Short Communication

Natural Product Communications

Volume 19(1): 1–10
The Leaf Oils of Beilschmiedia tonkinensis © The Author(s) 2024
Article reuse guidelines:
(Lecomte) Ridl. and Lindera gracilipes H. W. sagepub.com/journals-permissions
DOI: 10.1177/1934578X231224995
Li: Chemical Composition, Cytotoxicity, journals.sagepub.com/home/npx

Antimicrobial Activity, and Docking Study

Duong Quang Huan1 , Nguyen Dinh Luyen2, Nguyen Xuan Ha2 ,

Do Ngoc Dai3 , Nguyen Quang Hop1, Do Thi Lan Huong4
and Ninh The Son5

Abstract
Objective: The Lauraceae plants comprised high amounts of essential oils, some of which established pharmacological potentials such
as anticancer and antimicrobial activities. The Lauraceae essential oils are also used in cuisines and perfumes. The present study pro-
vides the chemical analysis of the leaf oils of Beilschmiedia tonkinensis and Lindera gracilipes, collected from Vietnam. Method: Chemical
components in the obtained oils were identified by the GC-FID/MS. The MTT and broth microdilution assays were used to evaluate
cytotoxic and antimicrobial effects, respectively. The protein interactions were viewed by a docking study. Result: Beilschmiedia ton-
kinensis leaf oil was characterized by sesquiterpene hydrocarbons (66.0%), in which bicyclogermacrene (23.3%), (E)-caryophyllene
(21.9%), caryophyllene oxide (9.9%), and spathulenol (6.0%) were the main components. The major chemical class in L gracilipes
leaf oil was still sesquiterpene hydrocarbons (64.2%) with bicyclogermacrene (32.2%) being the principal component. Both 2 oil
samples (IC50 41.2-44.12 µg/mL) were actively cytotoxic against the proliferation of A-549 cancer cells. In particular, B tonkinensis
leaf oil strongly controlled Hep-G2 and MCF-7 cancerous cells with the IC50 values of 20.6 and 9.36 µg/mL, respectively.
Beilschmiedia tonkinensis leaf oil also strongly inhibited the Gram (–) bacterium Pseudomonas aeruginosa and the fungus Aspergillus niger
with the MIC values of 16 and 32 µg/mL, respectively. By molecular docking approach, bicyclogermacrene interacted with the
p38α MAPK cancer protein (PDB ID: 4FA2) with a potential binding affinity of −8.019 kcal/mol, whereas (E)-caryophyllene
tends to bind 2 bacterial proteins P aeruginosa QS regulator (PDB ID: 6B8A) and glucosamine-6-phosphate synthase (GlmS)
(PDB ID: 2VF5) with the better binding affinities of −6.740 and −6.521 kcal/mol, respectively. The most preferable binding
mode was due to hydrophobic π-alkyl and alkyl interactions. Conclusion: The current result can be seen as a basic foundation
in the applications of essential oils of B tonkinensis and L gracilipes for anticancer and antimicrobial treatments. Further phytochemical
studies and mechanisms of action are needed.

Keywords
Beilschmiedia tonkinensis, Lindera gracilipes, essential oil, cytotoxic, antimicrobial, docking study

Received: August 5th, 2023; Accepted: December 19th, 2023.

Introduction 1
Faculty of Chemistry, Hanoi Pedagogical University 2 (HPU2), Vinhphuc, Viet
In the laurel family, Beilschmiedia is a genus containing about 278 Nam
2
plants of shrubs and medium trees, which are widely distributed Institute of Natural Products Chemistry, Vietnam Academy of Science and
Technology (VAST), Hanoi, Vietnam
in tropical regions.1 Various Beilschmiedia species have been tra- 3
Faculty of Agriculture, Forestry and Fishery, Nghe An University of
ditionally used to cure a variety of diseases, such as uterine Economics, Nghean, Vietnam
cancers, rheumatism, respiratory problems, infectious bacteria, 4
Faculty of Biology and Agriculture, HPU2, Vinhphuc, Viet Nam
malaria, and tuberculosis.2 It turns out that Beilschmiedia plants 5
Institute of Chemistry, VAST, Hanoi, Vietnam
are rich in essential oils. β-Caryophyllene (12.1%), β-eudesmol
Corresponding Author:
(24.1%), and δ-cadinene (28.7%) were the predominant compo- Ninh The Son, Institute of Chemistry, VAST, 18 Hoang Quoc Viet, Caugiay,
nents in the aerial part oils of Barclaya kunstleri, Byttneria maingayi, Hanoi 10000, Vietnam.
and Beilschmiedia penangiana, respectively.1 Beilschmiedia pulverulenta Email: yamantson@gmail.com

