Evaluation of polyphenol and antioxidant properties of Blumea balsamifera extract as potential therapeutic for breast cancer

Abstract

Blumea balsamifera (Bb) is a plant used as herbal medicine in Southeast Asia, and it has been used due to its antibacterial, antiinflammatory, anticancer, etc. However, there is currently limited evidence that Bb leaf extract from Batu, Indonesia, contains beneficial compounds against breast cancer. Hence, this study evaluates the active compounds in extract and their potential as therapeutic agents for breast cancer. The total phenolic and flavonoid content was determined based on quantified colourimetry analysis followed by DPPH assay to evaluate antioxidant activity and phytochemicals screening in the extract, which was characterised by LC-HRMS analysis. Furthermore, computational methods are used to predict the pharmacological properties of compounds in the extract, particularly against breast cancer. The results showed a total phenolic content of 103.85+1.5 mgGAE/g and a total flavonoid content of 225.99+17.68 mgQE/g, with an antioxidant activity of 255.17+13.11µg/mL. 11 compounds were identified, but only four (Aurantio-obtusin, Isorhamnetin, Quercetin, and Hemiphloin) were computationally analysed. Molecular docking and dynamics simulation indicate that these phytochemicals bind to their target, possibly limiting their activity. Therefore, Bb has potential as a natural product remedy for breast cancer and contributes significantly to our knowledge of the plant by providing essential data for its future development.

Full text

Evaluation of polyphenol and antioxidant
properties of Blumea balsamifera extract as
potential therapeutic for breast cancer
Fairuz Sarah Kamila1, Yuslinda Annisa1, Nuraini Rosyadah1, Feri Eko Hermanto2,
Muhammad Hermawan Widyananda1, Dinia Rizqi Dwijayanti1, and Nashi Widodo1
1Biology Department, Faculty of Mathematics and Natural Sciences, Universitas Brawijaya, Malang,
Indonesia
2Faculty of Animal Sciences, Universitas Brawijaya, Malang, Indonesia
Abstract. Blumea balsamifera (Bb) is a plant used as herbal medicine in
Southeast Asia, and it has been used due to its antibacterial, anti-
inflammatory, anticancer, etc. However, there is currently limited evidence
that Bb leaf extract from Batu, Indonesia, contains beneficial compounds
against breast cancer. Hence, this study evaluates the active compounds in
extract and their potential as therapeutic agents for breast cancer. The total
phenolic and flavonoid content was determined based on quantified
colourimetry analysis followed by DPPH assay to evaluate antioxidant
activity and phytochemicals screening in the extract, which was
characterised by LC-HRMS analysis. Furthermore, computational methods
are used to predict the pharmacological properties of compounds in the
extract, particularly against breast cancer. The results showed a total
phenolic content of 103.85+1.5 mgGAE/g and a total flavonoid content of
225.99+17.68 mgQE/g, with an antioxidant activity of 255.17+13.11µg/mL.
11 compounds were identified, but only four (Aurantio-obtusin,
Isorhamnetin, Quercetin, and Hemiphloin) were computationally analysed.
Molecular docking and dynamics simulation indicate that these
phytochemicals bind to their target, possibly limiting their activity.
Therefore, Bb has potential as a natural product remedy for breast cancer
and contributes significantly to our knowledge of the plant by providing
essential data for its future development.
1 Introduction
Reactive Nitrogen Species (RNS) and Reactive Oxygen Species (ROS) are byproducts of
normal cell metabolism. Natural antioxidant pathways can mitigate the negative effects of
ROS. These free radicals play a variety of roles in human health and disease. As a result, it
is possible to slow disease progression and lower the risk of chronic diseases by boosting the
body’s natural antioxidant defence system or supplementing with dietary antioxidants.
Antioxidants play a crucial role in preventing disease by neutralising harmful free radicals.
While synthetic antioxidants have potential drawbacks, natural antioxidants derived from
 Corresponding author: widodo@ub.ac.id
© The Authors, published by EDP Sciences. This is an open access article distributed under the terms of the Creative Commons
Attribution License 4.0 (https://creativecommons.org/licenses/by/4.0/).
