sianipar et al., 2016). Additionally, it may boost
mood and cognitive performance. In addition, some
research suggests that ethyl 9,12,15-octadecatrienoate
may have anticancer potential (Márquez-Fernández &
Camargo, 2019). In animal cancer models, linolenic acid,
a polyunsaturated fatty acid comparable to ethyl 9,12,15-
octadecatrienoate, was reported to have antitumor
effects in one study. Additional research is needed to
completely comprehend the potential advantages and
mechanisms of action of ethyl 9,12,15-octadecatrienoate
as a traditional medicine when found in lime leaves.
Methyl glycocholate, 3TMS Derivative
Methyl glycocholate, a 3TMS derivative, is a
chemical molecule discovered in the ethanol extract of
lime leaves of Citrus amblycarpa (Hassk.)(Budiarto et al.,
2017; Kasman et al., 2020). Methyl glycocholate
compound, a 3TMS derivative, belongs to the group of
terpenoids with antimicrobial activity. The compound
had a retention time of 24.57 minutes and a 3.59% area
peak. Methyl glycocholate, a 3TMS derivative, has also
been identified from green coffee bean hexane extract,
which can act as an antioxidant (Rosiak, 2020). In
addition, methyl glycocholate, a 3TMS derivative of lime
leaf ethanol extract, has potential as a larvicide and
bioinsecticide since it supplies contact and respiratory
toxins to the insect body. Due to the acidic nature of this
leaf, fragrant aromatics emit clearly, which has
ramifications for mosquito mortality and irritation; as a
result, it is efficiently utilized as an insecticide agent, but
more research is required (Kasman et al., 2020).
3,7-dimethyl oct-6-en-1-yl stearate
3 The chemical compound 3,7-dimethyloct-6-en-1-
yl stearate belongs to the class of fatty acid esters. It
consists of a molecule of stearic acid (a long-chain
saturated fatty acid) and a group of 3,7-dimethyloct-6-
en-1-yl (a branched unsaturated hydrocarbon chain)
(Sammi et al., 2016). Compound 3.7-dimethyloct-6-en-1-
yl stearate is a fragmentation of citronella seen at a
retention time of 26.45 with a relative area peak of
11.45%. Another name for this compound is citronellyl
stearate. Due to their propensity to combine with water
and oil, fatty acid esters are frequently employed in the
food and cosmetic industries as emulsifiers, thickeners,
and stabilizers (Watanabe et al., 2017). It is well known
for its emollient and conditioning characteristics, which
enhance the texture and look of the skin. This chemical
has been isolated from various natural sources,
including lime leaves (Citrus amblycarpa (Hassk.) ochse),
utilized in traditional medicine for possible health
advantages. In addition, they may have therapeutic
qualities, such as anti-inflammatory and antibacterial
activities (Sammi et al., 2016). However, the exact effects
of 3,7-dimethyloct-6-en-1-yl stearate still need to be fully
understood, and additional research is required to
evaluate its possible applications in traditional
medicine.
Ethyl iso-allocholate
The chemical compound ethyl iso-allocholate
belongs to the class of sterols. It has a similar structure
to other sterols, such as sitosterol and stigmasterol, and
is derived from cholesterol. Ethyl iso-allocholate has
been discovered in various natural sources, including
lime (Citrus aurantifolia) leaves, utilized in traditional
medicine for their possible health advantages. However,
there needs to be more investigation on the possible
advantages and mechanisms of action of this chemical as
a traditional medicine. According to several studies,
ethyl iso-allocholate is a steroid derivative that acts as an
antibacterial, diuretic, anti-inflammatory, and anti-
asthmatic agent (Muthulakshmi et al., 2012).

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
2001
Additionally, ethyl iso-allocholate was isolated from
traditional rice karungkavuni and is a selective inhibitor
of the enzyme dihydropteroate synthase in Escherichia
coli (Malathi et al., 2016). Phyllanthus nivosus contains
ethyl iso-allocate, which possesses anti-inflammatory
properties due to its high affinity for the target protein
caspase-1 (Johnson, 2020). Ipomoea obscura (L.) Ethyl iso-
allocholate has also been isolated and is known to have
the capacity to prevent the attachment of the SARS-CoV
viral genome to the target proteins angiotensin-
converting enzyme 2 (ACE2) and major protease (MPro)
(Poochi et al., 2020).
Rhodopin
Rhodopin has been isolated from the ethanol
extract of lime leaves (Citrus amblycarpa (Hassk.) ochse)
as a flavonoid. Flavonoids are recognized for their
antioxidant and anti-inflammatory qualities and are
commonly found in fruits, vegetables, and medicinal
plants, among other natural sources (Adnyana &
Sudaryati, 2023). Lime leaves have been utilized in
traditional medicine for their potential health
advantages, including as an anti-inflammatory agent, to
help digestion and to ease headaches. Rhodopin's
presence in the ethanol extract of lime leaves may
contribute to these possible medicinal effects. Rhodopin
possesses antioxidant characteristics, which may protect
against oxidative stress and reduce inflammation in the
body. Some studies suggest that it may limit the growth
and spread of cancer cells (Cuevas-Cianca et al., 2023;
Desai et al., 2018; Lim, 2012; Liu et al., 2022; Sammi et al.,
2016).