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(https://creativecommons.org/licenses/by-nc/4.0/) which permits non-commercial use, reproduction and distribution of the work without further permission
provided the original work is attributed as specified on the SAGE and Open Access page (https://us.sagepub.com/en-us/nam/open-access-at-sage).
2 Natural Product Communications
aerial part oil with eugenol (45.3%) being the main component
has good effects in antioxidative, antimicrobial, anti-
inflammatory, antityrosinase, and anticholinesterase examina-
tions.3 The cytotoxic activity of Beilschmiedia erythrophloia leaf
oil was due to the role of β-caryophyllene (22.6%) and
α-humulene (21.9%).4 Beilschmiedia glabra leaf and bark oils con-
taining β-eudesmol (15.4%-19.3%) exerted good antioxidative
effects.5
Lindera is a big genus of about 100 species of shrubs or
deciduous trees that are mostly native to tropical and subtrop-
ical regions.6 The medical and therapeutic benefits of Lindera
plants are well-known, in addition to their decorative and com-
mercial values. For instance, L aggregata parts have significant
effects on rheumatoid arthritis, chronic gastritis, and enuresis,
as well as other illnesses of the urinary system.6 Similar to
other genera of the family Lauraceae, Lindera plants are also
good reservoirs of essential oils. Lindera glauca fruit oil consisted
of more than 40 components, in which (E)-β-ocimene (41.5%)
and α-copaene (13.2%) were the major components.7 L. setchue-
nensis leaf oil was predominated by palmitic acid (26.6%).8
Lindera essential oils have also been paid attention to due to
their cytotoxic and antibacterial properties.9–11 The leaf oil of L
strychnifolia possessed IC50 values of 22.7 to 24.1 µg/mL toward
3 cancerous cell lines A-549, HeLa, and Hep-G2, whereas its
root oil generated IC50 values of 23.9 to 38.0 µg/mL.8
Lindera umbellate essential oil, also known as Kuromoji, caused
apoptosis in HL-60 cancer cells.10 Lindera nacusua leaf oil suc-
cessfully inhibited the bacterium Staphylococcus aureus and the
fungus Candida albicans with the inhibition zones of 23.7 and
23 mm, respectively.11
For the first time, our current study reports the chemical
analysis of essential oils of the leaves of 2 Vietnamese
Lauraceae species, Beilschmiedia tonkinensis, and Lindera gracilipes,
as well as the uses of these oils in cytotoxicity and antimicrobial
treatment.
Results and Discussion
Hydrodistillation of B tonkinensis fresh leaves gave a yellow oil
containing 45 identified components (Table 1). In a total of
96.2%, sesquiterpene hydrocarbons accounted for 66.0%, fol-
lowed by oxygenated sesquiterpenes (21.5%), monoterpene
hydrocarbons (8.2%), and nonterpenic components (0.5%). It
was also found that B tonkinensis leaf oil was predominated by
bicyclogermacrene (23.3%), (E)-caryophyllene (21.9%), caryo-
phyllene oxide (9.9%), and spathulenol (6.0%). There have
been a great number of components with more than 1.0%,
such as α-humulene (3.5%), (Z )-β-ocimene (2.9%), α-pinene
(2.2%), (E)-β-ocimene (2.0%), α-copaene (1.5%), and
δ-elemene (1.4%).
From Table 1, it turns out that L gracilipes leaf oil contained 55
identified components, which represented 91.6%. As can be
seen, the major chemical class was still sesquiterpene hydrocar-
bons (64.2%), whereas the remaining chemical classes consisted
of nonterpenic components (13.7%), oxygenated sesquiterpenes