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plants offer a promising alternative. Researchers are actively seeking new natural
antioxidants with potent radical scavenging properties and minimal side effects [1].
Blumea balsamifera (Bb), also known as ‘Sembung legi’ in Indonesian, was a plant used
as herbal medicine in several Asian countries, including Indonesia, China, Malaysia,
Thailand, Philippines, and Vietnam. This plant is used in traditional medicine due to its
pharmacological properties, such as antibacterial, anti-inflammatory, anticancer etc. [2]. It
belongs to the Blumea genera and can grow in tropical and subtropical environments. It grew
as an herbaceous plant, reaching a height of 3-4 meters and emitting a pungent camphor
aroma. Its leaves were long with pointed tips, serrated leaf margins, fine hairs on the leaf
surface, and scattered single leaflets [3, 4].
Bb contains many phytochemical compounds, including phenolic and flavonoid groups
[5]. Previous studies have shown dihydroflavonol contained in Bb leaf extract work
synergistically with Tumour necrosis factor-related apoptosis-inducing ligand (TRAIL)
protein to induce apoptosis of adult T-cell leukaemia/lymphoma (ATLL) cancer cells [6].
Another study showed that several compounds from the flavonoid group are toxic to the oral
cavity (KB) and lung cancer cell lines (NCI-H187). The compounds include luteolin-7-
methyl ether, quercetin, dihydroquercetin-7,4’-dimethyl ether, 5,7,3’5’-tetrahydroxyflavone,
and blumeatin [7].
The usage of natural products in medicine has led to the search for new potential drugs
derived from this plant. The medicinal value of plants depends on their phytochemicals,
which have various physiological effects on the human body. As a result, phytochemical
screening can identify compounds present in plants that could be used as the basis for current
medication development [8]. Phytochemical evaluation involves identifying and quantifying
bioactive compounds contained in herbal extracts using a tool, for example, liquid
chromatography combined with mass spectrometry [9]. LC-MS involves the process of
separating compounds through a chromatographic column that is simultaneously
characterised by m/z values. Both methods provide comprehensive data information on the
compounds in the analysed sample [10].
In recent years, in silico methods have emerged as powerful tools for predicting the
biological activities of compounds. These methods involve using computational techniques
to analyse the molecular structure and properties of compounds and predict their potential
interactions with biological targets. Molecular docking analysis predicts compound-protein
interactions using computer simulations. This method is important in the design of candidate
new drugs by simulating compound binding and its effect on protein structure. The
combination with molecular dynamics provides a deeper understanding of the dynamic
compound-protein interactions, helping in designing more effective and safe candidate drugs
[11].
While limited evidence exists for the efficacy of Bb leaves from Batu, Indonesia, against
breast cancer, the increasing incidence of this disease necessitates exploring novel treatment
options. Therefore, this study aimed to identify the phytochemicals and antioxidant activity
in B. balsamifera extract and explore the potential therapeutic of phenolic compounds in B.
balsamifera to inhibit breast cancer progression.
2 Materials and methods
2.1 Extraction
B. balsamifera leaves dried powder purchased from UPT Balai Materia Medica, Batu, East
Java. The extraction process is based on a previous study [12]. Dried leaves powder (5 g)
was inserted into the MAE vessel. Ethanol 96% (50 mL) was added into the MAE vessel
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containing the extract in a ratio of 1:10. The instrument is run by warming up to 50 °C for 5
minutes, holding set to 50 °C for 10 minutes, and cooling down for 5 minutes. The obtained
extract was filtered was put into a rotary vacuum evaporator at 50 rpm, 40 °C. The crude
extract was then stored at 4 °C before further use.