Several studies have indicated that rhodopin
compounds belong to the carotenoid group with
antioxidant activity (Hassan et al., 2021). Carotenoids
are a class of fat-soluble pigments that give various fruits
and vegetables in our daily meals their beautiful color,
including carrots, tomatoes, pumpkins, peppers, and
dark green leafy vegetables (Xavier & Pérez-Gálvez,
2016). Rhodopin is also found in photosynthetic bacteria
such as Rhodopseudomonas palustris (Faith-Anthony et al.,
2014), Rhodoplanes sp. (Takaichi et al., 2012),
Thermochromatium tepidum (Niedzwiedzki et al., 2011),
and Phaeospirillum oryzae (Lakshmi et al., 2011).
Rhodopin can act as an antioxidant and decrease the
proliferation of cells with low toxicity in carcinoma cell
cultures (Astiti & Ramona, 2021). While additional
research is required to completely comprehend the
potential advantages and mechanisms of action of
rhodopin as a traditional medicine, the substance's
antioxidant and anti-inflammatory qualities make it a
viable option for further investigation.
Tricyclo[20.8.0.0(7,16)]triacontane, 1(22),7(16)-diepoxy-
Tricyclo[20.8.0.0(7,16)] The chemical compound
1(22),7(16)-diepoxy-triacontane belongs to the class of
triterpenes. It is a cyclic hydrocarbon with a highly
branched structure found in various natural sources.
Tricyclo compounds [20.8.0.0 (7,16)] triacontane, 1 (22)
and 7 (16)-diepoxy-are compounds that have
antioxidant and antibacterial activity, seen at a retention
time of 28.97 with a relative peak area of 9.24%. As a
traditional medicine, little is known about the possible
advantages and mechanisms of action of
tricyclo[20.8.0.0(7,16)]tricontane,1(22),7(16)-diepoxy-.
Triterpenes have been reported to have anti-
inflammatory, anticancer, and antiviral effects, among
other potential medicinal applications (Budiarto et al.,
2021b; Budiarto & Sholikin, 2022; Khan et al., 2023).
Conclusion
Based on research findings using gas
chromatography‒mass spectrometry (GCMS), it was
shown that ethanol extracts of lime leaves (Citrus
amblycarpa (Hassk.) Ochse) contains 75 compound
components dominated by 11 principal components,
including citronellol, caryophyllene, hexadecanoic acid,
ethyl ester, 1-heptatriacotanol, Phytol, ethyl 9,12,15-oate;
methyl glycolate, a derivative of 3TMS; 3,7-dimethyloct-
6-en-1-yl stearate; methyl iso-allocholate; rhopin; and
tricyclo [20.8.0.0(7,16) triacontane, 1(22),7(16)-diepoxy-
The results of the content evaluation show that the
compound components in lime leaves have the potential
to be developed as traditional medicines for both
degenerative and infectious diseases. However, further
research is needed precisely to identify the therapeutic
effects of using compounds on lime leaves. Lime leaves
in Bali have been widely used as antioxidants, anticancer
agents, anti-inflammatory agents, and flavor enhancers
for food. In the future, further research is needed to
comprehensively evaluate the compound content of
lime leaves for their potential as candidates for
traditional medicine, especially in Bali.
Acknowledgments
The author wishes to thank the Hindu University of Indonesia
for sponsoring this research Additionally, we would like to
thank Gadjah Mada University's Integrated Research and
Testing Laboratory for assisting with sample testing.
Author Contributions
I.N.A: conceptualization, supervisor; N.K.A.J: resources,
commentary; A.A.A.S.S.W: data curation. All authors
performed initial manuscript writing, and final manuscript
approval.

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
2002
Funding
This research was funded through an internal grant from
Universitas Hindu Indonesia Denpasar for the 2021 fiscal year
with contract number 259B/LPPM/UNHI/VIII/2021.
Conflicts of Interest
The authors declare that there is no conflict of interest.
References
Abdullah, S. S. S., & Othman, M. K. I. (2018). Potential of
Kaffir Lime (Citrus hystrix) Peel Essential Oil as a
Cockroach Repellent. In Sustainable Technologies for
the Management of Agricultural Wastes (pp. 131–141).
Springer Singapore. https://doi.org/10.1007
/978-981-10-5062-6_10
Adnyana, I. M. D. M., & Sudaryati, N. L. G. (2023).
Phytochemical Analysis of the Antioxidant
Compounds of Baper Tea and Its Potential as an
Immunomodulatory agent and Candidate for
Standardized Herbal Medicine. Trends in Sciences,
20(1), 6391. https://doi.org/10.48048/tis.2023.
6391
Adnyana, I. M. D. M., Sudaryati, N. L. G., & Sitepu, I.
(2021). Toxicity of Legiayu incense as Insecticide
and Larvicide against Aedes aegypti Mosquitoes
Mortality. Indonesian Journal of Pharmacy, 32(4),
524–521. https://doi.org/10.22146/ijp.1814
Adorjan, B., & Buchbauer, G. (2010). Biological
Properties of Essential Oils: an Updated Review.