(7.0%), monoterpene hydrocarbons (4.7%), oxygenated mono-
terpenes (1.5%), and diterpene derivatives (0.6%). As compared
to B tonkinensis leaf oil, bicyclogermacrene in this sample was still
the main component with 32.2%. However, the amount of
(E)-caryophyllene was found to be only 1.0%. Lindera gracilipes
leaf oil has been also characterized by the appearance of other
identified components, such as (E)-4,8-dimethylnona-13,7-triene
(7.6%), (E,E)-α-farnesene (3.6%), dodecanal (2.8%), E-nerolidol
(2.3%), and γ-muurolene (2.0%).
Vietnamese Beilschmiedia and Lindera genera have drawn atten-
tion due to their essential oil resources. The leaf essential oils of
Vietnamese Beilschmiedia plants B erythrophloia, B fordii, B robusta,
and B yunnanensis were accompanied by the appearance of the
principal components bicyclogermacrene (30.5%), α-pinene
(45.1%), (E)-caryophyllene (22.5%), and 9-epi-(E)-caryophyllene
(21.2%), respectively.14,15 β-Caryophyllene (29.2%), α-humulene
(18.0%), and caryophyllene oxide (14.6%) can be seen as the
major components of L glauca leaf oil.15 The most dominant com-
ponent in the other Vietnamese Lindera species was camphor
(67.5%).16 Our current results provide new evidence about chem-
ical components analysis for 2 other perennial Lauraceae growing
in Vietnam.
Both 2 oil samples were submitted to cytotoxic and antimi-
crobial assays. From Table 2, the leaf oils of B tonkinensis and L
gracilipes exhibited cytotoxic activity against A-549 cancerous
cells with IC50 values of 44.12 and 41.2 µg/mL. Significantly,
B tonkinensis leaf oil successfully controlled the proliferation of
Hep-G2 and MCF-7 cancerous cells with the corresponding
IC50 values of 20.6 and 9.36 µg/mL. In contrast, L gracilipes
leaf oil did not exhibit activity (IC50 > 100 µg/mL). The
current results match well with previous reports. For instance,
Su and Ho revealed that B erythrophloia leaf oil was responsible
for the IC50 value of 38.8 µg/mL.3 As mentioned above, cyto-
toxicity of L strychnifolia leaf oil against A-549 and Hep-G2 cells
exerted the IC50 values of 23.1 and 22.7 µg/mL, respectively.7
Regarding antimicrobial assay, B tonkinensis leaf oil inhibited
the proliferation of 2 Gram (+) bacterial strains S aureus and
Bacillus subtilis with the same MIC value of 64 μg/mL, but the
second sample did not show activity (MIC > 400 μg/mL)
(Table 3). Both 2 oil samples were active against 2 Gram (–)
bacteria Escherichia coli and Pseudomonas aeruginosa. Especially, B
tonkinensis leaf oil showed strong activity against P aeruginosa
with the MIC value of 16 μg/mL, in comparison with that of
the standard tetracycline (MIC 8 μg/mL). Likewise, B tonkinensis
leaf oil has good results (MIC 32-64 μg/mL) against 2 fungi
Aspergillus niger and Fusarium oxysporum, in comparison with
those of L gracilipes leaf oil (MIC 64-128 μg/mL). In the final
case, B tonkinensis leaf oil possessed MIC values of 32 to 64
μg/mL against 2 yeasts C albicans and Saccharomyces cerevisiae,
whereas L gracilipes leaf oil only exhibited activity against S cere-
visiae with a MIC value of 128 μg/mL. In literature, the leaf and
bark oils of B madang were moderately active against B subtilis
and S aureus with a MIC value of 125 µg/mL.17 Beilschmiedia
fordii leaf oil generated remarkably strong antimicrobial activity
against S aureus, B cereus, and C albicans with a MIC value of 16
Huan et al 3