2.2 Total phenolic content
The total phenolic test is based on previous research protocols [12, 13]. The extract was 100
µg/mL, while gallic acid concentrations (standard) were 0.625, 1.25, 2.5, 5, 10, 20, 40 and
80 µg/mL. Extract and standard (10 µL each) were added to a 96-well plate, followed by
Folin-Ciocalteu (100 µL). Na2CO3 (100 µL) was added to the sample and incubated for 90
minutes at room temperature in a dark room. The absorbance of the sample was measured at
735 nm with a microplate reader. The total phenolic content of extracts was calculated by
converting the gallic acid standard curve (mg GAE/g).
2.3 Total flavonoid content
The total flavonoid test follows previous research protocols [14, 15]. The extract was used at
100 µg/mL, while the quercetin was used as a standard at 0.6125, 1.25, 5, 10, 20, 40, and 80
µg/mL. Extracts and quercetin (50 µL) were added to a 96-well plate, followed by ethanol
96% (150 µL) and AlCl3 (10 µL). CH3COONa 1 M (10 µL) was added to the sample and
incubated for 40 minutes at room temperature in a dark room. The absorbance of the sample
was measured at 405 nm using a microplate reader. The total flavonoid content of extracts
was calculated by converting the quercetin standard curve (mg QE/g).
2.4 Antioxidant activity
The antioxidant activity protocol was based on the previous study [12]. The extract
concentrations were 25, 50, 100, 200, and 400 µg/mL, while ascorbic acid as a standard at
0.625, 1.25, 2.5, 5, 10, 20, 40, and 80 µg/mL. Extracts and gallic acid (100 µL) were added
to a 96-well plate, followed by DPPH 0.4 mM solution (100 µL). The sample was incubated
for 30 minutes at room temperature in a dark room. The absorbance of the sample was
measured at 490 nm using a microplate reader. The antioxidant activity of extracts was
determined using the percentage IC50 value ratio of extract to ascorbic acid (%).
2.5 LC-HRMS analysis
The preparation protocol and compound profile analysis of the extract follows the
Metabolomics Laboratory protocol at Bogor Agricultural University (IPB, Bogor). Extract
(5 mg) was dissolved in methanol 1 mL, then filtered through a nylon membrane (0.2 µm).
LC-MS performed by Vanquish Tandem Q Exactive Plus Orbitrap HRMS UHPLC device
(Thermo Scientific, USA). The column used was an Accucore C18, with 100 x 2.1 mm, 1.5
µm (Thermo Scientific, USA). The Eluents were H2O + 0.1% formic acid (A) and acetonitrile
+ 0.1% formic acid (B). The eluent gradient ranged from 0-1 minute (A 5%), 1 to 25 minutes
(A 5-95%), 25 to 28 minutes (B 95%), and 28 to 33 minutes (B 5%). UHPLC was
programmed with a flow rate of 0.2 mL/min and a column temperature of 30 °C. Sample 2
µL was injected into a column on a UHPLC. HRMS was configured with a mass range of
100-1500 m/z and a negative ionisation mode. HRMS data was analysed using Compound
Discoverer 3.2 and mzCloud.
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2.6 Druglikeness, bioactivity, and target protein prediction
Eleven bioactive compounds were retrieved from LC-HRMS analysis. Subsequently, their
biological activities were analysed using the PASS server(http://way2drug.com/passonline/).
The assessment of biological activity was based on Pa values, with a cut-off set at Pa > 0.3.
The results of the biological activity analysis were displayed as a Heatmap using GraphPad
Prism. The pharmacological properties of the bioactive compounds were assessed using the
SwissADME web (http://www.swissadme.ch). The analysis used each compound’s
canonical smiles from the PubChem database and submitted to SwissADME. SWISS Target
Prediction (http://www.swisstargetprediction.ch/) was used to identify protein targets
associated with breast cancer in each compound. Target proteins were chosen based on their
association with breast cancer and frequent alteration.
2.7 Molecular docking and structural visualisation
Ligands were retrieved from PubChem and saved in three-dimensional SDF format. Ligand
energy was minimised by Open Babel in PyRx software. Native ligands as control were
extracted from each of the proteins using PyMol software. The three-dimensional protein
structure was sourced from the PDB RCSB database (https://www.rcsb.org.). Water
molecules and native ligands were removed from proteins using Biovia Discovery Studio
2019 Client software. The targeted docking process was performed between these
compounds and the active site of each protein using AutoDock Vina, which was integrated
into PyRx. The docking data were then visualised using the Biovia Discovery Studio 2019
Client software.