Flavour and Fragrance Journal, 25(6), 407–426.
https://doi.org/10.1002/ffj.2024
Agustina, E., Leksono, A. S., Gama, Z. P., Yanuwiadi, B.,
& Sukma, M. (2022). Potential of House Yard Plants
as an Alternative for Dengue Vector Control in the
Tsunami Area Settlement of Banda Aceh City.
Journal of Applied Pharmaceutical Science, 12(11),
122–137. https://doi.org/10.7324/JAPS.
2022.121114
Anuchapreeda, S., Chueahongthong, F.,
Viriyaadhammaa, N., Panyajai, P., Anzawa, R.,
Tima, S., Ampasavate, C., Saiai, A., Rungrojsakul,
M., Usuki, T., & Okonogi, S. (2020). Antileukemic
Cell Proliferation of Active Compounds from
Kaffir Lime (Citrus hystrix) Leaves. Molecules,
25(6), 1300. https://doi.org/10.3390/molecules25
061300
Astiti, N. P. A., & Ramona, Y. (2021). GC-MS Analysis of
Active and Applicable Compounds in Methanol
Extract of Sweet Star Fruit (Averrhoa carambola l.)
Leaves. HAYATI Journal of Biosciences, 28(1), 12–22.
https://doi.org/10.4308/hjb.28.1.12
Blum, L., Tafferner, N., Spring, I., Kurz, J., DeBruin, N.,
Geisslinger, G., Parnham, M. J., & Schiffmann, S.
(2018). Dietary Phytol Reduces Clinical Symptoms
in Experimental Autoimmune Encephalomyelitis
(EAE) at Least Partially by Modulating NOX2
Expression. Journal of Molecular Medicine, 96(10),
1131–1144. https://doi.org/10.1007/s00109-018-
1689-7
Buathong, R., & Duangsrisai, S. (2023). Plant Ingredients
in Thai Food: a Well-Rounded Diet for Natural
Bioactive Associated with Medicinal Properties.
PeerJ, 11, e14568. https://doi.org/10.
7717/peerj.14568
Budiarto, R., & Sholikin, M. M. (2022). Kaffir Lime
Essential Oil Variation in the Last Fifty Years: A
Meta-Analysis of Plant Origins, Plant Parts and
Extraction Methods. Horticulturae, 8(12), 1132.
https://doi.org/10.3390/horticulturae8121132
Budiarto, R., Poerwanto, R., Santosa, E., & Efendi, D.
(2017). The Potentials of Limau (Citrus amblycarpa
Hassk. Ochse) as A Functional Food and
Ornamental Mini Tree Based on Metabolomic and
Morphological Approaches. Journal of Tropical Crop
Science, 4(2), 49–57. https://doi.org/10.
29244/jtcs.4.2.49-57
Budiarto, R., Poerwanto, R., Santosa, E., & Efendi, D.
(2021a). Morphological Evaluation and
Determination Keys of 21 Citrus Genotypes at
Seedling Stage. Biodiversitas Journal of Biological
Diversity, 22(3), 1570–1579.
https://doi.org/10.13057/biodiv/d220364
Budiarto, R., Poerwanto, R., Santosa, E., Efedi, D., &
Agusta, A. (2021b). Sensory Evaluation of the
Quality of Kaffir Lime (Citrus hystrix DC.) Leaves
Exposed to Different Postharvest Treatments.
Journal of Tropical Crop Science, 8(02), 71–79.
https://doi.org/10.29244/jtcs.8.02.71-79
Budiarto, R., Poerwanto, R., Santosa, E., Efendi, D., &
Agusta, A. (2022). Comparative and Correlation
Analysis of Young and Mature Kaffir Lime (Citrus
hystrix DC) Leaf Characteristics. International
Journal of Plant Biology, 13(3), 270–280.
https://doi.org/10.3390/ijpb13030023
Cuevas-Cianca, S. I., Romero-Castillo, C., Gálvez-
Romero, J. L., Juárez, Z. N., & Hernández, L. R.
(2023). Antioxidant and Anti-Inflammatory
Compounds from Edible Plants with Anti-Cancer
Activity and Their Potential Use as Drugs.
Molecules, 28(3), 1488. https://doi.org/10.3390/
molecules28031488
Darwin, M., Mamondol, M. R., Sormin, S. A., Nurhayati,
Y., Tambunan, H., Sylvia, D., Adnyana, I. M. D. M.,
Prasetiyo, B., Vianitati, P., & Gebang, A. A. (2021).
Quantitative Approach Research Method (T. S.
Tambunan, Ed.; 1st ed.). CV Media Sains
Indonesia.
Dertyasasa, E. D., & Tunjung, W. A. S. (2017). Volatile
Organic Compounds of Kaffir Lime (Citrus hystrix

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
2003
DC.) Leaves Fractions and their Potency as
Traditional Medicine. Biosciences, Biotechnology
Research Asia, 14(4), 1235–1250. https://doi.org/
10.13005/bbra/2566
Desai, S. J., Prickril, B., & Rasooly, A. (2018). Mechanisms
of Phytonutrient Modulation of Cyclooxygenase-2
(COX-2) and Inflammation Related to Cancer.