Table 1. Chemical Constituents in the Leaf Oils of 2 Vietnamese Lauraceae Plants.a

Rt RIE RIL Constituents Beilschmiedia tonkinensis Lindera gracilipes
9.87 940 932 α-Pinene 2.2 1.2
10.36 956 946 Camphene 0.2
11.06 979 969 Sabinene 0.2
11.23 985 974 β-Pinene 0.4 0.7
11.48 993 988 Myrcene 0.2 0.2
12.03 1010 1002 α-Phellandrene 1.7
12.69 1029 1022 o-Cymene 0.3
12.88 1035 1024 Limonene 0.2 0.3
13.03 1039 1032 (Z)-β-Ocimene 2.9 0.3
13.40 1050 1044 (E)-β-Ocimene 2.0
14.89 1094 1086 Terpinolene 0.2
15.79 1119 1113 (E)-4,8-Dimethylnona-13,7-triene 7.6
21.85 1294 1282 (E)-Anethole 0.2
23.67 1348 1335 δ-Elemene 1.4 1.3
24.08 1361 1345 α-Cubebene 0.3
24.88 1385 1373 α-Ylangene 0.2
25.04 1390 1376 α-Copaene 1.5 0.5
25.32 1398 1385 α-Funebrene 0.6
25.46 1402 1387 β-Cubebene 1.7
25.50 1404 1389 cis-β-Elemene 1.1 1.9
25.78 1413 1397 Dodecanal 2.8
26.16 1425 1398 Cyperene 0.7
26.23 1427 1409 α-Gurjunene 0.3
26.37 1431 1410 α-Cedrene 0.1 0.4
26.59 1438 1417 (E)-Caryophyllene 21.9 1.0
26.78 1444 1431 β-Gurjunene 0.9
26.83 1446 1432 trans-α-Bergamotene 0.6 0.1
27.02 1452 1437 α-Guaiene 0.2
27.17 1456 1439 Aromadendrene 0.4 3.9
27.27 1460 1545 Selina-5,11-diene 0.7
27.48 1466 1448 cis-Muurola-3,5-diene 0.4
27.65 1472 1452 α-Humulene 3.5 0.7
27.86 1479 1464 9-epi-(E)-Caryophyllene 0.6 1.6
28.16 1488 1475 trans-Cadina-1(6),4-diene 0.2 0.5
28.24 1491 1478 γ-Muurolene 0.6 2.0
28.35 1494 1483 α-Amorphene 0.1 0.9
28.48 1498 1484 Germacrene D 2.9 3.3
28.68 1505 1491 β-Selinene 0.6 1.7
28.74 1507 1492 δ-Selinene 0.9
28.86 1511 1493 trans-Muurola-4(14),5-diene 0.2
28.91 1512 1495 γ-Amorphene 1.0
28.95 1514 1498 (E,E)-α-Farnesene 3.6
29.01 1516 1500 Bicyclogermacrene 23.3 32.2
29.18 1522 1516 δ-Amorphene 0.5
29.51 1533 1518 γ-Cadinene 1.4 0.8
29.64 1537 1522 δ-Cadinene 0.9 1.7
29.95 1547 1533 trans-Cadina-1,4-diene 0.3
30.11 1553 1537 α-Cadinene 0.4
30.29 1559 1544 α-Calacorene 0.3
30.33 1560 1545 Selina-3,7(11)-diene 0.4
30.47 1565 1548 Elemol 0.4
30.66 1571 1557 E-Nerolidol 0.1 2.3
30.84 1577 1559 Germacrene B 0.3
31.03 1583 1570 Dendrolasin 1.2
31.50 1599 1577 Spathulenol 6.0 0.4
31.66 1605 1581 Viridiflorol 0.9
31.70 1606 1582 Caryophyllene oxide 9.9
31.92 1614 1583 Cubeban-11-ol 1.1