2.8 Molecular dynamics
Molecular dynamics simulation was analysed using Yet Another Scientific Artificial Reality
Application (YASARA), a software commonly used for such simulations, with certain
modifications as described in [12]. The system parameters were tuned to match the cellular
physiological conditions, with settings at 310 °K, pH 7.4, 1 atm pressure, and a NaCl
concentration of 0.9%, maintained throughout a 50 ns period. The macros program included
"md_run" for executing simulations, "md_analyze" for assessing RMSD, and
"md_bindingenergy" for evaluating the binding energy of protein-ligand complexes during
molecular dynamics.
3 Results and discussions
3.1 Total phenolic content, total flavonoid content and antioxidant activity
Table 1 shows that B. balsamifera extract (BBE) has phenolic content in correlation with
flavonoid content. The concentrations of phenolic compounds and flavonoids in extracts vary
depending on the solvent used for extraction. This occurs because plant-derived compounds
are classified as polar or non-polar [16]. As a result, solvent selection is critical to the
efficiency of the polyphenol extraction process from plants. Furthermore, the precise
concentrations of phenols and flavonoids in the BBE may differ from those reported in
previous studies [17, 18]. Other factors, such as differences in growing locations, influence
component levels, and different growing environments can affect the plant’s metabolic
system, resulting in varying quantities of compounds produced [17].
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However, the phenolic and flavonoid content has a low correlation to the antioxidant
properties of BBE. The molecular structure of flavonoids has a significant influence on the
antioxidant activity of plant extracts. Key structural features contributing to this activity
include the ortho-dihydroxy structure on the B-ring, the 2,3 double bond with 4-oxo
functional group on the C-ring, and the 3- and 5- hydroxyl groups with 4-oxo functionalities
on the A and C rings. The ortho-dihydroxy structure on the B-ring enables efficient electron
donation and radical stabilisation, while the 2,3 double bond with the 4-oxo functional group
on the C-ring facilitates electron delocalisation. The 3- and 5-hydroxyl groups with 4-oxo
functionalities on the A and C rings are required for maximising radical scavenging potential
[19]. In addition, co-existing pigments in extracts that absorb in the same wavelength range
as DPPH radicals (around 517 nm) may interfere with the absorbance readings [20].
Consequently, high flavonoid content alone does not guarantee strong antioxidant activity,
as the specific molecular structure of flavonoids and the presence of interfering pigments are
significant factors.
Table 1. Total phenolic, flavonoid and antioxidant activity of BEE.
Assay
Value
TPC (mgGAE/g)
103.85 + 1.5
TFC (mgQE/g)
225.99 + 17.68
DPPH (µg/mL)
255.17 + 13.11
3.2 LC-HRMS analysis
The bioactive compounds detected in the B. balsamifera extract by LC-HRMS are listed in
Table 2. The compound 4,5-dicaffeoylquinic acid is known to have several bioactivities,
namely antioxidant, anti-inflammatory, prostate anticancer, anti-diabetic and cognitive
function improvement [21–23]. Aurantio-obtusin is known to be effective in inhibiting the
growth of liver cancer cells by decreasing fat production and inducing ferroptosis, which is
cell death due to the accumulation of Fe in the cells [24]. Lecanoric acid is effective in
inhibiting the growth of colon cancer cells by stopping the cell cycle in the M phase so that
the cancer cell division process is inhibited. In addition, the induction of death by this
compound is only specific to cancer cells [25]. Corchorifatty acid F showed efficacy as an
antifungal against the pathogenic fungus Pyricularia oryzae in resistant rice varieties [26].