Nutrition and Cancer, 70(3), 350–375.
https://doi.org/10.1080/01635581.2018.1446
091
Devassy, J. G., Leng, S., Gabbs, M., Monirujjaman, M., &
Aukema, H. M. (2016). Omega-3 Polyunsaturated
Fatty Acids and Oxylipins in Neuroinflammation
and Management of Alzheimer Disease. Advances
in Nutrition, 7(5), 905–916.
https://doi.org/10.3945/an.116.012187
Dewi, K. S., Julaeha, E., Farabi, K., & Supratman, U.
(2022). Flavonoids from Limau Peel (Citrus
amblycarpa (Hassk.) Ochse) and Their Antioxidant
Activity. Molekul, 17(2), 193.
https://doi.org/10.20884/1.jm.2022.17.2.5430
Ezinne, C., Osuocha, K. U., & Iwueke, A. V. (2020).
Phytochemical Profiling, Body Weight Effect and
Anti-Hypercholesterolemia Potentials of
Cnidoscolus aconitifolius Leaf Extracts in Male
Albino Rat. Journal of Pharmacognosy and
Phytotherapy, 12(2), 19–27. https://doi.org/10.
5897/jpp2016.0436
Faith-Anthony, A. O., Ibrahim, N., & Hamzah, A. (2014).
Clinical potentials of bacteriocarotenoids:
Rhodopin and β-carotene from phototrophic
Rhodopseudomonas palustris. IOSR J Dental Med
Sci, 13(12), 52-58. Retrieved from
https://www.iosrjournals.org/iosr-
jdms/papers/Vol13-issue12/Version-
8/M0131285258.pdf
Felim, J., Sim, M., Wijaya, S., Susanto, C., Sinamo, S., &
Florenly. (2021). A Promising Anti-Inflammatory
Drugs from Citrus Amblycarpa (Hassk.) Ochse
Seeds. 2021 IEEE International Conference on Health,
Instrumentation & Measurement, and Natural Sciences
(InHeNce), 1–6. https://doi.org/10.1109/
InHeNce52833.2021.9537212
Gallego-Landin, I., García-Baos, A., Castro-Zavala, A., &
Valverde, O. (2021). Reviewing the Role of the
Endocannabinoid System in the Pathophysiology
of Depression. Frontiers in Pharmacology, 12, 762738.
https://doi.org/10.3389/fphar.2021.76273
8
Ganesh, M., & Mohankumar, M. (2017). Extraction and
Identification of Bioactive Components in Sida
cordata (Burm.f.) Using Gas Chromatography–
Mass Spectrometry. Journal of Food Science and
Technology, 54(10), 3082–3091. https://doi.org/10.
1007/s13197-017-2744-z
Geddo, F. (2019). Pipenig®-FL, a Fluid Extract of Black
Pepper (Piper nigrum L.) with a High
Standardized Content of Trans-β-Caryophyllene,
Reduces Lipid Accumulation in 3T3-L1
Preadipocytes and Improves Glucose Uptake in
C2C12 Myotubes. Nutrients, 11(11).
https://doi.org/10.3390/nu11112788
Giacobbe, J., Benoiton, B., Zunszain, P., Pariante, C. M.,
& Borsini, A. (2020). The Anti-Inflammatory Role
of Omega-3 Polyunsaturated Fatty Acids
Metabolites in Pre-Clinical Models of Psychiatric,
Neurodegenerative, and Neurological Disorders.
Frontiers in Psychiatry, 11, 22. https://doi.org/10.
3389/fpsyt.2020.00122
Hadi, M. (2016). Analysis of Bioactive Chemical
Compounds of Nigella sativa Using Gas
Chromatography-Mass Spectrometry. Journal of
Pharmacognosy and Phytotherapy, 8(2), 8–24.
https://doi.org/10.5897/JPP2015.0364
Handayani, E. V. A. S. R. I., Nugraheni, E. R., &
Susilowati, A. R. I. (2019). Antibacterial and
Antifungal Activities of Essential Oil of
Tawangmangu Sweet Orange (Citrus sinensis)
Peel at Different Altitudes. Asian Journal of Natural
Product Biochemistry, 17(1), 47–54. https://doi.org
/10.13057/biofar/f170105
Harmayani, E., Anal, A. K., Wichienchot, S., Bhat, R.,
Gardjito, M., Santoso, U., Siripongvutikorn, S.,
Puripaatanavong, J., & Payyappallimana, U.
(2019). Healthy Food Traditions of Asia:
Exploratory Case Studies from Indonesia,
Thailand, Malaysia, and Nepal. Journal of Ethnic
Foods, 6, 1. https://doi.org/10.1186/s42779-019-
0002-x
Hassan, S. M., Ashour, M., Soliman, A. A. F., Hassanien,
H. A., Alsanie, W. F., Gaber, A., & Elshobary, M. E.
(2021). The Potential of a New Commercial
Seaweed Extract in Stimulating Morpho-
Agronomic and Bioactive Properties of Eruca
vesicaria (L.) cav. Sustainability (Switzerland), 13(8).
https://doi.org/10.3390/su13084485
Hien, T. T., Quyen, N. T. C., Minh, P. T. H., & Le, X. T.