(Continued)
4 Natural Product Communications

Table 1. Continued
Rt RIE RIL Constituents Beilschmiedia tonkinensis Lindera gracilipes
31.93 1614 1600 Guaiol 0.3
31.98 1616 1595 Tetradecanal 1.5
32.15 1622 1597 Rosifoliol 0.3
32.29 1627 1600 Cedrol 0.4 0.7
32.43 1632 1608 Humulene epoxide II 1.1
32.72 1633 1613 5-Guaiene-11-ol 0.2
32.87 1647 1627 1-epi-Cubenol 0.3
33.19 1659 1638 epi-α-Cadinol 0.9
33.60 1673 1649 β-Eudesmol 1.4
33.60 1673 1652 α-Cadinol 0.6
33.69 1676 1658 neo-Intermedeol 0.6
33.67 1675 1652 α-Eudesmol 1.6
36.59 1782 1759 Benzyl benzoate 0.5 1.5
37.55 1819 1818 Hexadecanal 0.3
41.41 1973 1960 m-Camphorene 0.1
44.80 2117 2122 Phytol 0.5
Total 96.2 91.6
Monoterpene hydrocarbons 8.2 4.7
Oxygenated monoterpenes 1.5
Sesquiterpene hydrocarbons 66.0 64.2
Oxygenated sesquiterpenes 21.5 7.0
Diterpene hydrocarbons 0.1
Oxygenated diterpenes 0.5
Non-terpenic compounds 0.5 13.7
Abbreviations: Rt, retention time; RIE, retention indices relative to n-alkanes (C7-C40) on HP-5 MS column; RIL, retention indices from Adams12 and the NIST
standard database.13
aBold: Compounds present in amounts greater than 5%.

Table 2. Cytotoxicity of 2 Tested Oil Samples.

Samples Concentration (µg/mL) A-549 Hep-G2 MCF-7
Beilschmiedia tonkinensis 256 80.12 ± 0.6 91.42 ± 0,5 94.16 ± 0.4
128 66.25 ± 0.1 67.80 ± 0,9 87.19 ± 0.2
64 51.35 ± 0.2 55.17 ± 0,7 80.98 ± 0.7
32 39.84 ± 0.2 50.24 ± 0.6 52.12 ± 0.1
IC50 (µg/mL) 44.12 20.6 9.36
Lindera gracilipes 256 85.87 ± 0.2 30.72 ± 0.7 30.89 ± 0.1
128 62.93 ± 0.5 28.43 ± 0,6 18.19 ± 0.1
64 51.75 ± 0.1 24.89 ± 0.1 10.02 ± 0.8
32 35.13 ± 0.4 21.13 ± 0.3 9.34 ± 0.5
IC50 41.2 > 100 > 100
Ellipticine IC50 0.40 0.41 0.43

µg/mL.15 In another example, B pulverulenta aerial part oil sup-
pressed B subtilis and S aureus with the MIC value of 62.5 µg/
mL, as well as monitoring the growth of C albicans and S cerevisiae
with a MIC of 125 µg/mL.2 Collectively, Beilschmiedia essential oils
are appropriate for further antimicrobial treatments (Figure 1).
Molecular Docking Study
It was supposed that 2 main components, namely bicycloger-
macrene and (E)-caryophyllene, established a great role in the
antimicrobial activity of B tonkinensis leaf oil. Hence, they were
subjected to experimental molecular docking. This procedure
aimed to determine their binding affinity and optimal docking
poses at the active sites of the target protein. The redocking
experiments were conducted using Auto Dock Vina v1.2.3 soft-
ware, resulting in the RMSD (root mean square deviation)
values of 0.261688, 0.286839, and 1.24192 Å when aligning
the ligands with their corresponding redocking poses
(Figure 2). These findings validated the reliability of the molec-
ular docking experiments. Additionally, the cocrystallized
ligands, within the protein’s crystal structure, human P38α
MAPK (PDB ID: 4FA2), MvfR (PDB ID: 6F8A), and
glucosamine-6-phosphate synthase (GlmS) (PDB ID: 2VF5)
have been considered as reference inhibitory components,
Huan et al 5

Table 3. Antimicrobial Activity of 2 Tested Oil Samples (MIC: μg/mL).