Isorhamnetin has anti-tumour activity against several cancer cells, including cervical,
lung, colon, breast, pancreatic, nasopharyngeal, liver, and gastric cancer cells. The anti-
tumour activity of this compound by inhibiting cancer cell proliferation induces apoptosis
and regulates tumour suppressor genes, proto-oncogene, and signalling pathways [27].
Tuberonic acid glucoside is a regulator of tuberonic acid levels in response to environmental
conditions [28]. Quercetin can induce the inhibition of important molecular pathways in cells,
such as MAPK/ERK1/2, p53, JAK/STAT, and others. These signalling pathways regulate
cell growth, induction of cell death and cell cycle arrest [29].Quercetin-3β-D-glucoside has
anti-arteriosclerotic, anti-inflammatory, antiviral, and antioxidant properties both in vivo and
in vitro. It also improves the anti-inflammatory properties of M2a macrophages and controls
the immune response to pro-inflammatory stimuli [30].
Salicylic acid can inhibit cancer cell growth by inducing stress on the endoplasmic
reticulum that induces death signals [31]. Chlorogenic acid, based on the previous study, has
the potential to be an anticancer by halting the cell cycle, inducing apoptosis, and suppressing
cancer cell proliferation [32]. Hemiphloin has pharmacological effects for humans, including
antioxidant, anti-inflammatory, anticancer, cardioprotective, and neuroprotective [33].
Overall, the results suggest that Bb extract contains bioactive compounds with potential
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health benefits. In addition, phenolic compounds found in the extract have promising
activities to inhibit cancer cells.
3.3 Druglikeness and bioactivity prediction
The general method to assess compound drug-likeness involves property-based filters and
substructure analysis, which set specific boundaries for certain molecular properties of
compound or drug candidates. According to the drug-likeness screening, all of the
compounds have passed the Lipinski rules [34] that are indicated in Table 3, indicating good
bioavailability and cell membrane penetration. While most compounds exhibited favourable
GIA, all of the compounds were unable to cross the BBB. Solubility was determined using
ESOL [34, 35], with most compounds classified as soluble, facilitating drug development
and formulation. ESOL (estimated solubility) is a computational chemistry model used to
predict a compound's solubility in water. The model belongs to the Quantitative Structure-
Property Relationship (QSPR) category, which is a model that links the chemical structure
of a compound with its solubility properties. The solubility scale is divided into several
categories, including Log S value < -10 (insoluble), -10 < Log S value < -6 (less soluble), -4
< Log S value < -2 (moderate soluble), -2 < Log S value < 0 (soluble), Log S value > 0 (very
soluble) [35]. P-gp interaction was evaluated, revealing that all of the compounds were not
substrates of P-gp. P-gp (permeability glycoprotein) influences the ADMET properties of
xenobiotics (drugs or chemicals) as it regulates the uptake and metabolism of compounds
within the cell, which acts similar to an efflux pump to remove the substrate from the cell
[34]. These findings suggest that B. balsamifera compounds have promising drug-like
properties.
The bioactivity screening results for four compounds through PASS Online, identified
four compounds activities were associated with the cancer pathway (Fig. 1). The compounds
were further screened to assess their potential bioactivity, with the aim of predicting their
capability to inhibit the progression of breast cancer. The bioactivities related to the
progression of breast cancer predicted from compounds were MMP9 inhibitor, JAK2
inhibitor, apoptosis agonist, caspase 3 stimulant, caspase 8 stimulant, TP53 enhancer,
antineoplastic (breast cancer), Bcl2 antagonist, Bcl-xL inhibitor, breast cancer-resistant
protein inhibitor, CDC25A inhibitor, CDC25B inhibitor, CDC25C inhibitor, CDC25
phosphatase inhibitor, CDK4/cyclin D1 inhibitor, CDK6 inhibitor, Topoisomerase I
inhibitor, Topoisomerase II inhibitor, Estrogen antagonist, Protein kinase C inhibitor,
antimutagenic, antineoplastic (breast cancer), anticarcinogenic, antimetastatic, and
chemosensitiser. Besides, bioactivities related to antioxidant activities were HIF-1α inhibitor,
free radical scavenger, and antioxidant.