(2020). Determine the Components of Kaffir Lime
Oil (Citrus Hystrix DC.) in the Microwave-Assisted
Extraction Process. IOP Conference Series: Materials
Science and Engineering, 991(1), 012018.
https://doi.org/10.1088/1757-899X/991/1/01201
8
Ho, Y. (2020). Anticancer Effect of Citrus hystrix dc. Leaf
Extract and Its Bioactive Constituents Citronellol
and, Citronellal on the Triple Negative Breast
Cancer MDA-MB-231 Cell Line. Pharmaceuticals,
13(12), 1–17. https://doi.org/10.
3390/ph13120476

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
2004
Islam, M. T., Ali, E. S., Uddin, S. J., Shaw, S., Islam, M.
A., Ahmed, M. I., Chandra Shill, M., Karmakar, U.
K., Yarla, N. S., Khan, I. N., Billah, M. M.,
Pieczynska, M. D., Zengin, G., Malainer, C.,
Nicoletti, F., Gulei, D., Berindan-Neagoe, I.,
Apostolov, A., Banach, M., … Atanasov, A. G.
(2018). Phytol: A Review of Biomedical Activities.
Food and Chemical Toxicology, 121, 82–94.
https://doi.org/10.1016/j.fct.2018.08.032
Johnson, S. A., Rodriguez, D., & Allred, K. (2020). A
Systematic Review of Essential Oils and the
Endocannabinoid System: A Connection Worthy
of Further Exploration. Evidence-Based
Complementary and Alternative Medicine, 1–13.
https://doi.org/10.1155/2020/8035301
Johnson, T. O. (2020). Biochemical Evaluation and
Molecular Docking Assessment of the Anti-
Inflammatory Potential of Phyllanthus nivosus
Leaf against Ulcerative colitis. Heliyon, 6(5).
https://doi.org/10.1016/j.heliyon.2020.e03893
Juliasih, N. K. A., & Adnyana, I. M. D. M. (2023).
Utilization of Pteridophyta Species in Cyathea
Park, Bali, as Traditional Medicine Agents: A Field
Study and Meta-Synthesis Review. Notulae Scientia
Biologicae, 15(2), 11522.
https://doi.org/10.55779/nsb15211522
Kasman, K. (2020). Identification of Active Compounds
of Ethanol Extract of Citrus Amblycarpa Leaves by
Analysis of Thin-Layer Chromatography and Gas
Chromatography-Mass Spectrometry as
Bioinsecticide Candidates for Mosquitoes. Open
Access Macedonian Journal of Medical Sciences, 8, 1–6.
https://doi.org/10.3889/OAMJMS.2020.5207
Kasman, K., Ishak, N. I., Hastutiek, P., & Suprihati, E.
(2021). Potential Extract Ethanol Citrus
Amblycarpa as a Bioinsecticide against Aedes
aegypti Larvae. Systematic Reviews in Pharmacy,
12(1), 1614–1618.
Kasman, K., Ishak, N. I., Hastutiek, P., Suprihati, E., &
Mallongi, A. (2020). Identification of Active
Compounds of Ethanol Extract of Citrus
Amblycarpa Leaves by Analysis of Thin-layer
Chromatography and Gas Chromatography-Mass
Spectrometry as Bioinsecticide Candidates for
Mosquitoes. Open Access Macedonian Journal of
Medical Sciences, 8(T2), 1–6. https://doi.org/10.
3889/oamjms.2020.5207
Khan, M. Z., Azad, A. K., Jan, S., Safdar, M., Bibi, S.,
Majid, A. M. S. A., Albadrani, G. M., Nouh, N. A.
T., Abdulhakim, J. A., & Abdel-Daim, M. M. (2023).
An Experimental and Computational Analysis of
Plant Compounds from Whole Urtica dioica L.
Plant’s Essential Oil for Antioxidant and
Antibacterial Activities. Metabolites, 13(4), 502.
https://doi.org/10.3390/metabo13040502
Kim, B.-R., Kim, H. M., Jin, C. H., Kang, S.-Y., Kim, J.-B.,
Jeon, Y. G., Park, K. Y., Lee, I.-S., & Han, A.-R.
(2020). Composition and Antioxidant Activities of
Volatile Organic Compounds in Radiation-Bred
Coreopsis Cultivars. Plants, 9(6), 717.
https://doi.org/10.3390/plants9060717
Koyama, S. (2019). Beta-Caryophyllene Enhances
Wound Healing Through Multiple Routes. PLoS
ONE, 14(12). https://doi.org/10.1371/journal.
pone.0216104
Kusumawati, I. G. A. W., Putra, I. M. W. A., &
Yogeswara, I. B. A. (2021). In Vitro ACE Inhibitory
Activity and Bioactive Compounds of Aqueous
Extract of Citrus amblycarpa. Traditional Medicine
Journal, 26(2), 118–122.
https://doi.org/10.22146/mot.65028
Lakshmi, K. V. N. S., Sasikala, C., Takaichi, S., &
Ramana, C. V. (2011). Phaeospirillum oryzae sp.
nov., a Spheroplastforming, Phototrophic
Alphaproteobacterium from a Paddy Soil.