Microbial strains Beilschmiedia tonkinensis Lindera gracilipes Streptomycin Tetracycline Nystatin
Gram (+) Bacillus subtilis ATCC 11774 64 >400 4
Staphylococcus aureus ATCC 29213 64 >400 8
Gram (–) Escherichia coli ATCC 8739 64 32 4
Pseudomonas aeruginosa ATCC 9027 16 32 8
Fungi Aspergillus niger ATCC 16404 32 64 16
Fusarium oxysporum ATCC 48112 64 128 8
Yeasts Candida albicans ATCC 14053 32 >400 4
Saccharomyces cerevisiae ATCC 9763 64 128 8

Figure 1. The major compounds in the leaf oil of Beilschmiedia tonkinensis (a) and Lindera gracilipes (b).

Figure 2. Validation of the docking protocol through the re-docking of the co-crystallized ligands in (A) 4FA2, (B) 6B8A, (C) 2VF5 proteins, and
superimposing their docked poses at the active binding pocket.

exhibiting respective binding affinities of −10.67, −10.42, and
−8.43 kcal/mol, respectively.
The p38 MAPK (p38 mitogen-activated protein kinase) is a
crucial signal transduction pathway that activates a complex
network of interacting proteins mainly responsible for various
cellular performances.18–21 Various studies have indicated that
p38α MAPK plays a significant role in cancer development, par-
ticularly showing substantial expression in invasive breast
cancer.22 Moreover, recent research has revealed that p38 α
inhibitors can be used for the deactivation of ER-negative
breast cancer and p53-mutated tumors.23 Therefore, the p38α
MAPK target was chosen for this study. To have an insight
understanding of the binding mode of the studied components
at the binding site of the p38α MAPK protein (PDB ID:
4FA2), molecular docking experiments were performed using
the AutoDock Vina program. The binding affinity of bicycloger-
macrene and (E)-caryophyllene was found to be −8.019 and
−7.955 kcal/mol, respectively (Table 4). The molecular interac-
tions of the docked poses within the active site region are
depicted in Figure 3. The main interactions observed between
6 Natural Product Communications

Table 4. Docking Results of the Major Compounds of Beilschmiedia tonkinensis Leaf Oils.
PDB ID Compound Binding affinity (kcal/mol) Alkyl and π–alkyl interactions
4FA2 Bicyclogermacrene −8.019 Met109, Leu108, Ala157, Ala51, Lys53, Ile84, Leu167, Leu171, Val38
(E)-Caryophyllene −7.955 Lys53, Ala51, Ile84, Val38, Leu171, Leu167, Met109, Ala157
SB239063 −10.67 Ala51, Lys53, Leu171
M64
Glucosamine-6-phosphate
6B8A Bicyclogermacrene −6.394 Leu189, Trp234, Val211, Ile236, Val170, Tyr258
(E)-Caryophyllene −6.740 Ile236, Val211, Trp234, Val170, Leu189, Tyr258, Ile263
SB239063
M64 −10.42 Ile263, Val170, Val211, Leu183, Ile149, Ala168
Glucosamine-6-phosphate
2VF5 Bicyclogermacrene −5.964 Leu601, Val605, Leu484
(E)-Caryophyllene −6.521 Cys300, Leu484
SB239063
M64
Glucosamine-6-phosphate −8.43

Figure 3. Binding interactions of compounds bicyclogermacrene and (E)-caryophyllene in the cavity of 4FA2 protein.