3.4 Molecular docking
Molecular docking was used to analyse the interactions between compounds in B.
balsamifera with target proteins selected based on SwissTarget (EGFR, ERα, IGF1R). EGFR
is a highly activated transmembrane protein receptor that can trigger a series of reactions in
cells that can promote uncontrolled cell growth and the formation of cancer cells [36]. ERα
plays a role in cell regulation related to proliferation and survival and induces cancer cell
growth and metastasis [37]. IGF1R plays a role in cell regulation related to survival, growth,
cell cycle, and differentiation of cancer cells [38]. Table 4 shows the binding affinity values
for all complexes. The complexes with the lowest binding affinity values for each compound
included EGFR-quercetin, ERα-hemiphloin, and IGF1R-isorhamnetin.
Fig. 2 depicts detailed interactions, with all three potential compounds binding to the
target protein’s active site. Quercetin binds to EGFR at the same residue as the inhibitor,
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including Leu718, Val726, Ala743, Ile744, Glu762, Met766, Leu788, Ile789, Thr790,
Leu792, Met793, Gly796, Leu844, and Thr854. Hemiphloin binds to ERα at the same residue
as the inhibitor, including Met343, Leu346, Thr347, Ala350, Glu353, Trp383, Leu384,
Leu387, Met388, Leu391, Leu525, and Met528. IGF1R binds to IGF1R at the same residue
as the inhibitor, including Leu975, Gln977, Gly978, Val983, Ala1001, Lys1003, Val1033,
Met1049, Glu1050, Leu1051, Met1052, Gly1055, Asp1056, Met1112, Met1126, Thr1127,
and Ile1130. The active compound has the same interaction position as an inhibitor. It is
considered to have similar activity to interfere with the protein’s function.
3.5 Molecular dynamics
Molecular dynamic simulation was used to determine the stability of the protein-ligand
complexes between three potential compounds and their targets [39]. The simulations
evaluated root mean square deviation (RMSD) of protein-ligand complexes, ligand
movement RMSD and molecular dynamics binding energy. The RMSD values of the three
selected complexes during simulations displayed low fluctuations, indicating that the ligand-
protein complexes' structure and the ligand's movement in the binding pocket of the target
protein during the simulation remained stable (Fig. 3). The binding energy values during the
molecular dynamic simulation displayed stable interaction between the ligand-protein with
the more positive value means more stable the ligand-protein interaction [39]. The binding
energy values show no fluctuations, suggesting the complex interaction was stable. Overall,
the molecular dynamics reveal that the interaction of active compounds to target protein
remains stable.
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Table 2. Identified compounds in B. balsamifera based LC-HRMS analysis.
Name
Formula
m/z
Exper.
mass
Cal. mass
Mass error
Fragment
product
Ion
Mode
Class
Ref.
4,5-
Dicaffeoylquinic
acid
C25H24O12
516
516.1261
516.1257
0.775
354. 09143;
191.05513;
179.03386
[M−H]−
phenolic
[40]
Aurantio-obtusin
C17H14O7
330.0675
330.0739
330.0740
-0.3029
314.04266;
299.01913;
271.02429;
243. 02872
[M−H]−
Anthraquinone
[41,42]
Lecanoricacid
C16H14O7
318.0662
318.0737
318.0740
-0.9431
167.03383;
149.02328;
123.04377
[M−H]−
phenolic
[43–45]
Corchorifatty acid
F
C18H32O5
328.2237
328.2251
328.2250
0.3046
229.14493;
211.13298;
183.13773;
97.05895;
85.02833;
57.03311
[M−H]−
fatty acid
[46]
Isorhamnetin
C16H12O7
316.0583
316.0583
316.0583
0.00
330.02695;
271.02402
[M−H]−
flavonoid
[47,48]
Tuberonic acid
glucoside
C18H28O9
388.1733
388.1733
388.1733
0.00
207.10168;
163.11220
[M−H]−
terpenoid
[49]
Quercetin
C15H10O7
302.18
302.0427
302.0427
0.00
273.04022;
229.04938;
151.00256;
121.02830;
107.00581
[M−H]−
flavonoid
[50]
Quercetin-3β-D-
glucoside
C21H20O12
464.0936
464.0955
464.0955
0.00
301.03433;
300.02676;
271.02368
[M−H]−
flavonoid
[46,47]
Salicylic acid
C7H6O3
138.0311
138.0311
138.0317
-4.3468
108.89841;
93.03335
[M−H]−
phenolic
[46]
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Name
Formula
m/z
Exper.