International Journal of Systematic and Evolutionary
Microbiology, 61(7), 1656–1661. https://doi.org/
10.1099/ijs.0.025544-0
Lim, T. K. (2012). Citrus amblycarpa. In Edible Medicinal
and Non-Medicinal Plants (pp. 623–624). Springer
Netherlands. https://doi.org/10.1007/978-94-007
-4053-2_72
Liu, S., Lou, Y., Li, Y., Zhang, J., Li, P., Yang, B., & Gu, Q.
(2022). Review of Phytochemical and Nutritional
Characteristics and Food Applications of Citrus L.
Fruits. Frontiers in Nutrition, 9, 968604.
https://doi.org/10.3389/fnut.2022.968604
Liu, Z., Ren, Y., Meng, L., Li, L., Beatson, R., Deng, J.,
Zhang, T., Liu, J., & Han, X. (2021). Epigenetic
Signaling of Cancer Stem Cells During
Inflammation. Frontiers in Cell and Developmental
Biology, 9, 772211. https://doi.org/10.3389/fcell.
2021.772211
Lopez-Romero, J. C., González-Ríos, H., Borges, A., &
Simões, M. (2015). Antibacterial Effects and Mode
of Action of Selected Essential Oils Components
against Escherichia coli and Staphylococcus
aureus. Evidence-Based Complementary and
Alternative Medicine, 1–9. https://doi.org/
10.1155/2015/795435
Lu, J. X. (2019). Citronellal Prevents Endothelial
Dysfunction and Atherosclerosis in Rats. Journal of
Cellular Biochemistry, 120(3), 3790–3800. https://
doi.org/10.1002/jcb.27660
Lubinska-Szczygieł, M., Różańska, A., Namieśnik, J.,
Dymerski, T., Shafreen, R. B., Weisz, M., Ezra, A.,
& Gorinstein, S. (2018). Quality of Limes Juices
Based on the Aroma and Antioxidant Properties.
Food Control, 89, 270–279.
https://doi.org/10.1016/j.foodcont.2018.02.005

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
2005
Machado, K. D. C. (2018). A Systematic Review on the
Neuroprotective Perspectives of Beta-
Caryophyllene. Phytotherapy Research 32(12), 2376–
2388. https://doi.org/10.1002/ptr.6199
Makeen, H. A. (2020). Documentation of Bioactive
Principles of the Exudate Gel (EG) from the Stem
of Caralluma retrospiciens (Ehrenb) and in Vitro
Antibacterial Activity – Part A. Arabian Journal of
Chemistry, 13(8), 6672–6681. https://doi.org/10.
1016/j.arabjc.2020.06.022
Malathi, K., Anbarasu, A., & Ramaiah, S. (2016). Ethyl
Iso-Allocholate from a Medicinal Rice
Karungkavuni Inhibits Dihydropteroate Synthase
in Escherichia coli: A Molecular Docking and
Dynamics Study. Indian Journal of Pharmaceutical
Sciences, 78(6), 780–788. https://doi.org/10.
4172/pharmaceutical-sciences.1000184
Márquez-Fernández, M. E., & Camargo, M. (2019).
Therapeutic Potential of Omega Fatty Acids in
Breast Cancer. Review. Revista Vitae, 26(1), 23–43.
https://doi.org/10.17533/udea.vitae.v26n1a04
Moni, S. S., Sultan, M. H., Makeen, H. A., Madkhali, O.
A., Bakkari, M. A., Alqahtani, S. S., Alshahrani, S.,
Menachery, S. J., Alam, M. I., Elmobark, M. E., ur
Rehman, Z., Alam, M. S., Faqihi, A. Y., Mogaidi, H.
M., Khobrani, H., Albana, E., & Sharahily, A. M.
(2021). Bioactive Principles in Exudate Gel from the
Leaf of Aloe fleurentiniorum, Traditionally Used
as Folkloric Medicine by Local People of Aridah
and Fayfa Mountains, Saudi Arabia. Arabian
Journal of Chemistry, 14(11), 103400.
https://doi.org/10.1016/j.arabjc.2021.103400
Mulyani, S. (2014). Antibacterial Activity and GC-MS
Analysis of the Citrus amblycarpa (Hassk) Ochse
Essential Oil. Indonesian Journal of
PharmacyARMACY, 20(3), 127–132. https://doi.
org/10.14499/indonesianjpharm20iss3pp127-132
Muthulakshmi, A., Jothibai-Margret, R., & Mohan, V. R.
(2012). GC-MS Analysis of Bioactive Components
of Feronia elephantum Correa (Rutaceae). Journal
of Applied Pharmaceutical Science, 2(2), 69–74.