the 2 studied components were predominantly alkyl-type interac- an alkyl interaction with Leu108 residue was only observed in the
tions, focusing primarily on amino acid residues Ala51, Met109, complex formed by 4FA2 protein and bicyclogermacrene, but
Ala157, Lys53, Ile84, Leu167, Leu171, and Val38. Furthermore, not in the complex formed by 4FA2 protein and
Huan et al
Figure 4. Binding interactions of compounds bicyclogermacrene and (E)-caryophyllene in the cavity of 6B8A protein.
(E)-caryophyllene. Notably, the residues Ala51, Lys53, Leu171,
Leu167, and Met109 are crucial positions within the active site
of the 4FA2 protein.24 These findings demonstrate that these
main bioactive components can effectively interact with and
inhibit the p38 α MAPK pathway.
Regarding antimicrobacterial activity, P aeruginosa QS regulator
(or a multiple virulence factor regulator (MvfR)) (PDB ID: 6B8A)
and GlmS (PDB ID: 2VF5) are the 2 protein targets selected in this
study. MvfR is an important target for the control and treatment of
P aeruginosa infection caused by the direct and indirect activation of
a variety of virulence genes, responsible for the formation and
development of biofilms.25 Besides, GlmS played a crucial role
in microbial cell membranes by transferring fructose-6-phosphate
to glucosamine-6-phosphate in the presence of glutamine.26
Therefore, in this study, docking experiments were applied to
evaluate the behavior of the studied components in the active
site of the 6B8A and 2VF5 proteins. From Table 4, the binding
affinity of bicyclogermacrene and (E)-caryophyllene was −6.394
and −6.74 kcal/mol for 6B8A protein, and −5.964 and −6.521
7
kcal/mol for 2VF5 protein, respectively. (E)-Caryophyllene has
been shown to inhibit proteins 6B8A and 2VF5 more effectively
than bicyclogermacrene. The diagram interaction analysis shows
that the main interactions between the 2 studied components
were the alkyl and π–alkyl interaction types (Figure 4). In the
active region of the protein 6B8A, both components formed
interactions with residues Leu189, Tyr258, Trp234, Val211,
Ile236, and Val170, and further (E)-caryophyllene also formed
with the residue Ile263. It should be noted that Tyr258 Val211,
Ile236, Val170, and Ile263 are important amino acids in the
active region of the protein 6BA8. Similarly, observations of
interactions between the protein 2VF5 and selected components
also indicated that the main type of interaction is the alkyl
type (Figure 5). Bicyclogermacrene formed these 3 interactions
at the amino acid residue sites of Leu601, Leu484, and Valu605
while (E)-caryophyllene formed interactions with Leu484 and
Cys300. In summary, docking results have demonstrated the
antimicrobial effect of the major components of B tonkinensis
essential oils.
8 Natural Product Communications

Figure 5. Binding interactions of compounds bicyclogermacrene and (E)-caryophyllene in the cavity of 2VF5 protein.