mass
Cal. mass
Mass error
Fragment
product
Ion
Mode
Class
Ref.
Chlorogenic acid
C16H18O9
354.0878
354.0952
354.0951
0.2824
191.05511;
179.03352;
161.02330;
135.04399
[M−H]−
phenolic
[47]
Hemiphloin
C21H22O10
434.1154
434.1213
434.1213
0.00
343.08188;
313.07135;
271.06116;
151.00255;
119.04906
[M−H]−
flavonoid
[47]
Table 3. Druglikeness prediction of BEE selected compounds.
Compounds
Physicochemical Properties (Lipinski rule)
Water Solubility
Pharmacokinetics
TPSA
MLOGP
HBA
HBD
MW
Log S
(ESOL)
Class
GI
BBB
P-gp
Aurantio-
obtusin
113.29
Ų
-0.22
7
3
330.07387
-3.8
Soluble
High
No
No
Isorhamnetin
120.36
Ų
-0.31
7
4
316.0583
-3.36
Soluble
High
No
No
Quercetin
131.36
Ų
-0.56
7
5
302.04265
-3.16
Soluble
High
No
No
Hemiphloin
177.14
Ų
-1.83
10
7
434.12133
-2.58
Soluble
Low
No
No
TPSA: Topological polar surface area; MW: molecular weight; GI: gastrointestinal tract; BBB: blood brain barrier; P-gp: P-glycoprotein
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Fig 1. Bioactivity related to breast cancer prediction of selected compounds. BC = breast cancer.
Table 4. Molecular docking results between bioactive compounds and their target proteins
Compound
Protein target
PDB ID
Inhibitor
Ref.
Binding affinity (kcal/mol)
Protein-compound
Protein-inhibitor
Aurantio-obtusin
EGFR
4I23
Dacomitinib
[51]
-7.88
-8.158
ERα
3ERT
Tamoxifen
[52]
-5.194
-9.428
Isorhamnetin
EGFR
4I23
Dacomitinib
[51]
-8.367
-8.158
IGF1R
2OJ9
Benzimidazole
[53]
-7.978
-9.478
Quercetin
EGFR
4I23
Dacomitinib
[51]
-8.861
-8.158
IGF1R
2OJ9
Benzimidazole
[53]
-7.475
-9.478
Hemiphloin
ERα
3ERT
Tamoxifen
[52]
-6.776
-9.428
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Fig 2. Molecular interaction of the ligand with target. A) EGFR, B) ERα, C) IGF1R.
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Fig 3. Molecular dynamics of the ligand with target. A) EGFR, B) ERα, C) IGF1R
From above to below – RMSD backbone, RMSD ligand movement, and Binding energy
12
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4 Conclusion
The findings of this study demonstrate that B. balsamifera has a diverse range of identified
compounds with health-promoting properties. Furthermore, the phenolics in BBE show
promising anticancer activity. The molecular docking and molecular dynamics simulations
provide further evidence for the potential therapeutic applications of BBE for breast cancer.
Further research is needed to explore the potential health benefits of BBE fully and
investigate its mechanisms of action. In vitro and in vivo studies are also necessary to
evaluate the efficacy and safety of BBE extract before it is used in further therapeutic
applications.
This research was funded by Professor Grants from LPPM (Grants No.
02172.4/UN10.F0901/B/KS2024), Universitas Brawijaya, Malang. We also thank the AI Center of
Universitas Brawijaya for providing access to high-performance computing facilities to perform
molecular dynamics simulations.
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