Retrieved from
https://japsonline.com/abstract.php?article_id=3
82
Niedzwiedzki, D. M., Fuciman, M., Kobayashi, M.,
Frank, H. A., & Blankenship, R. E. (2011). Ultrafast
Time-Resolved Spectroscopy of the Light-
Harvesting Complex 2 (LH2) from the
Photosynthetic Bacterium Thermochromatium
tepidum. Photosynthesis Research, 110(1), 49–60.
https://doi.org/10.1007/s11120-011-9692-7
Ollitrault, P., Curk, F., & Krueger, R. (2020). Citrus
Taxonomy. In The Genus Citrus (Issue 1943, pp. 57–
81). Elsevier Inc. https://doi.org/10.1016/
B978-0-12-812163-4.00004-8
Panakkal, E. J., Tantayotai, P., Adediran, A. A.,
Phusantisampan, T., & Sriariyanun, M. (2021).
Chemical Profiles of Herbal Compress and Their
Persistence Affected by Steaming and Storage
Time. Journal of Chemistry, 2021, 1–10.
https://doi.org/10.1155/2021/2743855
Parisotto-Peterle, J. (2020). Healing Activity of Hydrogel
Containing Nanoemulsified β-Caryophyllene.
European Journal of Pharmaceutical Sciences, 148.
https://doi.org/10.1016/j.ejps.2020.105318
Pedraza-Hernández, J., Elghandour, M. M. M. Y.,
Khusro, A., Salem, M. Z. M., Camacho-Diaz, L. M.,
Barbabosa-Pliego, A., & Salem, A. Z. M. (2021).
Assessment on Bioactive Role of Moringa Oleifera
Leaves as Anthelmintic Agent and Improved
Growth Performance in Goats. Tropical Animal
Health and Production, 53(2).
https://doi.org/10.1007/s11250-021-02745-9
Poochi, S. P., Easwaran, M., Balasubramanian, B.,
Anbuselvam, M., Meyyazhagan, A., Park, S.,
Bhotla, H. K., Anbuselvam, J., Arumugam, V. A.,
Keshavarao, S., Kanniyappan, G. V., Pappusamy,
M., & Kaul, T. (2020). Employing Bioactive
Compounds Derived from Ipomoea obscura (L.) to
Evaluate Potential Inhibitor for SARS‐CoV‐2 Main
Protease and ACE2 Protein. Food Frontiers, 1(2),
168–179. https://doi.org/10.1002/fft2.29
Putra, G. M. D., Satriawati, D. A., Astuti, N. K. W., &
Yadnya-Putra, A. A. G. R. (2018). Standarisasi dan
Skrining Fitokimia Ekstrak Etanol 70% Daun Jeruk
Limau (Citrus amblycarpa (Hassk.) Osche). Jurnal
Kimia, 187. https://doi.org/10.24843/jchem
.2018.v12.i02.p15
Rinkenbaugh, A., & Baldwin, A. (2016). The NF-κB
Pathway and Cancer Stem Cells. Cells, 5(2), 16.
https://doi.org/10.3390/cells5020016
Rosiak, A. (2020). Antioxidant properties of green coffee
extract. Forests, 11(5), 557.
https://doi.org/10.3390/f11050557
Sammi, S. R., Mishra, D. P., Trivedi, S., Smita, S. S.,
Nagar, A., Tandon, S., & Pandey, R. (2016). Citrus
hystrix-derived 3,7-dimethyloct-6-enal and 3,7-
dimethyloct-6-enyl acetate ameliorate
acetylcholine deficits. RSC Advances, 6(73), 68870–
68884. https://doi.org/10.1039/C6RA12522K
Santos, P. L., Matos, J. P. S. C. F., Picot, L., Almeida, J. R.
G. S., Quintans, J. S. S., & Quintans-Júnior, L. J.
(2019). Citronellol, a Monoterpene Alcohol with
Promising Pharmacological Activities - A
Systematic Review. Food and Chemical Toxicology,
123, 459–469. https://doi.org/10.1016/j.fct.2018.
11.030
Saxena, A. (2018). Antimalarial Activity of Phytol
Derivatives: in Vitro and in Vivo Study. Medicinal

Jurnal Penelitian Pendidikan IPA (JPPIPA)
April 2024, Volume 10, Issue 4, 1994-2006
2006
Chemistry Research, 27(5), 1345–1354.
https://doi.org/10.1007/s00044-017-2132-2
Shetty, S., Mahin-Syed-Ismail, P., Varghese, S., Thomas-
George, B., Kandathil- Thajuraj, P., Baby, D.,
Haleem, S., Sreedhar, S., & Devang-Divakar, D.
(2016). Antimicrobial Effects of Citrus sinensis Peel
Extracts against Dental Caries Bacteria: An in Vitro
Study. Journal of Clinical and Experimental Dentistry,
8(1), e71–e77. https://doi.org/10.4317/
jced.52493
Sianipar, N. F., Purnamaningsih, R., & Rosaria, R. (2016).