Conclusion bicyclogermacrene was the main contributor to MCF-7 cytotox-

This is the first time that chemical components of the leaf oils of
2 Vietnamese Lauraceae plants B tonkinensis and L gracilipes were
identified by the GC-FID/MS analytical procedure. For both 2
oils, sesquiterpene hydrocarbons (64.2%-66.0%) were the main
chemical class, as well as bicyclogermacrene (23.3%-32.2%),
can be seen as the main components. Both 2 oil samples exhib-
ited cytotoxicity toward A-549 cancer cell lines. In particular, B
tonkinensis leaf oil strongly controlled the proliferation of
Hep-G2 and MCF-7 with the IC50 of 9.36 to 20.6 µg/mL.
Two oil samples exhibited antimicrobial activity at different
levels, in which B tonkinensis leaf oil showed strong activity
against the Gram (–) bacterial strain P aeruginosa and fungal
strain A niger with MIC values of 16 to 32 µg/mL. Molecular
docking studies showed a good binding affinity, in which
icity activity, sharing many important residues with the p38α
MAPK active site, whereas (E)-caryophyllene remarkably
bound 2 bacterial proteins P aeruginosa QS regulator and
glucosamine-6-phosphate synthase (GlmS). As a result, they
may act as strong inhibitors when synergizing with other
minor components.
Experimental
Materials
Beilschmiedia tonkinensis fresh leaves were gathered from Pu
Huong-Nghean, Vietnam, whereas L gracilipes fresh leaves
were collected from Thanh Chuong-Nghean, Vietnam, in
March 2023. Botanical taxonomy was confirmed by the
Huan et al
coauthor Dr Do Ngoc Dai. Two voucher specimens, BTL-2023
(B tonkinensis) and LGL-2023 (L gracilipes), have been deposited
in the faculty of chemistry, HUP2 University.
Distillation
The fresh leaves of B tonkinensis (1.0 kg) were cut into small
pieces, and then submitted to water distillation using a
Clevenger-type apparatus for 3.2 h, to afford a yellowish oil
(0.6% w/w). The obtained oil was dried over Na2SO4 and
maintained in a sealed vial at −10 °C for further analysis. The
same manner was assigned to the fresh leaves of L gracilipes
(1.0 kg), to yield a yellowish oil with 0.5% w/w.
The GC-FID/MS Analyses
Chemical components in the studied essential oils were identi-
fied by the GC-FID/MS (gas chromatography-flame ionization
detection/mass spectrophotometry) analyses as previously
reported.27–29 The GC analysis was performed using an HP
7890A Plus Agilent Technologies gas aided by an FID and
coupled with an HP-5MS column (60 m × 0.25 mm i.d., s
0.25 μm film thickness). The temperature of the oven was
kept at 55 °C for 3 min and then operated at 240 °C and a
rate of 3 °C/min. The corresponding temperatures of the injec-
tor and detector were 260 °C and 270 °C. Helium (1.0 mL/min)
was used as a carrier gas. The injection volume of the oil sample
was 1.0 µL.
The GC-MS analysis has been carried out using the same
HP-5MS column and coupled with a mass HP 5973 MSD spec-
trometer. The temperature of the oven was 55 °C to 240 °C at a
rate of 3 °C/min. He (1 mL/min), split ratio 9:1, ionization
energy of 70 eV, emission current of 40 mA, sampling rate of
1.0 scan/s, and mass scanning of 50 to 450 amu were estab-
lished. An EI source at 240 °C was used, while the interface
temperature was 270 °C. The retention index (RI) of each com-
ponent was calculated by a comparison between the experimen-
tal result and a homologous series of n-alkanes C7-C40. Each
chemical was identified by comparing its RI value to that in lit-
erature.12,13 Based on the NIST 20 and WILEY 12 Libraries,
the MS fragmentations were compared to those of other essen-
tial oils with known compositions, whereas the peak area in the
GC-FID was used to calculate the relative percentage (%)
without using any correction factor.
Cytotoxic and Antimicrobial Assays
Cytotoxic and antimicrobial experiments were deduced from
our previous reports.27–31
Docking Calculation
Docking simulations have been conducted to consider the inter-
actions between the main components of B tonkinensis leaf oil
and the targeted proteins. Auto Dock Vina v1.2.332,33 was
9
employed for the simulations, utilizing a united atom scoring
function and generating a maximum of 20 binding modes.
The global search exhaustiveness was set to 400. The structures
of the target proteins have been collected from the RCSB PDB
database34 and underwent preprocessing steps, including the
deletion of water molecules, the addition of hydrogen atoms
and missed residues, and the Kollman charge, before initiating
the molecular docking process. Gasteiger charges were incorpo-
rated into the ligand molecules, which were then converted to
the PDBQT format. Auto Dock Tools v.1.5.6 was utilized to
confirm a grid box of size 22 × 22 × 22 Å with a grid spacing
of 1 Å. During docking, the receptor molecule was held rigid
while the ligand molecules remained flexible. The best
docking poses were examined to identify conformers with the
lowest binding energy and to analyze 2-dimensional interac-
tions. The docked conformations and 3-dimensional target-
ligand interactions were visualized using BIOVIA Discovery
Studio v2021.
Authors’s note
Ninh The Son is also affiliated with Graduate University of Science and
Technology, VAST, Caugiay, Hanoi, Vietnam.
Declaration of Conflicting Interests
The author(s) declared no potential conflicts of interest with respect to
the research, authorship, and/or publication of this article.
Funding
The author(s) disclosed receipt of the following financial support for
the research, authorship, and/or publication of this article: This
research was funded by Hanoi Pedagogical University 2 foundation
for Sciences and Technology Development via grant number:
HPU2.2023-UT-05.
ORCID iDs
Duong Quang Huan https://orcid.org/0000-0002-7661-5533
Do Ngoc Dai https://orcid.org/0000-0002-7741-9454
Nguyen Xuan Ha https://orcid.org/0000-0002-8779-256X
Ninh The Son https://orcid.org/0000-0002-2413-7496
Supplemental Material
Supplemental material for this article is available online.
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