Bioactive Compounds of Fourth Generation
Gamma-Irradiated Typhonium flagelliforme
Lodd. Mutants Based on Gas Chromatography-
Mass Spectrometry. IOP Conference Series: Earth and
Environmental Science, 41, 012025.
https://doi.org/10.1088/1755-1315/41/1/012025
Siti, H. N., Mohamed, S., & Kamisah, Y. (2022). Potential
Therapeutic Effects of Citrus hystrix DC and Its
Bioactive Compounds on Metabolic Disorders.
Pharmaceuticals, 15(2), 167. https://doi.
org/10.3390/ph15020167
Soetjipto, H., & Martono, Y. (2017). Plant Essential Oils
Potency as Natural Antibiotic in Indonesian
Medicinal Herb of “Jamu.” IOP Conference Series:
Materials Science and Engineering, 172, 012022.
https://doi.org/10.1088/1757-899X/172/1/01202
2
Soltanian, S. (2020). Phytol Down-Regulates Expression
of Some Cancer Stem Cell Markers and Decreases
Side Population Proportion in Human Embryonic
Carcinoma NCCIT Cells. Nutrition and Cancer, 1–
14. https://doi.org/10.1080/01635581.2020.17956
95
Stasiulewicz, A., Znajdek, K., Grudzień, M., Pawiński,
T., & Sulkowska, J. I. (2020). A Guide to Targeting
the Endocannabinoid System in Drug Design.
International Journal of Molecular Sciences, 21(8),
2778. https://doi.org/10.3390/ijms21082778
Suresh, A., Velusamy, S., Ayyasamy, S., & Rathinasamy,
M. (2021). Techniques for Essential Oil Extraction
from Kaffir Lime and Its Application in Health
Care Products—A Review. Flavour and Fragrance
Journal, 36(1), 5–21.
https://doi.org/10.1002/ffj.3626
Takaichi, S., Sasikala, C., Ramana, C. V., Okamura, K., &
Hiraishi, A. (2012). Carotenoids in Rhodoplanes
species: Variation of Compositions and Substrate
Specificity of Predicted Carotenogenesis Enzymes.
Current Microbiology, 65(2), 150–155.
https://doi.org/10.1007/s00284-012-0139-y
Talon, M., Wu, G. A., Gmitter, F. G., & Rokhsar, D. S.
(2020). The Origin of Citrus. In The Genus Citrus.
Elsevier Inc. https://doi.org/10.1016/B978-0-12-
812163-4.00002-4
Tambunan, G. C. A., Dutt, A., Nadhifa, S., Amelia, F., &
Girsang, E. (2020). The In Vitro Anti-Diabetic
Activity of Lime Peels (Citrus amblycarpa (Hassk.)
Ochse). Journal of Health Sciences, 13(01), 26–33.
https://doi.org/10.33086/jhs.v13i01.1437
Toro, V. P. (2019). Larvicidal Effects of GC-MS Fractions
from Leaf Extracts of Cassia uniflora Mill non
Spreng. Journal of Biosciences, 44(1).
https://doi.org/10.1007/s12038-018-9823-9
Usman, M. A. (2021). Phytol Suppresses Parasitemia and
Ameliorates Anaemia and Oxidative Brain
Damage in Mice Infected with Plasmodium
berghei. Experimental Parasitology, 224.
https://doi.org/10.1016/j.exppara.2021.108097
Watanabe, T., Kawai, T., & Nonomura, Y. (2017). Effects
of Fatty Acid Addition to Oil-in-water Emulsions
Stabilized with Sucrose Fatty Acid Ester. Journal of
Oleo Science, 67(3), 307–313. https://doi.org/10.
5650/jos.ess17097
Wulandari, Y. W., Supriyadi, S., & Anwar, C. (2019).
Comparison between Hydrodistillation with
Steam Explosion and Conventional
Hydrodistillation in Kaffir Lime Oil Extraction.
AgriTECH, 39(4), 306. https://doi.org/10.22146/
agritech.38497
Xavier, A. A. O., & Pérez-Gálvez, A. (2016). Carotenoids
as a Source of Antioxidants in the Diet. Subcellular
Biochemistry, 79, 359–375.
https://doi.org/10.1007/978-3-319-39126-7_14
Xu, X., & Yan, J. (2021). β-Caryophyllene May Attenuate
Hyperoxaluria-Induced Kidney Dysfunction in
Rats by Regulating Stress Marker KIM-1/MCP-1
and NF-κB Signaling Pathway. Journal of
Biochemical and Molecular Toxicology, 35(11).
https://doi.org/10.1002/jbt.22891
Yu, W.-N., Lai, Y.-J., Ma, J.-W., Ho, C.-T., Hung, S.-W.,
Chen, Y.-H., Chen, C.-T., Kao, J.-Y., & Way, T.-D.
(2019). Citronellol Induces Necroptosis of Human
Lung Cancer Cells via TNF-α Pathway and
Reactive Oxygen Species Accumulation. In Vivo,
33(4), 1193–1201. https://doi.org/10.21873/
invivo.11590
Zou, S., & Kumar, U. (2018). Cannabinoid Receptors and
the Endocannabinoid System: Signaling and
Function in the Central Nervous System.
International Journal of Molecular Sciences, 19(3), 833.
https://doi.org/10.3390/ijms19030833